title: PeerJ description: Articles published in PeerJ link: https://peerj.com/articles/index.rss3?journal=peerj&month=2020-10 creator: info@peerj.com PeerJ errorsTo: info@peerj.com PeerJ language: en title: Integrated network pharmacology and molecular docking approaches to reveal the synergistic mechanism of multiple components in Venenum Bufonis for ameliorating heart failure link: https://peerj.com/articles/10107 last-modified: 2020-10-30 description: Venenum Bufonis (VB), also called Chan Su in China, has been extensively used as a traditional Chinese medicine (TCM) for treating heart failure (HF) since ancient time. However, the active components and the potential anti-HF mechanism of VB remain unclear. In the current study, the major absorbed components and metabolites of VB after oral administration in rats were first collected from literatures. A total of 17 prototypes and 25 metabolites were gathered. Next, a feasible network-based pharmacological approach was developed and employed to explore the therapeutic mechanism of VB on HF based on the collected constituents. In total, 158 main targets were screened out and considered as effective players in ameliorating HF. Then, the VB components–main HF putative targets–main pathways network was established, clarifying the underlying biological process of VB on HF. More importantly, the main hubs were found to be highly enriched in adrenergic signalling in cardio-myocytes. After verified by molecular docking studies, four key targets (ATP1A1, GNAS, MAPK1 and PRKCA) and three potential active leading compounds (bufotalin, cinobufaginol and 19-oxo-bufalin) were identified, which may play critical roles in cardiac muscle contraction. This study demonstrated that the integrated strategy based on network pharmacology and molecular docking was helpful to uncover the synergistic mechanism of multiple constituents in TCM. creator: Wei Ren creator: Zhiqiang Luo creator: Fulu Pan creator: Jiali Liu creator: Qin Sun creator: Gang Luo creator: Raoqiong Wang creator: Haiyu Zhao creator: Baolin Bian creator: Xiao Xiao creator: Qingrong Pu creator: Sijin Yang creator: Guohua Yu uri: https://doi.org/10.7717/peerj.10107 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Ren et al. title: Biases in genome reconstruction from metagenomic data link: https://peerj.com/articles/10119 last-modified: 2020-10-30 description: BackgroundAdvances in sequencing, assembly, and assortment of contigs into species-specific bins has enabled the reconstruction of genomes from metagenomic data (MAGs). Though a powerful technique, it is difficult to determine whether assembly and binning techniques are accurate when applied to environmental metagenomes due to a lack of complete reference genome sequences against which to check the resulting MAGs.MethodsWe compared MAGs derived from an enrichment culture containing ~20 organisms to complete genome sequences of 10 organisms isolated from the enrichment culture. Factors commonly considered in binning software—nucleotide composition and sequence repetitiveness—were calculated for both the correctly binned and not-binned regions. This direct comparison revealed biases in sequence characteristics and gene content in the not-binned regions. Additionally, the composition of three public data sets representing MAGs reconstructed from the Tara Oceans metagenomic data was compared to a set of representative genomes available through NCBI RefSeq to verify that the biases identified were observable in more complex data sets and using three contemporary binning software packages.ResultsRepeat sequences were frequently not binned in the genome reconstruction processes, as were sequence regions with variant nucleotide composition. Genes encoded on the not-binned regions were strongly biased towards ribosomal RNAs, transfer RNAs, mobile element functions and genes of unknown function. Our results support genome reconstruction as a robust process and suggest that reconstructions determined to be >90% complete are likely to effectively represent organismal function; however, population-level genotypic heterogeneity in natural populations, such as uneven distribution of plasmids, can lead to incorrect inferences. creator: William C. Nelson creator: Benjamin J. Tully creator: Jennifer M. Mobberley uri: https://doi.org/10.7717/peerj.10119 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Nelson et al. title: Revalidation of morphological characteristics and multiplex PCR for the identification of three congener invasive Liriomyza species (Diptera: Agromyzidae) in China link: https://peerj.com/articles/10138 last-modified: 2020-10-30 description: Due to varietal differences, diminutive size, and similar morphological characters, it is difficult to classify and identify Liriomyza spp., a genus comprised of economically-important, highly-polyphagous insect pests. In this study, we reconfirmed the morphological characteristics of three closely-related invasive leafminers, L. trifolii, L. sativae, and L. huidobrensis. Morphological results showed that characteristics imparted by the male genitalia were the most reliable morphological features for identification. The colors exhibited by vertical setae were variable among species, and the ratio of the length of the ultimate section of vein CuA1 divided by penultimate section also varied within species. Although the patterns of abdominal tergites were diverse among Liriomyza spp., L. trifolii exhibited a unique pattern with a yellow patch at the 5th black visible tergite; this pattern can be profiled as a prominent characteristic for morphological identification. In order to identify the three Liriomyza spp. quickly and accurately, we developed an improved molecular identification method using multiplex PCR based on the gene encoding mitochondrial cytochrome oxidase I (COI); this method enabled direct identification based on the size of amplified products. The results of this study provide a valuable reference for the identification of Liriomyza spp., which will ultimately improve our ability to control individual species. creator: Ya-Wen Chang creator: Jing-Yun Chen creator: Si-Zhu Zheng creator: Yuan Gao creator: Yunfang Chen creator: Yanfeng Deng creator: Yu-Zhou Du uri: https://doi.org/10.7717/peerj.10138 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chang et al. title: Social cognition and social functioning in people with borderline personality disorder and their first-degree relatives link: https://peerj.com/articles/10212 last-modified: 2020-10-30 description: BackgroundA few papers studying healthy, first-degree relatives of people with borderline personality disorder (BPD) have found that this group presents attention and memory problems. However, current research has not analyzed their social cognition.Materials and MethodsWe designed an age-, gender- and education-level matched case-control study involving 57 people with BPD, 32 of their first-degree relatives, and 57 healthy controls in Spain in 2018–2019. All were assessed for social cognition and functioning using the Movie for Assessment of Social Cognition and the Social Functioning Scale; other potential confounders were also collected (marital status, occupation and household variables).ResultsThere were differences in the social cognition domain of overmentalizing errors, with the BPD group scoring significantly higher than controls; however, there was no significant difference with relatives; in the social functioning domain of family relationships, with the controls showing the highest scores. Social engagement/withdrawal, interpersonal behavior, independence-competence, prosocial activities, full scale and categorization domains showed the same pattern: the BPD group had lower scores than their relatives and the controls. Relatives were significantly different from BPD patients in family relationships, social engagement/withdrawal and interpersonal behavior, as well as on the full Social Functioning Scale (both as a linear and categorical variable). However, only controls showed differences with relatives in family relationships.ConclusionsAll in all, relatives show similar levels of social cognition and functioning compared with controls, and people with BPD show some alterations in different domains of both social cognition and functioning. creator: Esther Ortega-Díaz creator: Jonatan García-Campos creator: José María Rico-Gomis creator: Carlos Cuesta-Moreno creator: Antonio Palazón-Bru creator: Gabriel Estañ-Cerezo creator: José Antonio Piqueras-Rodríguez creator: Jesús Rodríguez-Marín uri: https://doi.org/10.7717/peerj.10212 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Ortega-Díaz et al. title: Genetic variations and dog breed identification using inter-simple sequence repeat markers coupled with high resolution melting analysis link: https://peerj.com/articles/10215 last-modified: 2020-10-30 description: The identification of differing physical characteristics of dogs is an uncomplicated and straightforward way to categorize dog breeds. However, many dog owners and veterinarians still struggle to distinguish between pure breed and mixed variations in certain breeds of dogs. Presently, the absence of the tools and methods needed to confirm a pure breed dog is a significant problem since the only method available to validate pure or mongrel breeds is the official pedigree system. Inter-simple sequence repeat markers have been successfully used to assess genetic variations and differentiations. Notably, inter-simple sequence repeat markers coupled with high resolution melting analysis were effectively used for the breed identification of 43 breeds of dogs (total 463 dogs). The 10 primers chosen for analysis resulted in a range of 31–78.6% of breed discrimination when using one primer, while a combination of two primers was able to successfully discriminate between all of the 43 dog breeds (100%). Shannon’s index information (I = 2.586 ± 0.034) and expected heterozygosity (He = 0.908 ± 0.003) indicated a high level of genetic diversity among breeds. The fixation index (Fst) revealed a value of 10.4%, demonstrating that there was a high level of genetic subdivision between populations. This study showed that inter-simple sequence repeat marker analysis was effective in demonstrating high genetic diversity among varying breeds of dogs, while a combination of Inter-simple sequence repeat marker analysis and high resolution melting analysis could provide an optional technique for researchers to effectively identify breeds through genetic variations. creator: Wannapimol Kriangwanich creator: Korakot Nganvongpanit creator: Kittisak Buddhachat creator: Puntita Siengdee creator: Siriwadee Chomdej creator: Siriluck Ponsuksili creator: Chatchote Thitaram uri: https://doi.org/10.7717/peerj.10215 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kriangwanich et al. title: Construction of fibronectin conditional gene knock-out mice and the effect of fibronectin gene knockout on hematopoietic, biochemical and immune parameters in mice link: https://peerj.com/articles/10224 last-modified: 2020-10-30 description: Fibronectin (FN) is a multi-functional glycoprotein that primarily acts as a cell adhesion molecule and tethers cells to the extra cellular matrix. In order to clarify the effect of FN deficiency on hematopoiesis, biochemical and immune parameters in mice. We constructed a tamoxifen-induced conditional (cre-loxp system) fibronectin knock-out (FnKO) mouse model on a C57BL/6 background, and monitored their behavior, fertility, histological, hematopoietic, biochemical and immunological indices. We found that the Fn KO mice had reduced fertility, high platelet counts, smaller bone marrow megakaryocytes and looser attachment between the hepatocyte and vascular endothelial junctions compared to the wild type (WT) mice. In contrast, the behavior, hematological counts, serum biochemical indices and vital organ histology were similar in both Fn KO and WT mice. This model will greatly help in elucidating the role of FN in immune-related diseases in future. creator: Xiaohong Yuan creator: Shu Yang creator: Wen Li creator: Jinggang Li creator: Jia Lin creator: Yong Wu creator: Yuanzhong Chen uri: https://doi.org/10.7717/peerj.10224 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yuan et al. title: Cytological and molecular characterizations of a novel 2A nullisomic line derived from a widely-grown wheat cultivar Zhoumai 18 conferring male sterility link: https://peerj.com/articles/10275 last-modified: 2020-10-30 description: A dwarf, multi-pistil and male sterile dms mutant was previously reported by us. However, the genetic changes in this dms are unclear. To examine the genetic changes, single nucleotide polymorphism (SNP) association, chromosome counting, and high-resolution chromosome fluorescence in situ hybridization (FISH) techniques were employed. By comparing tall plants (T) with dwarf plants (D) in the offspring of dms mutant plants, SNP association analysis indicated that most SNPs were on chromosome 2A. There were three types in offspring of dms plants, with 42, 41 and 40 chromosomes respectively. High-resolution chromosome painting analysis demonstrated that T plants had all 42 wheat chromosomes; the medium plants (M) had 41 chromosomes, lacking one chromosome 2A; while D plants had 40 wheat chromosomes, and lacked both 2A chromosomes. These data demonstrated that dms resulted from a loss of chromosome 2A. We identified 23 genes on chromosome 2A which might be involved in the development of stamens or pollen grains. These results lay a solid foundation for further analysis of the molecular mechanisms of wheat male sterility. Because D plants can be used as a female parent to cross with other wheat genotypes, dms is a unique germplasm for any functional study of chromosome 2A and wheat breeding specifically targeting genes on 2A. creator: Zhixin Jiao creator: Xinxin Zhu creator: Huijuan Li creator: Zhitao Liu creator: Xinyi Huang creator: Nan Wu creator: Junhang An creator: Junchang Li creator: Jing Zhang creator: Yumei Jiang creator: Qiaoyun Li creator: Zengjun Qi creator: Jishan Niu uri: https://doi.org/10.7717/peerj.10275 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Jiao et al. title: On local scouring downstream small water structures link: https://peerj.com/articles/10282 last-modified: 2020-10-30 description: BackgroundIn order to regulate water flow, hydraulic structures such as weirs or checks, frequently equipped with gates, are used. Water can flow below or over the gate or, simultaneously, over and below the gate. Both diversifications of hydraulic gradient, being an effect of damming up a river by the structure and shear stresses at the bed, which exceeds the critical shear stress value, invoke the local scouring downstream the structure. This phenomenon has been studied in laboratory and field conditions for many years, however Researchers do not agree on the parameters that affect the size of the local scour and the intensity of its formation. There are no universal methods for estimating its magnitude However, solutions are sought in the form of calculation formulas typical for the method of flow through the structure, taking into account the parameters that characterize a given structure. These formulas are based on factors that affect the size of the local scours, that is, their dimensions and location. Examples of such formulas are those contained in this article: Franke (1960), Straube (1963), Tarajmovič (1966), Rossinski & Kuzmin (1969) equations. The need to study this phenomenon results from the prevalence of hydrotechnical structures equipped with gates (from small gated checks to large weirs) and from potential damage that may be associated with excessive development of local erosion downstream, including washing of foundations and, consequently, loss of stability of the structure.MethodsThis study verifies empirical formulas applied to estimate the geometry parameters of a scour hole on a laboratory model of a structure where water is conducted downstream the gate with bottom reinforcements of various roughness. A specially designed remote-controlled measuring device, equipped with laser scanner, was applied to determine the shape of the sandy bottom. Then the formula optimization is conducted, using Monte Carlo sampling method, followed by verification of field conditions.ResultsThe suitability of a specially designed device, equipped with laser scanner for measuring the bottom shape in laboratory conditions was demonstrated. Simple formula describing local scour geometry in laboratory conditions was derived basing on the Straube formula. The optimized formula was verified in field conditions giving very good comparative results. Therefore, it can be applied in engineering and designing practices. creator: Marta Kiraga creator: Zbigniew Popek uri: https://doi.org/10.7717/peerj.10282 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kiraga and Popek title: A review of Euryoryzomys legatus (Rodentia, Sigmodontinae): morphological redescription, cytogenetics, and molecular phylogeny link: https://peerj.com/articles/9884 last-modified: 2020-10-29 description: The taxonomic history of Euryoryzomys legatus has been complex and controversial, being either included in the synonymy of other oryzomyine species or considered as a valid species, as in the most recent review of the genus. Previous phylogenetic analyses segregated E. legatus from E. russatus, its putative senior synonym, but recovered it nested within E. nitidus. A general lack of authoritative evaluation of morphological attributes, details of the chromosome complement, or other data types has hampered the ability to choose among alternative taxonomic hypotheses, and thus reach a general consensus for the status of the taxon. Herein we revisit the status of E. legatus using an integrated approach that includes: (1) a morphological review, especially centered on specimens from northwestern Argentina not examined previously, (2) comparative cytogenetics, and (3) phylogenetic reconstruction, using mitochondrial genes. Euryoryzomys legatus is morphologically and phylogenetically distinct from all other species-level taxa in the genus, but its 2n=80, FN=86 karyotype is shared with E. emmonsae, E. nitidus, and E. russatus. Several morphological and morphometric characters distinguish E. legatus from other species of Euryoryzomys, and we provide an amended diagnosis for the species. Morphological characters useful in distinguishing E. legatus from E. nitidus, its sister taxon following molecular analyses, include: larger overall size, dorsal fur with a strong yellowish brown to orange brown tinge, flanks and cheeks with an orange lateral line, ventral color grayish-white with pure white hairs present only on the chin, presence of a thin blackish eye-ring, tail bicolored, presence of an alisphenoid strut and a well-developed temporal and lambdoid crests in the skull, and a labial cingulum on M3. Molecular phylogenetic analyses recovered E. legatus as a monophyletic group with high support nested within a paraphyletic E. nitidus; genetic distances segregated members of both species, except for an exemplar of E. nitidus. Our integrated analyses reinforce E. legatus as a full species, but highlight that E. macconnelli, E. emmonsae, and E. nitidus each may be a species complex and worthy of systematic attention. Finally, we also evaluated the chromosome evolution of the genus within a phylogenetic context. creator: Mariana D. Guilardi creator: Pablo Jayat creator: Marcelo Weksler creator: James L. Patton creator: Pablo Edmundo Ortiz creator: Keila Almeida creator: Maria José de J. Silva uri: https://doi.org/10.7717/peerj.9884 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Guilardi et al. title: A 14 immune-related gene signature predicts clinical outcomes of kidney renal clear cell carcinoma link: https://peerj.com/articles/10183 last-modified: 2020-10-29 description: Kidney renal clear cell carcinoma (KIRC) is the leading cause of kidney cancer-related deaths. Currently, there are no studies in tumor immunology investigating the use of signatures as a predictor of overall survival in KIRC patients. Our study attempts to establish an immune-related gene risk signature to predict clinical outcomes in KIRC. A total of 528 patients from The Cancer Genome Atlas (TCGA) database were included in our analysis and randomly divided into training (n = 315) and testing sets (n = 213). We collected 1,534 immune-related genes from the Immunology Database and Analysis Portal as candidates to construct our signature. LASSO-COX was used to find gene models with the highest predictive ability. We used survival and Cox analysis to test the model’s independent prognostic ability. Univariate analysis identified 650 immune-related genes with prognostic abilities. After 1,000 iterations, we choose 14 of the most frequent and stable immune-related genes as our signature. We found that the signature was associated with M stage, T stage, and pathological staging. More importantly, the signature can independently predict clinical prognosis in KIRC patients. Gene Set Enrichment Analysis (GSEA) showed an association between our signature and critical metabolism pathways. Our research established a model based upon 14 immune-related genes that predicted the prognosis of KIRC patients based on tumor immune microenvironments. creator: Yong Zou creator: Chuan Hu uri: https://doi.org/10.7717/peerj.10183 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zou and Hu title: Comparative analysis of the metabolically active microbial communities in the rumen of dromedary camels under different feeding systems using total rRNA sequencing link: https://peerj.com/articles/10184 last-modified: 2020-10-29 description: Breakdown of plant biomass in rumen depends on interactions between bacteria, archaea, fungi, and protozoa; however, the majority of studies of the microbiome of ruminants, including the few studies of the rumen of camels, only studied one of these microbial groups. In this study, we applied total rRNA sequencing to identify active microbial communities in 22 solid and liquid rumen samples from 11 camels. These camels were reared at three stations that use different feeding systems: clover, hay and wheat straw (G1), fresh clover (G2), and wheat straw (G3). Bacteria dominated the libraries of sequence reads generated from all rumen samples, followed by protozoa, archaea, and fungi respectively. Firmicutes, Thermoplasmatales, Diplodinium, and Neocallimastix dominated bacterial, archaeal, protozoal and fungal communities, respectively in all samples. Libraries generated from camels reared at facility G2, where they were fed fresh clover, showed the highest alpha diversity. Principal co-ordinate analysis and linear discriminate analysis showed clusters associated with facility/feed and the relative abundance of microbes varied between liquid and solid fractions. This provides preliminary evidence that bacteria dominate the microbial communities of the camel rumen and these communities differ significantly between populations of domesticated camels. creator: Alaa Emara Rabee creator: Robert Forster creator: Chijioke Elekwachi creator: Ebrahim Sabra creator: Mebarek Lamara uri: https://doi.org/10.7717/peerj.10184 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Rabee et al. title: Polylactic acid as a suitable material for 3D printing of protective masks in times of COVID-19 pandemic link: https://peerj.com/articles/10259 last-modified: 2020-10-29 description: A critical lack of personal protective equipment has occurred during the COVID-19 pandemic. Polylactic acid (PLA), a polyester made from renewable natural resources, can be exploited for 3D printing of protective face masks using the Fused Deposition Modelling technique. Since the possible high porosity of this material raised questions regarding its suitability for protection against viruses, we have investigated its microstructure using scanning electron microscopy and aerosol generator and photometer certified as the test system according to the standards EN 143 and EN 149. Moreover, the efficiency of decontaminating PLA surfaces by conventional chemical disinfectants including 96% ethanol, 70% isopropanol, and a commercial disinfectant containing 0.85% sodium hypochlorite has been determined. We confirmed that the structure of PLA protective masks is compact and can be considered a sufficient barrier protection against particles of a size corresponding to microorganisms including viruses. Complete decontamination of PLA surfaces from externally applied Staphylococcus epidermidis, Escherichia coli, Candida albicans and SARS-CoV-2 was achieved using all disinfectants tested, and human adenovirus was completely inactivated by sodium hypochlorite-containing disinfectant. Natural contamination of PLA masks worn by test persons was decontaminated easily and efficiently by ethanol. No disinfectant caused major changes to the PLA surface properties, and the pore size did not change despite severe mechanical damage of the surface. Therefore, PLA may be regarded as a suitable material for 3D printing of protective masks during the current or future pandemic crises. creator: Eva Vaňková creator: Petra Kašparová creator: Josef Khun creator: Anna Machková creator: Jaroslav Julák creator: Michal Sláma creator: Jan Hodek creator: Lucie Ulrychová creator: Jan Weber creator: Klára Obrová creator: Karin Kosulin creator: Thomas Lion creator: Vladimír Scholtz uri: https://doi.org/10.7717/peerj.10259 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Vaňková et al. title: The invasive red-eared slider turtle is more successful than the native Chinese three-keeled pond turtle: evidence from the gut microbiota link: https://peerj.com/articles/10271 last-modified: 2020-10-29 description: BackgroundThe mutualistic symbiosis between the gut microbial communities (microbiota) and their host animals has attracted much attention. Many factors potentially affect the gut microbiota, which also varies among host animals. The native Chinese three-keeled pond turtle (Chinemys reevesii) and the invasive red-eared slider turtle (Trachemys scripta elegans) are two common farm-raised species in China, with the latter generally considered a more successful species. However, supporting evidence from the gut microbiota has yet to be collected.MethodsWe collected feces samples from these two turtle species raised in a farm under identical conditions, and analyzed the composition and relative abundance of the gut microbes using bacterial 16S rRNA sequencing on the Roach/454 platform.ResultsThe gut microbiota was mainly composed of Bacteroidetes and Firmicutes at the phylum level, and Porphyromonadaceae, Bacteroidaceae and Lachnospiraceae at the family level in both species. The relative abundance of the microbes and gene functions in the gut microbiota differed between the two species, whereas alpha or beta diversity did not. Microbes of the families Bacteroidaceae, Clostridiaceae and Lachnospiraceae were comparatively more abundant in C. reevesii, whereas those of the families Porphyromonadaceae and Fusobacteriaceae were comparatively more abundant in T. s. elegans. In both species the gut microbiota had functional roles in enhancing metabolism, genetic information processing and environmental information processing according to the Kyoto Encyclopedia of Genes and Genomes database. The potential to gain mass is greater in T. s. elegans than in C. reevesii, as revealed by the fact that the Firmicutes/Bacteroidetes ratio was lower in the former species. The percentage of human disease-related functional genes was lower in T. s. elegans than in C. reevesii, presumably suggesting an enhanced potential to colonize new habitats in the former species. creator: Yan-Fu Qu creator: Yan-Qing Wu creator: Yu-Tian Zhao creator: Long-Hui Lin creator: Yu Du creator: Peng Li creator: Hong Li creator: Xiang Ji uri: https://doi.org/10.7717/peerj.10271 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Qu et al. title: Genetic variability in Ruditapes decussatus clam combined with Perkinsus infection level to support founder population selection for a breeding program link: https://peerj.com/articles/9728 last-modified: 2020-10-28 description: Clam farmers worldwide face several challenges, including irregular seed supply and high mortalities due to pathogenic organisms such as Perkinsus olseni. In Europe, there is a high unmet consumer demand for native clam species such as Ruditapes decussatus. The high market value of R. decussatus makes the culture of this species potentially more attractive than that culture of the alien species Ruditapes philippinarum. Thus, there is a market opportunity in breeding and producing R. decussatus at an industrial scale. A selective breeding program to improve R. decussatus performance will be carried out in Portugal; and the first critical step to develop such a breeding program is the establishment of a founder population. In this study, intra- and interpopulation genetic diversity was assessed using 13 microsatellite markers in eight natural beds located in Portugal, Spain and Italy. Also, allele and genotypic frequencies of each microsatellite locus were assessed discriminating between clams infected and non-infected by P. olseni. All locations showed similar values for several genetic diversity parameters. Analyses of population differentiation (FST, Bayesian clustering and AMOVAs) revealed five genetically differentiated regions: Rías Altas and Rías Baixas (NW Spain), North/Central Coast of Portugal, Gulf of Cadiz and Adriatic Sea. Significant differences in the allelic and genotypic frequency distribution between infected clams and non-infected ones at four microsatellite loci are reported suggesting that resistance to the disease could have a genetic basis. Moreover, a positive or negative relationship between the frequency of certain alleles and the parasite infection was inferred. Further studies should confirm the potential use of those alleles as genetic markers for P. olseni infection. Integrating results of genetic diversity within and between populations and Perkinsus infection levels, a founder population for a R. decussatus breeding program is proposed, composed by individuals from Barallobre (Rías Altas), Pontevedra or Cangas (Rías Baixas), Óbidos (North/Central Coast of Portugal), Algarve (Gulf of Cadiz) and Venice (Adriatic Sea). creator: Andreia Cruz creator: Fiz da Costa creator: Jenyfer Fernández-Pérez creator: Ana Nantón creator: Sergio Fernández-Boo creator: Ana Insua creator: Josefina Méndez uri: https://doi.org/10.7717/peerj.9728 license: https://creativecommons.org/licenses/by-nc/4.0 rights: © 2020 Cruz et al. title: A shift away from mutualism under food-deprived conditions in an anemone-dinoflagellate association link: https://peerj.com/articles/9745 last-modified: 2020-10-28 description: The mutualistic symbiosis between anthozoans and intra-gastrodermal dinoflagellates of the family Symbiodiniaceae is the functional basis of all coral reef ecosystems, with the latter providing up to 95% of their fixed photosynthate to their hosts in exchange for nutrients. However, recent studies of sponges, jellyfish, and anemones have revealed the potential for this mutualistic relationship to shift to parasitism under stressful conditions. Over a period of eight weeks, we compared the physiological conditions of both inoculated and aposymbiotic anemones (Exaiptasia pallida) that were either fed or starved. By the sixth week, both fed groups of anemones were significantly larger than their starved counterparts. Moreover, inoculated and starved anemones tended to disintegrate into “tissue balls” within eight weeks, and 25% of the samples died; in contrast, starved aposymbiotic anemones required six months to form tissue balls, and no anemones from this group died. Our results show that the dinoflagellates within inoculated anemones may have posed a fatal metabolic burden on their hosts during starvation; this may be because of the need to prioritize their own metabolism and nourishment at the expense of their hosts. Collectively, our study reveals the potential of this dynamic symbiotic association to shift away from mutualism during food-deprived conditions. creator: Shao-En Peng creator: Alessandro Moret creator: Cherilyn Chang creator: Anderson B. Mayfield creator: Yu-Ting Ren creator: Wan-Nan U. Chen creator: Mario Giordano creator: Chii-Shiarng Chen uri: https://doi.org/10.7717/peerj.9745 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Peng et al. title: TVA-based modeling of short-term memory capacity, speed of processing and perceptual threshold in chronic stroke patients undergoing cognitive training: case-control differences, reliability, and associations with cognitive performance link: https://peerj.com/articles/9948 last-modified: 2020-10-28 description: Attentional deficits following stroke are common and pervasive, and are important predictors for functional recovery. Attentional functions comprise a set of specific cognitive processes allowing to attend, filter and select among a continuous stream of stimuli. These mechanisms are fundamental for more complex cognitive functions such as learning, planning and cognitive control, all crucial for daily functioning. The distributed functional neuroanatomy of these processes is a likely explanation for the high prevalence of attentional impairments following stroke, and underscores the importance of a clinical implementation of computational approaches allowing for sensitive and specific modeling of attentional sub-processes. The Theory of Visual Attention (TVA) offers a theoretical, computational, neuronal and practical framework to assess the efficiency of visual selection performance and parallel processing of multiple objects. Here, in order to assess the sensitivity and reliability of TVA parameters reflecting short-term memory capacity (K), processing speed (C) and perceptual threshold (t0), we used a whole-report paradigm in a cross-sectional case-control comparison and across six repeated assessments over the course of a three-week computerized cognitive training (CCT) intervention in chronic stroke patients (> 6 months since hospital admission, NIHSS ≤ 7 at hospital discharge). Cross-sectional group comparisons documented lower short-term memory capacity, lower processing speed and higher perceptual threshold in patients (n = 70) compared to age-matched healthy controls (n = 140). Further, longitudinal analyses in stroke patients during the course of CCT (n = 54) revealed high reliability of the TVA parameters, and higher processing speed at baseline was associated with larger cognitive improvement after the intervention. The results support the feasibility, reliability and sensitivity of TVA-based assessment of attentional functions in chronic stroke patients. creator: Geneviève Richard creator: Anders Petersen creator: Kristine Moe Ulrichsen creator: Knut K. Kolskår creator: Dag Alnæs creator: Anne-Marthe Sanders creator: Erlend S. Dørum creator: Hege Ihle-Hansen creator: Jan E. Nordvik creator: Lars T. Westlye uri: https://doi.org/10.7717/peerj.9948 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Richard et al. title: Unsupervised inference approach to facial attractiveness link: https://peerj.com/articles/10210 last-modified: 2020-10-28 description: The perception of facial attractiveness is a complex phenomenon which depends on how the observer perceives not only individual facial features, but also their mutual influence and interplay. In the machine learning community, this problem is typically tackled as a problem of regression of the subject-averaged rating assigned to natural faces. However, it has been conjectured that this approach does not capture the complexity of the phenomenon. It has recently been shown that different human subjects can navigate the face-space and “sculpt” their preferred modification of a reference facial portrait. Here we present an unsupervised inference study of the set of sculpted facial vectors in such experiments. We first infer minimal, interpretable and accurate probabilistic models (through Maximum Entropy and artificial neural networks) of the preferred facial variations, that encode the inter-subject variance. The application of such generative models to the supervised classification of the gender of the subject that sculpted the face reveals that it may be predicted with astonishingly high accuracy. We observe that the classification accuracy improves by increasing the order of the non-linear effective interaction. This suggests that the cognitive mechanisms related to facial discrimination in the brain do not involve the positions of single facial landmarks only, but mainly the mutual influence of couples, and even triplets and quadruplets of landmarks. Furthermore, the high prediction accuracy of the subjects’ gender suggests that much relevant information regarding the subjects may influence (and be elicited from) their facial preference criteria, in agreement with the multiple motive theory of attractiveness proposed in previous works. creator: Miguel Ibanez-Berganza creator: Ambra Amico creator: Gian Luca Lancia creator: Federico Maggiore creator: Bernardo Monechi creator: Vittorio Loreto uri: https://doi.org/10.7717/peerj.10210 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Ibanez-Berganza et al. title: Long-term participation in collaborative fisheries research improves angler opinions on marine protected areas link: https://peerj.com/articles/10146 last-modified: 2020-10-28 description: Recent marine spatial planning efforts, including the management and monitoring of marine protected areas (MPAs), increasingly focus on the importance of stakeholder engagement. For nearly 15 years, the California Collaborative Fisheries Research Program (CCFRP) has partnered volunteer anglers with researchers, the fishing industry, and resource managers to monitor groundfishes in California’s network of MPAs. While the program has succeeded in generating sustained biological observations, we know little about volunteer angler demography or the impact of participation on their perceptions and opinions on fisheries data or MPAs. In this study we surveyed CCFRP volunteers to learn about (a) volunteer angler demographics and attitudes toward groundfish management and stock health, (b) volunteer angler motivations for joining and staying in the program, and (c) whether participation in the program influenced volunteer angler opinions on the quality of fisheries data used in resource management and the establishment of MPAs in California. CCFRP volunteers were older and had higher fishing avidity than average within the California recreational angling community. Many self-identified as more conservation-minded than their peers in the recreational fishing community and had positive views of California groundfish management and stock health. Participation in science and giving back to fisheries resources were major motivating factors in their decision to become and remain CCFRP volunteers. Angler opinions toward MPAs were more positive after volunteering with CCFRP. Those who had volunteered for seven or more years with CCFRP were more likely than not to gain a positive opinion of MPAs. Our survey results provide evidence that long-term engagement of stakeholders in collaborative research positively influences stakeholder opinions regarding marine resource management, and highlights CCFRP’s success in engaging citizen science stakeholders in collaborative fisheries research. creator: Erica T. Mason creator: Allison N. Kellum creator: Jennifer A. Chiu creator: Grant T. Waltz creator: Samantha Murray creator: Dean E. Wendt creator: Richard M. Starr creator: Brice X. Semmens uri: https://doi.org/10.7717/peerj.10146 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Mason et al. title: Transcriptome-wide identification of MAPKKK genes in bermudagrass (Cynodon dactylon L.) and their potential roles in low temperature stress responses link: https://peerj.com/articles/10159 last-modified: 2020-10-28 description: As upstream components of MAPK cascades, mitogen-activated protein kinase kinase kinases (MAPKKKs) act as adaptors linking upstream signaling steps to the core MAPK cascades. MAPK cascades are universal modules of signal transduction in eukaryotic organisms and play crucial roles in plant development processes and in responses to biotic and abiotic stress and signal transduction. Members of the MAPKKK gene family have been identified in several plants,however, MAPKKKs have not been systematically studied in bermudagrass (Cynodon dactylon L.). In this study, 55 potential CdMAPKKKs were produced from bermudagrass transcriptome data, of which 13 belonged to the MEKK, 38 to the Raf, and 4 to the ZIK subfamily. Multiple alignment and conserved motif analysis of CdMAPKKKs supported the evolutionary relationships inferred from phylogenetic analyses. Moreover, the distribution pattern in Poaceae species indicated that members of the MAPKKK family were conserved among almost all diploid species, and species-specific polyploidy or higher duplication ratios resulted in an expansion of the MAPKKK family. In addition, 714 co-functional links which were significantly enriched in signal transduction, responses to temperature stimuli, and other important biological processes of 55 CdMAPKKKs were identified using co-functional gene networks analysis; 30 and 19 co-functional genes involved in response to cold or heat stress, respectively, were also identified. Results of promoter analyses, and interaction network investigation of all CdMAPKKKs based on the rice homologs suggested that CdMAPKKKs are commonly associated with regulation of numerous biological processes. Furthermore, 12 and 13 CdMAPKKKs were significantly up- and downregulated, respectively, in response to low temperature stress; among them, six CdMAPKKKs were significantly induced by low temperature stress, at least at one point in time. This is the first study to conduct identification and functional analysis of the MAPKKK gene family in bermudagrass, and our results provide a foundation for further research on the functions of CdMAPKKKs in response to low temperature stress. creator: Wei Wang creator: An Shao creator: Erick Amombo creator: Shugao Fan creator: Xiao Xu creator: Jinmin Fu uri: https://doi.org/10.7717/peerj.10159 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: Assessment of nationally representative dietary studies in the Gulf Cooperation Council: a scoping review link: https://peerj.com/articles/10163 last-modified: 2020-10-28 description: BackgroundObesity is at a record high in Gulf Cooperation Council (GCC) countries and is expected to continue increasing. Diet is a major contributor to this disease, but there is inadequate nationally representative dietary research from these countries. The aim was to quantify the number dietary studies using food frequency questionnaires (FFQs) that have been conducted in individual GCC countries and to assess the quality of eligible studies.MethodologyFour databases (PubMed, Web of Science, MEDLINE, and DOAJ) were searched for keywords; records were screened for eligible studies and data were abstracted on study characteristics (publication year, geographical locations, sample size, units of measurement, number of foods examined, number of Arab foods and key findings). Quality was assessed using an adapted Newcastle-Ottawa Quality Assessment Scale for cross-sectional studies.ResultsOnly seven studies were eligible from four of six GCC countries (Saudi Arabia, Bahrain, Kuwait and Qatar). All eligible studies used FFQs, but only 29% used a validated questionnaire, one being in Arabic, and none of the studies used any additional tools to measure diet. Fifty-seven percent of studies made an effort to include local foods. The majority of studies (71%) either measured frequency or quantity of food consumed, but only 29% attempted to account for both frequency and quantity.ConclusionsThe quality of studies varied and major weaknesses of FFQ validity and adaptability have been highlighted. More dietary investigations are needed using validated FFQs that have been adapted to the local GCC diets. Using reference tools will allow for better dietary estimations. creator: Rukshana Hoque creator: Erin Strotheide creator: Juliann Saquib creator: Nazmus Saquib uri: https://doi.org/10.7717/peerj.10163 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Hoque et al. title: Random forest algorithms for recognizing daily life activities using plantar pressure information: a smart-shoe study link: https://peerj.com/articles/10170 last-modified: 2020-10-28 description: BackgroundWearable activity trackers are regarded as a new opportunity to deliver health promotion interventions. Indeed, while the prediction of active behaviors is currently primarily relying on the processing of accelerometer sensor data, the emergence of smart clothes with multi-sensing capacities is offering new possibilities. Algorithms able to process data from a variety of smart devices and classify daily life activities could therefore be of particular importance to achieve a more accurate evaluation of physical behaviors. This study aims to (1) develop an activity recognition algorithm based on the processing of plantar pressure information provided by a smart-shoe prototype and (2) to determine the optimal hardware and software configurations.MethodSeventeen subjects wore a pair of smart-shoe prototypes composed of plantar pressure measurement insoles, and they performed the following nine activities: sitting, standing, walking on a flat surface, walking upstairs, walking downstairs, walking up a slope, running, cycling, and completing office work. The insole featured seven pressure sensors. For each activity, at least four minutes of plantar pressure data were collected. The plantar pressure data were cut in overlapping windows of different lengths and 167 features were extracted for each window. Data were split into training and test samples using a subject-wise assignment method. A random forest model was trained to recognize activity. The resulting activity recognition algorithms were evaluated on the test sample. A multi hold-out procedure allowed repeating the operation with 5 different assignments. The analytic conditions were modulated to test (1) different window lengths (1–60 seconds), (2) some selected sensor configurations and (3) different numbers of data features.ResultsA window length of 20 s was found to be optimum and therefore used for the rest of the analysis. Using all the sensors and all 167 features, the smart shoes predicted the activities with an average success of 89%. “Running” demonstrated the highest sensitivity (100%). “Walking up a slope” was linked with the lowest performance (63%), with the majority of the false negatives being “walking on a flat surface” and “walking upstairs.” Some 2- and 3-sensor configurations were linked with an average success rate of 87%. Reducing the number of features down to 20 does not alter significantly the performance of the algorithm.ConclusionHigh-performance human behavior recognition using plantar pressure data only is possible. In the future, smart-shoe devices could contribute to the evaluation of daily physical activities. Minimalist configurations integrating only a small number of sensors and computing a reduced number of selected features could maintain a satisfying performance. Future experiments must include a more heterogeneous population. creator: Dian Ren creator: Nathanael Aubert-Kato creator: Emi Anzai creator: Yuji Ohta creator: Julien Tripette uri: https://doi.org/10.7717/peerj.10170 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Ren et al. title: Systematic revision of the genus Isodacrys Sharp, 1911 (Coleoptera: Curculionidae: Entiminae: Tanymecini) link: https://peerj.com/articles/10191 last-modified: 2020-10-28 description: The genus Isodacrys Sharp, 1911 is revised. Twenty species of the genus are recognized ranging from south United States of America, Mexico, Guatemala and Honduras of which eight are herein described as new. These species are Isodacrys antrum Cortés-Hernández, new species (Mexico: Tamaulipas, Chiapas; Guatemala: Baja Verapaz); Isodacrys carlae Cortés-Hernández, new species (Mexico: Coahuila, Hidalgo, Nuevo León, San Luis Potosí, Tamaulipas); Isodacrys confusum Cortés-Hernández, new species (Mexico: Tamaulipas); Isodacrys fasciatum Cortés-Hernández, new species (Mexico: Coahuila, Durango, Nuevo León); Isodacrys frontalis Cortés-Hernández, new species (Mexico: Oaxaca; Guatemala: Sacatepéquez, Guatemala); Isodacrys kuchii Cortés-Hernández, new species (Mexico: Puebla); Isodacrys obrienorum Cortés-Hernández, new species (Guatemala: Totonicapán, Jalapa, San Marcos); and Isodacrys okuiltontli Cortés Hernández, new species (Mexico: Oaxaca). Insights into the monophyly of Isodacrys and its phylogenetic relationships with other Tanymecini based on adult morphology are given by implementing a phylogenetic analysis of 43 terminals (21 ingroup, 22 outgroup) coded for 72 adult morphological characters. Characters were discussed and highlighted for the inclusion in the phylogenetic analysis. Final analysis yielded two most-parsimonious cladograms of 242 steps, which support the monophyly of Isodacrys. Isodillex Cortés-Hernández, new genus is here described to accommodate Isodillex minutum (Sharp, 1911), new combination and Isodillex plumosum Cortés-Hernández, new species (Mexico: Zacatecas). Isodillex was recovered as sister group of Isodacrys. Key to separate Isodacrys species, occurrence map and habitus photographs are also provided. creator: Kevin A. Cortés-Hernández creator: Juan J. Morrone uri: https://doi.org/10.7717/peerj.10191 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Cortés-Hernández and Morrone title: Transcriptome analysis of diploid and triploid Populus tomentosa link: https://peerj.com/articles/10204 last-modified: 2020-10-28 description: Triploid Chinese white poplar (Populus tomentosa Carr., Salicaceae) has stronger advantages in growth and better stress resistance and wood quality than diploid P. tomentosa. Using transcriptome sequencing technology to identify candidate transcriptome-based markers for growth vigor in young tree tissue is of great significance for the breeding of P. tomentosa varieties in the future. In this study, the cuttings of diploid and triploid P. tomentosa were used as plant materials, transcriptome sequencing was carried out, and their tissue culture materials were used for RT-qPCR verification of the expression of genes. The results showed that 12,240 differentially expressed genes in diploid and triploid P. tomentosa transcripts were annotated and enriched into 135 metabolic pathways. The top six pathways that enriched the most significantly different genes were plant-pathogen interaction, phenylpropanoid biosynthesis, MAPK signalling pathway-plant, ascorbate and aldarate metabolism, diterpenoid biosynthesis, and the betalain biosynthesis pathway. Ten growth-related genes were selected from pathways of plant hormone signal transduction and carbon fixation in photosynthetic organisms for RT-qPCR verification. The expression levels of MDH and CYCD3 in tissue-cultured and greenhouse planted triploid P. tomentosa were higher than those in tissue-cultured diploid P. tomentosa, which was consist ent with the TMM values calculated by transcriptome. creator: Wen Bian creator: Xiaozhen Liu creator: Zhiming Zhang creator: Hanyao Zhang uri: https://doi.org/10.7717/peerj.10204 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Bian et al. title: Identification and expression analysis of the WRKY gene family during different developmental stages in Lycium ruthenicum Murr. fruit link: https://peerj.com/articles/10207 last-modified: 2020-10-28 description: BackgroundThe WRKY gene family, one of the major transcription factor families in plants, plays crucial regulatory roles in physiological and biological developmental processes, and the adaptation of plants to the environment. However, the systematic study of WRKY structure, expression profiling, and regulatory functions has not been extensively reported in Lycium ruthenicum, although these aspects have been comprehensively studied in most plant species.MethodsIn this study, the WRKY genes were identified from a L. ruthenicum transcriptome database by using bioinformatics. The identification, phylogenetic analysis, zinc-finger structures, and conserved motif prediction were extensively explored. Moreover, the expression levels of 23 selected genes with fragments per kilobase of exons per million mapped reads (FPKM) >5 were assayed during different fruit developmental stages with real-time quantitative polymerase chain reaction (RT-qPCR).ResultsA total of 73 putative WRKY proteins in the L. ruthenicum transcriptome database were identified and examined. Forty-four proteins with the WRKY domain were identified and divided into three major groups with several subgroups, in accordance with those in other plant species. All 44 LrWRKY proteins contained one or two conserved WRKY domains and a zinc-finger structure. Conserved motif prediction revealed conservation of the WRKY DNA-binding domain in L. ruthenicum proteins. The selected LrWRKY genes exhibited discrete expression patterns during different fruit developmental stages. Interestingly, five LrWRKYs (-20, -21, -28, -30, and -31) were expressed remarkably throughout the fruit developmental stages.DiscussionOur results reveal the characteristics of the LrWRKY gene family, thus laying a foundation for further functional analysis of the WRKY family in L. ruthenicum. creator: Richard John Tiika creator: Jia Wei creator: Rui Ma creator: Hongshan Yang creator: Guangxin Cui creator: Huirong Duan creator: Yanjun Ma uri: https://doi.org/10.7717/peerj.10207 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Tiika et al. title: Influences of heel height on human postural stability and functional mobility between inexperienced and experienced high heel shoe wearers link: https://peerj.com/articles/10239 last-modified: 2020-10-28 description: BackgroundHigh heel shoes (HHS) can affect human postural control because elevated heel height (HH) may result in plantar flexed foot and limit ankle joint range of motion during walking. Effects of HH and HHS wearing experience on postural stability during self-initiated and externally triggered perturbations are less examined in the literature. Hence, the objective of the present study is to investigate the influences of HH on human postural stability during dynamic perturbations, perceived stability, and functional mobility between inexperienced and experienced HHS wearers.MethodsA total of 41 female participants were recruited (21 inexperienced HHS wearers and 20 experienced HHS wearers). Sensory organization test (SOT), motor control test (MCT), and limits of stability (LOS) were conducted to measure participant’s postural stability by using computerized dynamic posturography. Functional reach test and timed up and go test were performed to measure functional mobility. The participants’ self-perceived stability was assessed by visual analog scale. Four pairs of shoes with different HH (i.e., 0.8, 3.9, 7.0, and 10.1 cm) were applied to participants randomly. Repeated measures analysis of variance was conducted to detect the effects of HH and HHS wearing experience on each variable.ResultsDuring self-initiated perturbations, equilibrium score remarkably decreased when wearing 10.1 cm compared with flat shoes and 3.9 cm HHS. The contribution of vision to postural stability was larger in 10.1 cm HHS than in flat shoes. The use of ankle strategy worsened when HH increased to 7 cm. Similarly, the directional control of the center of gravity (COG) decreased for 7 cm HHS in LOS. Experienced wearers showed significantly higher percentage of ankle strategy and COG directional control than novices. Under externally triggered perturbations, postural stability was substantially decreased when HH reached 3.9 cm in MCT. No significant difference was found in experienced wearers compared with novices in MCT. Experienced wearers exhibited considerably better functional mobility and perceived stability with increased HH.ConclusionsThe use of HHS may worsen dynamic postural control and functional mobility when HH increases to 3.9 cm. Although experienced HHS wearers exhibit higher proportion of ankle strategy and COG directional control, the experience may not influence overall human postural control. Sensory organization ability, ankle strategy and COG directional control might provide useful information in developing a safety system and prevent HHS wearers from falling. creator: Yiyang Chen creator: Jing Xian Li creator: Lin Wang uri: https://doi.org/10.7717/peerj.10239 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: Effects of clay materials and moisture levels on habitat preference and survivorship of Formosan subterranean termite, Coptotermes formosanus Shiraki (Blattodea: Rhinotermitidae) link: https://peerj.com/articles/10243 last-modified: 2020-10-28 description: Formosan subterranean termites, Coptotermes formosanus Shiraki, usually transport clay materials into tree hollows and bait stations. Our previous research showed that C. formosanus preferred to aggregate in the locations containing field-collected clay samples, but it was not clear whether this preference was influenced by clay types and/or moisture. In the present study, we conducted multiple-choice tests under low-moisture (25% moisture) or moderate-moisture (50% moisture) conditions to evaluate the aggregation and wood-feeding preferences of C. formosanus responding to hollow wooden cylinders (simulation of tree hollows) or baiting containers (simulation of bait stations) filled with different clay materials (bentonite , kaolin, chlorite, illite, or attapulgite), soil, or unfilled. Under low-moisture conditions, the majority of termites were found in the wooden cylinders or baiting containers filled with bentonite. Under moderate-moisture conditions, however, termites preferred to aggregate in wooden cylinders filled with chlorite or attapulgite; the percentages of termites that stayed in baiting containers filled with chlorite, attapulgite or soil were similar, which were significantly higher than those that filled with kaolin, illite, or unfilled. We then conducted no-choice tests to study the effect of clay materials on termites. Under low-moisture conditions, clay filled in the baiting containers significantly increased survivorship and body water percentage (an indicator of termite vigor) of termites, whereas no similar effect was detected under moderate-moisture conditions. This study demonstrated that both clay type and moisture affect termites’ preference. creator: Zhengya Jin creator: Jian Chen creator: Xiujun Wen creator: Cai Wang uri: https://doi.org/10.7717/peerj.10243 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Jin et al. title: Treatment with Bifidobacteria can suppress Aβ accumulation and neuroinflammation in APP/PS1 mice link: https://peerj.com/articles/10262 last-modified: 2020-10-28 description: BackgroundAlzheimer’s disease (AD), being a complex disorder, is affected either by genetic or environmental factors or both. It is observed that there is an excessive accumulation of amyloid β (Aβ) in the extracellular space of the brain. AD is the first neurodegenerative disease in the elderly, and so far there is no effective treatment. In recent years, many studies have reported that Alzheimer’s disease has a relationship with gut microflora, indicating that regulating gut microbiota could offer therapeutic intervention for AD. This study explored the effect Bifidobacteria has in averting AD.MethodsWT and APP/PS1 mice were used for the experiments. The mice were randomly assigned to four groups: WT group, WT + Bi group, AD group (APP/PS1 mouse) and AD + Bi group (Bifidobacteria-treated APP/PS1 mouse). Treatment with Bifidobacteria lasted for 6 months and mice were prepared for immunohistochemistry, immunofluorescence, Thioflavin S staining, Western blotting, PCR and Elisa quantitative assay.ResultsThe results show that after 6 months of treatment with Bifidobacteria signiis to be lesficantly reduces Aβ deposition in cortex and hippocampus of AD mice. The level of insoluble Aβ in the hippocampus and cortex of AD+Bi mice was decreased compared with AD mice. Meanwhile, a significant decrease in the level of soluble Aβ in the cortex of AD+Bi mice but not in the hippocampus was observed. The activation of microglia and the release of inflammatory factors were also determined in this study. From the results, Bifidobacteria inhibited microglial activation and reduced IL-1β, TNF-α, IL-4, IL-6 and INF-γ release. Altogether, these results implied that Bifidobacteria can alleviate the pathological changes of AD through various effects. creator: Qiong Wu creator: Qifa Li creator: Xuan Zhang creator: Michael Ntim creator: Xuefei Wu creator: Ming Li creator: Li Wang creator: Jie Zhao creator: Shao Li uri: https://doi.org/10.7717/peerj.10262 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wu et al. title: Contamination and oxidative stress biomarkers in estuarine fish following a mine tailing disaster link: https://peerj.com/articles/10266 last-modified: 2020-10-28 description: BackgroundThe Rio Doce estuary, in Brazil, was impacted by the deposition of iron mine tailings, caused by the collapse of a dam in 2015. Based on published baseline datasets, the estuary has been experiencing chronic trace metal contamination effects since 2017, with potential bioaccumulation in fishes and human health risks. As metal and metalloid concentrations in aquatic ecosystems pose severe threats to the aquatic biota, we hypothesized that the trace metals in estuarine sediments nearly two years after the disaster would lead to bioaccumulation in demersal fishes and result in the biosynthesis of metal-responsive proteins.MethodsWe measured As, Cd, Cr, Cu, Fe, Mn, Pb, Se and Zn concentrations in sediment samples in August 2017 and compared to published baseline levels. Also, trace metals (As, Cd, Cr, Cu, Fe, Hg, Mn, Pb, Se and Zn) and protein (metallothionein and reduced glutathione) concentrations were quantified in the liver and muscle tissues of five fish species (Cathorops spixii, Genidens genidens, Eugerres brasilianus, Diapterus rhombeus and Mugil sp.) from the estuary, commonly used as food sources by local populations.ResultsOur results revealed high trace metal concentrations in estuarine sediments, when compared to published baseline values for the same estuary. The demersal fish species C. spixii and G. genidens had the highest concentrations of As, Cr, Mn, Hg, and Se in both, hepatic and muscle, tissues. Trace metal bioaccumulation in fish was correlated with the biosynthesis of metallothionein and reduced glutathione in both, liver and muscle, tissues, suggesting active physiological responses to contamination sources. The trace metal concentrations determined in fish tissues were also present in the estuarine sediments at the time of this study. Some elements had concentrations above the maximum permissible limits for human consumption in fish muscles (e.g., As, Cr, Mn, Se and Zn), suggesting potential human health risks that require further studies. Our study supports the high biogeochemical mobility of toxic elements between sediments and the bottom-dwelling biota in estuarine ecosystems. creator: Fabrício Â. Gabriel creator: Rachel Ann Hauser-Davis creator: Lorena Soares creator: Ana Carolina A. Mazzuco creator: Rafael Christian Chavez Rocha creator: Tatiana D. Saint Pierre creator: Enrico Saggioro creator: Fabio Verissimo Correia creator: Tiago O. Ferreira creator: Angelo F. Bernardino uri: https://doi.org/10.7717/peerj.10266 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Gabriel et al. title: Cytotoxic effect of Montivipera bornmuelleri’s venom on cancer cell lines: in vitro and in vivo studies link: https://peerj.com/articles/9909 last-modified: 2020-10-27 description: BackgroundMontivipera bornmuelleri’s venom has shown immunomodulation of cytokines release in mice and selective cytotoxicity on cancer cells in a dose-dependent manner, highlighting an anticancer potential. Here, we extend these findings by elucidating the sensitivity of murine B16 skin melanoma and 3-MCA-induced murine fibrosarcoma cell lines to M. bornmuelleri’s venom and its effect on tumor growth in vivo.MethodsThe toxicity of the venom on B16 and MCA cells was assessed using flow cytometry and xCELLigence assays. For in vivo testing, tumor growth was followed in mice after intratumoral venom injection.ResultsThe venom toxicity showed a dose-dependent cell death on both B16 and MCA cells. Interestingly, overexpression of ovalbumin increased the sensitivity of the cells to the venom. However, the venom was not able to eradicate induced-tumor growth when injected at 100 µg/kg. Our study demonstrates a cytotoxic effect of M. bornmuelleri’s venom in vitro which, however, does not translate to an anticancer action in vivo. creator: Carol Haddoub creator: Mohamad Rima creator: Sandrine Heurtebise creator: Myriam Lawand creator: Dania Jundi creator: Riyad Sadek creator: Sebastian Amigorena creator: Ziad Fajloun creator: Marc C. Karam uri: https://doi.org/10.7717/peerj.9909 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Haddoub et al. title: Reliability of a standing isokinetic shoulder rotators strength test using a functional electromechanical dynamometer: effects of velocity link: https://peerj.com/articles/9951 last-modified: 2020-10-27 description: BackgroundThe evaluation of the force in internal rotation (IR) and external rotation (ER) of the shoulder is commonly used to diagnose possible pathologies or disorders in the glenohumeral joint and to assess patient’s status and progression over time. Currently, there is new technology of multiple joint isokinetic dynamometry that allows to evaluate the strength in the human being. The main purpose of this study was to determine the absolute and relative reliability of concentric and eccentric internal and external shoulder rotators with a functional electromechanical dynamometer (FEMD).MethodsThirty-two male individuals (21.46 ±  2.1 years) were examined of concentric and eccentric strength of shoulder internal and external rotation with a FEMD at velocities of 0.3 m s−1 and 0.6 m s−1. Relative reliability was determined by intraclass correlation coefficients (ICC). Absolute reliability was quantified by standard error of measurement (SEM) and coefficient of variation (CV). Systematic differences across velocities testing circumstances, were analyzed with dependent t tests or repeated-measures analysis of variance in case of 2 or more than 2 conditions, respectively.ResultsReliability was high to excellent for IR and ER on concentric and eccentric strength measurements, regardless of velocity used (ICC: 0.81–0.98, CV: 5.12–8.27% SEM: 4.06–15.04N). Concentric outcomes were more reliable than eccentric due to the possible familiarization of the population with the different stimuli.ConclusionAll procedures examined showed high to excellent reliability for clinical use. However, a velocity of 0.60 m s−1 should be recommended for asymptomatic male patients because it demands less time for evaluation and patients find it more comfortable. creator: Dario Martinez-Garcia creator: Angela Rodriguez-Perea creator: Paola Barboza creator: David Ulloa-Díaz creator: Daniel Jerez-Mayorga creator: Ignacio Chirosa creator: Luis Javier Chirosa Ríos uri: https://doi.org/10.7717/peerj.9951 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Martinez-Garcia et al. title: Functional traits of broad-leaved monocot herbs in the understory and forest edges of a Costa Rican rainforest link: https://peerj.com/articles/9958 last-modified: 2020-10-27 description: Broad-leaved monocot herbs are widespread and dominant components of the shaded understories of wet neotropical forests. These understory habitats are characterized by light limitation and a constant threat of falling branches. Many shaded understory herb species have close relatives that occupy forest edges and gaps, where light availability is higher and defoliation threat is lower, creating an opportunity for comparative analysis of functional traits in order to better understand the evolutionary adaptations associated with this habitat transition. We documented ecological, morphological and ecophysiological traits of multiple herb species in six monocot families from each of these two habitats in the wet tropical rainforest at the La Selva Biological Station, Costa Rica. We found that a mixture of phylogenetic canalization and ecological selection for specific habitats helped explain patterns of functional traits. Understory herbs were significantly shorter and had smaller leaves than forest edge species. Although the mean number of leaves per plant and specific leaf area did not differ between the two groups, the larger leaf size of forest edge species gave them more than three times the mean plant leaf area. Measures of leaf water content and nitrogen content varied within both groups and mean values were not significantly different. Despite the high leaf nitrogen contents, the maximum photosynthetic rates of understory herbs were quite low. Measures of δ13C as an analog of water use efficiency found significantly lower (more negative) values in understory herbs compared to forest edge species. Clonality was strongly developed in several species but did not show strong phylogenetic patterns. This study highlights many functional traits that differ between broad-leaved monocot species characteristic of understory and forest edge habitats, as well as traits that vary primarily by phylogenetic relatedness. Overall, plant functional traits do not provide a simple explanation for the relative differences in abundance for individual understory and forest edge species with some occurring in great abundance while others are relatively rare. creator: Philip W. Rundel creator: Arielle M. Cooley creator: Katharine L. Gerst creator: Erin C. Riordan creator: M. Rasoul Sharifi creator: Jennifer W. Sun creator: J. Alexandra Tower uri: https://doi.org/10.7717/peerj.9958 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Rundel et al. title: Co-application of bio-fertilizer and salicylic acid improves growth, photosynthetic pigments and stress tolerance in wheat under drought stress link: https://peerj.com/articles/9960 last-modified: 2020-10-27 description: Drought stress hampers the growth and productivity of wheat crop worldwide. Thus far, different strategies have been proposed to improve drought tolerance in wheat but the combined application of plant growth-promoting rhizobacteria formulated bio-fertilizer (BF) and salicylic acid (SA) has not been thoroughly explored yet. Therefore, a pot experiment was conducted to observe the effect of SA, BF, and their combination on wheat plants under optimal and drought stress conditions. Seeds priming was done with BF (107 CFU mL−1). After 2 weeks of germination, SA (one mM) was applied as a foliar spray. Drought stress was applied by withholding water supply at three-leaf stage (30 d old plants) for the next 15 d until soil moisture dropped to 10%. Foliar application of SA increased the bacterial population of BF significantly compared to the sole application of BF under irrigated as well as drought stress conditions. Co-application of BF and foliar spray of SA induced drought tolerance in wheat plants by enhancing plant biomass, photosynthetic pigments, relative water content and osmolytes, and activities of the defense-related system. Plants treated with SA and BF together under drought stress had significantly increased leaf water status, Chl a, Chl b, and carotenoids synthesis by 238%, 125%, 167%, and 122%, respectively. Moreover, the co-application of SA and BF showed maximum SOD, POD, APX, and CAT activities by 165%, 85%, 156%, and 169% in the leaves while 153%, 86%, 116% and 200% in roots under drought stress. Similarly, the combined treatment exhibited a pronounced decrease in MDA content by 54% while increased production of proteins and proline by 145% and 149%, respectively. Our results showed that the co-application of SA and BF induced better drought tolerance as compared with the sole application of SA or BF. The results obtained herein suggest that combined application of BF and SA can be applied to the wheat crop to greatly improve drought tolerance in field conditions. creator: Ammar Azmat creator: Humaira Yasmin creator: Muhammad Nadeem Hassan creator: Asia Nosheen creator: Rabia Naz creator: Muhammad Sajjad creator: Noshin Ilyas creator: Malik Nadeem Akhtar uri: https://doi.org/10.7717/peerj.9960 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Azmat et al. title: Optimised biomolecular extraction for metagenomic analysis of microbial biofilms from high-mountain streams link: https://peerj.com/articles/9973 last-modified: 2020-10-27 description: Glacier-fed streams (GFS) are harsh ecosystems dominated by microbial life organized in benthic biofilms, yet the biodiversity and ecosystem functions provided by these communities remain under-appreciated. To better understand the microbial processes and communities contributing to GFS ecosystems, it is necessary to leverage high throughput sequencing. Low biomass and high inorganic particle load in GFS sediment samples may affect nucleic acid extraction efficiency using extraction methods tailored to other extreme environments such as deep-sea sediments. Here, we benchmarked the utility and efficacy of four extraction protocols, including an up-scaled phenol-chloroform protocol. We found that established protocols for comparable sample types consistently failed to yield sufficient high-quality DNA, delineating the extreme character of GFS. The methods differed in the success of downstream applications such as library preparation and sequencing. An adapted phenol-chloroform-based extraction method resulted in higher yields and better recovered the expected taxonomic profile and abundance of reconstructed genomes when compared to commercially-available methods. Affordable and straight-forward, this method consistently recapitulated the abundance and genomes of a mock community, including eukaryotes. Moreover, by increasing the amount of input sediment, the protocol is readily adjustable to the microbial load of the processed samples without compromising protocol efficiency. Our study provides a first systematic and extensive analysis of the different options for extraction of nucleic acids from glacier-fed streams for high-throughput sequencing applications, which may be applied to other extreme environments. creator: Susheel Bhanu Busi creator: Paraskevi Pramateftaki creator: Jade Brandani creator: Stilianos Fodelianakis creator: Hannes Peter creator: Rashi Halder creator: Paul Wilmes creator: Tom J. Battin uri: https://doi.org/10.7717/peerj.9973 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Busi et al. title: Vertical distribution of polycyclic aromatic hydrocarbons in the brackish sea water column: ex situ experiment link: https://peerj.com/articles/10087 last-modified: 2020-10-27 description: BackgroundOil spills can cause severe damage within a marine ecosystem. Following a spill, the soluble fraction of polycyclic aromatic hydrocarbons is rapidly released into the water column. These remain dissolved in seawater over an extended period of time, even should the insoluble fraction be removed. The vertical distribution of the aromatic hydrocarbon component and how these become transferred is poorly understood in brackish waters. This study examines the vertical distribution of polycyclic aromatic hydrocarbons having been released from a controlled film of spilled oil onto the surface of brackish water.MethodsThe study was undertaken under controlled conditions so as to minimize the variability of environmental factors such as temperature and hydrodynamics. The distribution of polycyclic aromatic hydrocarbons was measured in the dissolved and suspended phases throughout the 1 m water column with different intensity of water sampling: 1, 2, 4, 7, 72, 120, 336, 504 and 984 h.ResultsThe total concentration of polycyclic aromatic hydrocarbons ranged from 19.01 to 214.85 ng L–1 in the dissolved phase and from 5.14 to 63.92 ng L–1 in the suspended phase. These hydrocarbons were released immediately following a controlled spill attaining 214.9 ng L–1 in the dissolved phase and 54.4 ng L–1 in the suspended phase near the cylinder bottom after 1–2 h. The 2–3 ring polycyclic aromatic hydrocarbons dominated in the dissolved phase (60–80%), whereas the greater amount of 4–6 ring polycyclic aromatic hydrocarbons (55–90%) occurred in the suspended phase. A relatively low negative correlation (rS = –0.41) was determined between the concentration of phenanthrene and suspended matter, whereas a high negative correlation (r =  − 0.79) was found between the concentration of pyrene and suspended matter. Despite the differences in the relationships between the concentration ratio and amount of suspended matter the obtained regressions allow roughly to predict the concentration of polycyclic aromatic hydrocarbons. creator: Zilvinas Kryzevicius creator: Kristina Mickuviene creator: Martynas Bucas creator: Monika Vilkiene creator: Audrone Zukauskaite uri: https://doi.org/10.7717/peerj.10087 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kryzevicius et al. title: Effects of different drying methods on smears of canine blood and effusion fluid link: https://peerj.com/articles/10092 last-modified: 2020-10-27 description: BackgroundGlass slide preparations from a variety of specimens (blood, masses, effusions) are commonly made as part of the diagnostic work-up, however the effects of various drying methods in veterinary practice and diagnostic laboratory settings is not clear.ObjectiveCompare the effects of four drying methods on results of microscopic examination of canine blood smears and direct smears of pleural or peritoneal effusion fluid.MethodsTwelve canine blood samples (6 from healthy dogs, 6 from sick dogs) and 6 canine peritoneal or pleural effusion samples. Four smears were prepared from each of the 18 samples and dried using the following methods: air-dry, hair dryer with or without heat, and heat block at 58 °C. Observers, blinded to the drying method, independently reviewed the slides microscopically, using a scoring system to evaluate cell morphology and (for blood smears) echinocyte numbers; scoring results were analyzed statistically.ResultsFor blood smears, several comparisons showed more adverse effects on morphology using the heat block method than for one or more other drying methods. For effusion fluid smears, RBCs dried with the heat block or air-dry methods had more poorly preserved morphology than RBCs dried by the hair dryer method without heat.Conclusions and clinical relevanceThe results (1) indicate that different drying methods had a significant effect, (2) support using a hair dryer without heat for both blood smears and effusion fluid smears, and (3) discourage using a 58 °C heat block. creator: Fiamma G. De Witte creator: Aimee Hebrard creator: Carolyn N. Grimes creator: Kristin Owens creator: Deanna M. Schaefer creator: Xiaojuan Zhu creator: Michael M. Fry uri: https://doi.org/10.7717/peerj.10092 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 De Witte et al. title: The osteology and phylogenetic position of the loricatan (Archosauria: Pseudosuchia) Heptasuchus clarki, from the ?Mid-Upper Triassic, southeastern Big Horn Mountains, Central Wyoming (USA) link: https://peerj.com/articles/10101 last-modified: 2020-10-27 description: Loricatan pseudosuchians (known as “rauisuchians”) typically consist of poorly understood fragmentary remains known worldwide from the Middle Triassic to the end of the Triassic Period. Renewed interest and the discovery of more complete specimens recently revolutionized our understanding of the relationships of archosaurs, the origin of Crocodylomorpha, and the paleobiology of these animals. However, there are still few loricatans known from the Middle to early portion of the Late Triassic and the forms that occur during this time are largely known from southern Pangea or Europe. Heptasuchus clarki was the first formally recognized North American “rauisuchian” and was collected from a poorly sampled and disparately fossiliferous sequence of Triassic strata in North America. Exposed along the trend of the Casper Arch flanking the southeastern Big Horn Mountains, the type locality of Heptasuchus clarki occurs within a sequence of red beds above the Alcova Limestone and Crow Mountain formations within the Chugwater Group. The age of the type locality is poorly constrained to the Middle—early Late Triassic and is likely similar to or just older than that of the Popo Agie Formation assemblage from the western portion of Wyoming. The holotype consists of associated cranial elements found in situ, and the referred specimens consist of crania and postcrania. Thus, about 30% of the osteology of the taxon is preserved. All of the pseudosuchian elements collected at the locality appear to belong to Heptasuchus clarki and the taxon is not a chimera as previously hypothesized. Heptasuchus clarki is distinct from all other archosaurs by the presence of large, posteriorly directed flanges on the parabasisphenoid and a distinct, orbit-overhanging postfrontal. Our phylogenetic hypothesis posits a sister-taxon relationship between Heptasuchus clarki and the Ladinian-aged Batrachotomus kupferzellensis from current-day Germany within Loricata. These two taxa share a number of apomorphies from across the skull and their phylogenetic position further supports ‘rauisuchian’ paraphyly. A minimum of three individuals of Heptasuchus are present at the type locality suggesting that a group of individuals died together, similar to other aggregations of loricatans (e.g., Heptasuchus, Batrachotomus, Decuriasuchus, Postosuchus). creator: Sterling J. Nesbitt creator: John M. Zawiskie creator: Robert M. Dawley uri: https://doi.org/10.7717/peerj.10101 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Nesbitt et al. title: Novel targets for engineering Physostegia chlorotic mottle and tomato brown rugose fruit virus-resistant tomatoes: in silico prediction of tomato microRNA targets link: https://peerj.com/articles/10096 last-modified: 2020-10-27 description: BackgroundPhysostegia chlorotic mottle virus (PhCMoV; genus: Alphanucleorhabdovirus, family: Rhabdoviridae) and tomato brown rugose fruit virus (ToBRFV; genus: Tobamovirus, family: Virgaviridae) are newly emerging plant viruses that have a dramatic effect on tomato production. Among various known virus-control strategies, RNAi-mediated defence has shown the potential to protect plants against various pathogens including viral infections. Micro(mi)RNAs play a major role in RNAi-mediated defence.MethodsUsing in silico analyses, we investigated the possibility of tomato-encoded miRNAs (TomiRNA) to target PhCMoV and ToBRFV genomes using five different algorithms, i.e., miRanda, RNAhybrid, RNA22, Tapirhybrid and psRNATarget.ResultsThe results revealed that 14 loci on PhCMoV and 10 loci on ToBRFV can be targeted by the TomiRNAs based on the prediction of at least three algorithms. Interestingly, one TomiRNA, miR6026, can target open reading frames from both viruses, i.e., the phosphoprotein encoding gene of PhCMoV, and the two replicase components of ToBRFV. There are currently no commercially available PhCMoV- or ToBRFV-resistant tomato varieties, therefore the predicted data provide useful information for the development of PhCMoV- and ToBFRV-resistant tomato plants. creator: Yahya Zakaria Abdou Gaafar creator: Heiko Ziebell uri: https://doi.org/10.7717/peerj.10096 license: https://creativecommons.org/licenses/by-nc/4.0 rights: ©2020 Gaafar and Ziebell title: The Bacteria Genome Pipeline (BAGEP): an automated, scalable workflow for bacteria genomes with Snakemake link: https://peerj.com/articles/10121 last-modified: 2020-10-27 description: Next generation sequencing technologies are becoming more accessible and affordable over the years, with entire genome sequences of several pathogens being deciphered in few hours. However, there is the need to analyze multiple genomes within a short time, in order to provide critical information about a pathogen of interest such as drug resistance, mutations and genetic relationship of isolates in an outbreak setting. Many pipelines that currently do this are stand-alone workflows and require huge computational requirements to analyze multiple genomes. We present an automated and scalable pipeline called BAGEP for monomorphic bacteria that performs quality control on FASTQ paired end files, scan reads for contaminants using a taxonomic classifier, maps reads to a reference genome of choice for variant detection, detects antimicrobial resistant (AMR) genes, constructs a phylogenetic tree from core genome alignments and provide interactive short nucleotide polymorphism (SNP) visualization across core genomes in the data set. The objective of our research was to create an easy-to-use pipeline from existing bioinformatics tools that can be deployed on a personal computer. The pipeline was built on the Snakemake framework and utilizes existing tools for each processing step: fastp for quality trimming, snippy for variant calling, Centrifuge for taxonomic classification, Abricate for AMR gene detection, snippy-core for generating whole and core genome alignments, IQ-TREE for phylogenetic tree construction and vcfR for an interactive heatmap visualization which shows SNPs at specific locations across the genomes. BAGEP was successfully tested and validated with Mycobacterium tuberculosis (n = 20) and Salmonella enterica serovar Typhi (n = 20) genomes which are about 4.4 million and 4.8 million base pairs, respectively. Running these test data on a 8 GB RAM, 2.5 GHz quad core laptop took 122 and 61 minutes on respective data sets to complete the analysis. BAGEP is a fast, calls accurate SNPs and an easy to run pipeline that can be executed on a mid-range laptop; it is freely available on: https://github.com/idolawoye/BAGEP. creator: Idowu B. Olawoye creator: Simon D.W. Frost creator: Christian T. Happi uri: https://doi.org/10.7717/peerj.10121 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Olawoye et al. title: The size of signal detection and emission organs in a synchronous firefly: sexual dimorphism, allometry and assortative mating link: https://peerj.com/articles/10127 last-modified: 2020-10-27 description: The size of the organs responsible for emitting and detecting sexual communication signals is a likely target for selection. Communication via bioluminescent signals in synchronous fireflies is a promising model to test hypotheses regarding differences between males and females in the effect of the size of signal emission and detection organs on fitness components. Synchronous firefly species congregate in large numbers during the mating season, displaying bioluminescent signals aimed at potential mates during relatively short nightly periods. Operational sex ratios are male-biased and, thus, the so-called typical sex roles (indiscriminate males and choosy females) are expected to evolve. We studied the synchronous firefly Photinus palaciosi, a species that during the mating season congregates in forests of central Mexico offering a magnificent natural show that attracts numerous tourists. P. palaciosi females have reduced wings (brachyptery) and cannot fly. Our field study tested the hypothesis that the male-biased operational sex ratio and the short daily mating period result in strong male-male competition that selects for males with larger lanterns and larger eyes, and against male mate choice, whereas female-female mate competition is absent and, thus, no selection on lantern or eye size is expected. Even though lantern, eye or body size do not predict the probability of being found in copula for either sex, sexual dimorphism in these features, along with allometric slopes of lantern size and assortative mating in terms of relative lantern size, support not only the hypothesis of intense sexual selection among males, but the possibility of subtle mechanisms of sexual selection among females. Trade-offs between investment in signaling (lanterns) versus detection (eyes) structures, or with pressures different from sexual selection such as those imposed by predators, are also likely to be important in shaping the evolution of sexual signaling in these fireflies. creator: Tania López-Palafox creator: Rogelio Macías-Ordóñez creator: Carlos R. Cordero uri: https://doi.org/10.7717/peerj.10127 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 López-Palafox et al. title: Integrated transcriptome meta-analysis of pancreatic ductal adenocarcinoma and matched adjacent pancreatic tissues link: https://peerj.com/articles/10141 last-modified: 2020-10-27 description: A comprehensive meta-analysis of publicly available gene expression microarray data obtained from human-derived pancreatic ductal adenocarcinoma (PDAC) tissues and their histologically matched adjacent tissue samples was performed to provide diagnostic and prognostic biomarkers, and molecular targets for PDAC. An integrative meta-analysis of four submissions (GSE62452, GSE15471, GSE62165, and GSE56560) containing 105 eligible tumor-adjacent tissue pairs revealed 344 differentially over-expressed and 168 repressed genes in PDAC compared to the adjacent-to-tumor samples. The validation analysis using TCGA combined GTEx data confirmed 98.24% of the identified up-regulated and 73.88% of the down-regulated protein-coding genes in PDAC. Pathway enrichment analysis showed that “ECM-receptor interaction”, “PI3K-Akt signaling pathway”, and “focal adhesion” are the most enriched KEGG pathways in PDAC. Protein-protein interaction analysis identified FN1, TIMP1, and MSLN as the most highly ranked hub genes among the DEGs. Transcription factor enrichment analysis revealed that TCF7, CTNNB1, SMAD3, and JUN are significantly activated in PDAC, while SMAD7 is inhibited. The prognostic significance of the identified and validated differentially expressed genes in PDAC was evaluated via survival analysis of TCGA Pan-Cancer pancreatic ductal adenocarcinoma data. The identified candidate prognostic biomarkers were then validated in four external validation datasets (GSE21501, GSE50827, GSE57495, and GSE71729) to further improve reliability. A total of 28 up-regulated genes were found to be significantly correlated with worse overall survival in patients with PDAC. Twenty-one of the identified prognostic genes (ITGB6, LAMC2, KRT7, SERPINB5, IGF2BP3, IL1RN, MPZL2, SFTA2, MET, LAMA3, ARNTL2, SLC2A1, LAMB3, COL17A1, EPSTI1, IL1RAP, AK4, ANXA2, S100A16, KRT19, and GPRC5A) were also found to be significantly correlated with the pathological stages of the disease. The results of this study provided promising prognostic biomarkers that have the potential to differentiate PDAC from both healthy and adjacent-to-tumor pancreatic tissues. Several novel dysregulated genes merit further study as potentially promising candidates for the development of more effective treatment strategies for PDAC. creator: Sevcan Atay uri: https://doi.org/10.7717/peerj.10141 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Atay title: Developing mathematical model for diurnal dynamics of photosynthesis in Saccharum officinarum responsive to different irrigation and silicon application link: https://peerj.com/articles/10154 last-modified: 2020-10-27 description: In the dynamic era of climate change, agricultural farming systems are facing various unprecedented problems worldwide. Drought stress is one of the serious abiotic stresses that hinder the growth potential and crop productivity. Silicon (Si) can improve crop yield by enhancing the efficiency of inputs and reducing relevant losses. As a quasi-essential element and the 2nd most abundant element in the Earth’s crust, Si is utilized by plants and applied exogenously to combat drought stress and improve plant performance by increasing physiological, cellular and molecular responses. However, the physiological mechanisms that respond to water stress are still not well defined in Saccharum officinarum plants. To the best of our knowledge, the dynamics of photosynthesis responsive to different exogenous Si levels in Saccharum officinarum has not been reported to date. The current experiment was carried out to assess the protective role of Si in plant growth and photosynthetic responses in Saccharum officinarum under water stress conditions. Saccharum officinarum cv. ‘GT 42’ plants were subjected to drought stress conditions (80–75%, 55–50% and 35–30% of soil moisture) after ten weeks of normal growth, followed by the soil irrigation of Si (0, 100, 300 and 500 mg L−1) for 8 weeks. The results indicated that Si addition mitigated the inhibition in Saccharum officinarum growth and photosynthesis, and improved biomass accumulation during water stress. The photosynthetic responses (photosynthesis, transpiration and stomatal conductance) were found down-regulated under water stress, and it was significantly enhanced by Si application. No phytotoxic effects were monitored even at excess (500 mg L−1). Soil irrigation of 300 mg L−1 of Si was more effective as 100 and 500 mg L−1 under water stress condition. It is concluded that the stress in Saccharum officinarum plants applied with Si was alleviated by improving plant fitness, photosynthetic capacity and biomass accumulation as compared with the control. Thus, this study offers new information towards the assessment of growth, biomass accumulation and physiological changes related to water stress with Si application in plants. creator: Krishan K. Verma creator: Kai-Chao Wu creator: Chhedi Lal Verma creator: Dong-Mei Li creator: Mukesh Kumar Malviya creator: Rajesh Kumar Singh creator: Pratiksha Singh creator: Gan-Lin Chen creator: Xiu Peng Song creator: Yang Rui Li uri: https://doi.org/10.7717/peerj.10154 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Verma et al. title: Comparing sediment DNA extraction methods for assessing organic enrichment associated with marine aquaculture link: https://peerj.com/articles/10231 last-modified: 2020-10-27 description: Marine sediments contain a high diversity of micro- and macro-organisms which are important in the functioning of biogeochemical cycles. Traditionally, anthropogenic perturbation has been investigated by identifying macro-organism responses along gradients. Environmental DNA (eDNA) analyses have recently been advocated as a rapid and cost-effective approach to measuring ecological impacts and efforts are underway to incorporate eDNA tools into monitoring. Before these methods can replace or complement existing methods, robustness and repeatability of each analytical step has to be demonstrated. One area that requires further investigation is the selection of sediment DNA extraction method. Environmental DNA sediment samples were obtained along a disturbance gradient adjacent to a Chinook (Oncorhynchus tshawytscha) salmon farm in Otanerau Bay, New Zealand. DNA was extracted using four extraction kits (Qiagen DNeasy PowerSoil, Qiagen DNeasy PowerSoil Pro, Qiagen RNeasy PowerSoil Total RNA/DNA extraction/elution and Favorgen FavorPrep Soil DNA Isolation Midi Kit) and three sediment volumes (0.25, 2, and 5 g). Prokaryotic and eukaryotic communities were amplified using primers targeting the 16S and 18S ribosomal RNA genes, respectively, and were sequenced on an Illumina MiSeq. Diversity and community composition estimates were obtained from each extraction kit, as well as their relative performance in established metabarcoding biotic indices. Differences were observed in the quality and quantity of the extracted DNA amongst kits with the two Qiagen DNeasy PowerSoil kits performing best. Significant differences were observed in both prokaryotes and eukaryotes (p < 0.001) richness among kits. A small proportion of amplicon sequence variants (ASVs) were shared amongst the kits (~3%) although these shared ASVs accounted for the majority of sequence reads (prokaryotes: 59.9%, eukaryotes: 67.2%). Differences were observed in the richness and relative abundance of taxonomic classes revealed with each kit. Multivariate analysis showed that there was a significant interaction between “distance” from the farm and “kit” in explaining the composition of the communities, with the distance from the farm being a stronger determinant of community composition. Comparison of the kits against the bacterial and eukaryotic metabarcoding biotic index suggested that all kits showed similar patterns along the environmental gradient. Overall, we advocate for the use of Qiagen DNeasy PowerSoil kits for use when characterizing prokaryotic and eukaryotic eDNA from marine farm sediments. We base this conclusion on the higher DNA quality values and richness achieved with these kits compared to the other kits/amounts investigated in this study. The additional advantage of the PowerSoil Kits is that DNA extractions can be performed using an extractor robot, offering additional standardization and reproducibility of results. creator: John K. Pearman creator: Nigel B. Keeley creator: Susanna A. Wood creator: Olivier Laroche creator: Anastasija Zaiko creator: Georgia Thomson-Laing creator: Laura Biessy creator: Javier Atalah creator: Xavier Pochon uri: https://doi.org/10.7717/peerj.10231 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Pearman et al. title: Reinstatement of Phascolosoma (Phascolosoma) varians Keferstein, 1865 (Sipuncula: Phascolosomatidae) based on morphological and molecular data link: https://peerj.com/articles/10238 last-modified: 2020-10-27 description: Phascolosoma (P.) varians, a sipunculan species known from the Greater Caribbean, was designated as a synonym of Phascolosoma (P.) nigrescens, which was originally described from Fiji. Their synonymy was primarily based upon an interpretation that these two species were morphologically indistinguishable. After its designation as a synonym, no further detailed analyses of morphological or molecular characteristics were performed to corroborate the assumed widespread distribution of Phascolosoma (P.) nigrescens. In this study, Phascolosoma (P.) varians is redescribed, and notable differences between this species and its proposed senior synonym are presented. These two species differ in the shape of their hooks, the spatial attachment of nephridia to the body wall, and the morphology of the contractile vessel. Additionally, there is high genetic divergence between nucleotide sequences within their respective cytochrome c oxidase subunit 1 (COI) genes, which supports the morphological data. Herein, the synonymy of Phascolosoma (P.) varians with Phascolosoma (P.) nigrescens is rejected due to morphological and molecular differences. Furthermore, the assumed widespread distribution of Phascolosoma (P.) nigrescens is still considered as questionable. creator: Itzahi Silva-Morales uri: https://doi.org/10.7717/peerj.10238 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Silva-Morales title: An improved modified early warning score that incorporates the abdomen score for identifying multiple traumatic injury severity link: https://peerj.com/articles/10242 last-modified: 2020-10-27 description: BackgroundRapid identification of trauma severity is essential for the timely triage of multiple trauma patients. Tools such as the modified early warning score (MEWS) are used for determining injury severity. Although the conventional MEWS is a good predictor of mortality, its performance assessing injury severity is moderate. This study hypothesized that adding an injury site severity-related score (e.g., abdomen score) may enhance the capability of the MEWS for identifying severe trauma.MethodTo validate the hypothesis, we propose an improved modified early warning score called MEWS-A, which incorporates an injury site-specific severity-related abdomen score to MEWS. The utility of MEWS and MEWS-A were retrospectively evaluated and compared for identifying trauma severity in adult multiple trauma patients admitted to the emergency department.ResultsWe included 1,230 eligible multiple trauma patients and divided them into minor and severe trauma groups based on the injury severity score. Results of logistic regression and receiver operating characteristic (ROC) curve analyses showed that the MEWS-A had a higher area under the ROC curve (AUC: 0.81 95% CI [0.78–0.83]) than did the MEWS (AUC: 0.77 95% CI [0.74–0.79]), indicating that the MEWS-A is superior to the MEWS in identifying severe trauma. The optimal MEWS-A cut-off score is 4, with a specificity of 0.93 and a sensitivity of 0.54. MEWS-A ≥ 4 can be used as a protocol for decision-making in the emergency department.ConclusionsOur study suggests that while the conventional MEWS is sufficient for predicting mortality risk, adding an injury site-specific score (e.g., abdomen score) can enhance its performance in determining injury severity in multiple trauma patients. creator: Xiaobin Jiang uri: https://doi.org/10.7717/peerj.10242 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Jiang title: Gynomonoecy in a mycoheterotrophic orchid Eulophia zollingeri with autonomous selfing hermaphroditic flowers and putatively outcrossing female flowers link: https://peerj.com/articles/10272 last-modified: 2020-10-27 description: Most orchid species exhibit an extreme case of hermaphroditism, owing to the fusion of male and female organs into a gynostemium. Exceptions to this rule have only been reported from the subtribes Catasetinae and Satyriinae. Here, I report an additional orchidaceous example whose flowers are not always hermaphroditic. In several Japanese populations of Eulophia zollingeri (Rchb.f.) J.J.Sm, a widespread Asian and Oceanian orchid, some flowers possess both the anther (i.e., anther cap and pollinaria) and stigma, whereas others possess only the stigma. Therefore, pollination experiments, an investigation of floral morphology and observations of floral visitors were conducted to understand the reproductive biology of E. zollingeri in Miyazaki Prefecture, Japan. It was confirmed that E. zollingeri studied here possesses a gynomonoecious reproductive system, a sexual system in which a single plant has both female flowers and hermaphroditic flowers. In addition, hermaphroditic flowers often possess an effective self-pollination system while female flowers could avoid autogamy but suffered from severe pollinator limitation, due to a lack of agamospermy and low insect-mediated pollination. The present study represents the first documented example of gynomonoecy within Orchidaceae. Gynomonoecy in E. zollingeri may be maintained by the tradeoff in reproductive traits between female flowers (with low fruit set but potential outcrossing benefits) and hermaphroditic flowers (with high fruit set but inbreeding depression in selfed offspring). This mixed mating is probably important in mycoheterotrophic E. zollingeri because it occurs in shaded forest understorey with a paucity of pollinators. creator: Kenji Suetsugu uri: https://doi.org/10.7717/peerj.10272 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Suetsugu title: Haiti has more forest than previously reported: land change 2000–2015 link: https://peerj.com/articles/9919 last-modified: 2020-10-26 description: Estimates of forest cover have important political, conservation, and funding implications, but methods vary greatly. Haiti has often been cited as one of the most deforested countries in the world, yet estimates of forest cover range from <1% to 33%. Here, we analyze land change for seven land cover classes (forest, shrub land, agriculture/pasture, plantation, urban/infrastructure, barren land, and water) between 2000 and 2015 using Landsat imagery (30 m resolution) in the Google Earth Engine platform. Forest cover was estimated at 26% in 2000 and 21% in 2015. Although forest cover is declining in Haiti, our quantitative analysis resulted in considerably higher forest cover than what is usually reported by local and international institutions. Our results determined that areas of forest decline were mainly converted to shrubs and mixed agriculture/pasture. An important driver of forest loss and degradation could be the high demand for charcoal, which is the principal source of cooking fuel. Our results differ from other forest cover estimates and forest reports from national and international institutions, most likely due to differences in forest definition, data sources, spatial resolution, and methods. In the case of Haiti, this work demonstrates the need for clear and functional definitions and classification methods to accurately represent land use/cover change. Regardless of how forests are defined, forest cover in Haiti will continue to decline unless corrective actions are taken to protect remaining forest patches. This can serve as a warning of the destructive land use patterns and can help us target efforts for better planning, management, and conservation. creator: Ose Pauleus creator: T. Mitchell Aide uri: https://doi.org/10.7717/peerj.9919 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Pauleus and Aide title: Effects of changes in isotopic baselines on the evaluation of food web structure using isotopic functional indices link: https://peerj.com/articles/9999 last-modified: 2020-10-26 description: BackgroundThis study aimed to assess whether ecological inferences from isotopic functional indices (IFIs) are impacted by changes in isotopic baselines in aquatic food webs. We used sudden CO2-outgassing and associated shifts in DIC-δ13C brought by waterfalls as an excellent natural experimental set-up to quantify impacts of changes in algal isotopic baselines on ecological inferences from IFIs.MethodsCarbon (δ13C) and nitrogen (δ15N) stable isotopic ratios of invertebrate communities sharing similar structure were measured at above- and below-waterfall sampling sites from five rivers and streams in Southern Quebec (Canada). For each sampled invertebrate community, the six Laymans IFIs were then calculated in the δ -space (δ13C vs. δ15N).ResultsAs expected, isotopic functional richness indices, measuring the overall extent of community trophic space, were strongly sensitive to changes in isotopic baselines unlike other IFIs. Indeed, other IFIs were calculated based on the distribution of species within δ-space and were not strongly impacted by changes in the vertical or horizontal distribution of specimens in the δ-space. Our results highlighted that IFIs exhibited different sensitivities to changes in isotopic baselines, leading to potential misinterpretations of IFIs in river studies where isotopic baselines generally show high temporal and spatial variabilities. The identification of isotopic baselines and their associated variability, and the use of independent trophic tracers to identify the actual energy pathways through food webs must be a prerequisite to IFIs-based studies to strengthen the reliability of ecological inferences of food web structural properties. creator: Simon Belle creator: Gilbert Cabana uri: https://doi.org/10.7717/peerj.9999 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Belle and Cabana title: Conservation genomics and pollination biology of an endangered, edaphic-endemic, octoploid herb: El Dorado bedstraw (Galium californicum subsp. sierrae; Rubiaceae) link: https://peerj.com/articles/10042 last-modified: 2020-10-26 description: El Dorado bedstraw (Galium californicum subsp. sierrae) is a federally endangered dioecious, octoploid, perennial herb found only in the Pine Hill region of El Dorado County, CA, USA. Like many species of Galium, El Dorado bedstraw is capable of both sexual and asexual reproduction, spreading via stem-layering as well as seeds. El Dorado bedstraw is also dioecious, and thus dependent on pollinators to transfer pollen from male to female stems. The capacity for asexual reproduction has conservation implications for this plant, due to the potential for populations to become dominated by a small number of clones in the absence of recruitment from seeds. No previous work has examined either the population genetics or pollination biology of this plant. Here, double-digest restriction site-associated DNA sequencing was used to develop a genetic dataset for a sample of El Dorado bedstraw (12 individuals from each of seven locations). Genomic data was used to calculate population genetic statistics and quantify the degree to which clonality affects the sampled populations. Visual observation of insect visitors at every sampling location was used to assess the potential for pollen transfer within and among locations. A total of 23 clonal colonies were detected across 82 successfully sequenced stems, consisting of an average of 2.4 individuals (range: 2–6). Significant isolation by distance among locations was detected using a Mantel test. Insect pollinators were from eleven families, consisting mainly of small species with weak flight. It is recommended that clonality and small-scale population differentiation be taken into account in conservation measures. creator: Dylan Burge uri: https://doi.org/10.7717/peerj.10042 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Burge title: New record of podocopid ostracods from Cretaceous amber link: https://peerj.com/articles/10134 last-modified: 2020-10-26 description: Burmese Cretaceous amber (∼99 Ma, Myanmar) is famous for the preservation of a wide range of fauna and flora, including representatives of marine, freshwater and terrestrial groups. Here, we report on three ostracod specimens, that came visible as syninclusions to an aquatic isopod. The three specimens represent three different taxa, that were found preserved in a single piece of amber. One of the described specimens was studied using µCT scanning data. On the basis of general carapace morphology we assign all three to the group Podocopida, and (tentatively) its ingroup Cypridocopina. A lack of visibility of more particular diagnostic features such as adductor muscle scars and details of the marginal zone precludes a further identification, but we discuss possible affinities with either the marine-brackish group Pontocypridoidea or the non-marine group Cypridoidea. The taphonomy indicates that the studied ostracods had been subject to limited (if any) post-mortem transport, which could be consistent with marginal marine environments. creator: He Wang creator: Mario Schädel creator: Benjamin Sames creator: David J. Horne uri: https://doi.org/10.7717/peerj.10134 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: Spatial modeling could not differentiate early SARS-CoV-2 cases from the distribution of humans on the basis of climate in the United States link: https://peerj.com/articles/10140 last-modified: 2020-10-26 description: The SARS-CoV-2 coronavirus is wreaking havoc globally, yet, as a novel pathogen, knowledge of its biology is still emerging. Climate and seasonality influence the distributions of many diseases, and studies suggest at least some link between SARS-CoV-2 and weather. One such study, building species distribution models (SDMs), predicted SARS-CoV-2 risk may remain concentrated in the Northern Hemisphere, shifting northward in summer months. Others have highlighted issues with SARS-CoV-2 SDMs, notably: the primary niche of the virus is the host it infects, climate may be a weak distributional predictor, global prevalence data have issues, and the virus is not in population equilibrium. While these issues should be considered, we believe climate’s relationship with SARS-CoV-2 is still worth exploring, as it may have some impact on the distribution of cases. To further examine if there is a link to climate, we build model projections with raw SARS-CoV-2 case data and population-scaled case data in the USA. The case data were from across March 2020, before large travel restrictions and public health policies were impacting cases across the country. We show that SDMs built from population-scaled case data cannot be distinguished from control models (built from raw human population data), while SDMs built on raw case data fail to predict the known distribution of cases in the U.S. from March. The population-scaled analyses indicate that climate did not play a central role in early U.S. viral distribution and that human population density was likely the primary driver. We do find slightly more population-scaled viral cases in cooler areas. Ultimately, the temporal and geographic constraints on this study mean that we cannot rule out climate as a partial driver of the SARS-CoV-2 distribution. Climate’s role on SARS-CoV-2 should continue to be cautiously examined, but at this time we should assume that SARS-CoV-2 will continue to spread anywhere in the U.S. where governmental policy does not prevent spread. creator: Robert Harbert creator: Seth W. Cunningham creator: Michael Tessler uri: https://doi.org/10.7717/peerj.10140 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Harbert et al. title: Towards a unified generic framework to define and observe contacts between livestock and wildlife: a systematic review link: https://peerj.com/articles/10221 last-modified: 2020-10-26 description: Wild animals are the source of many pathogens of livestock and humans. Concerns about the potential transmission of economically important and zoonotic diseases from wildlife have led to increased surveillance at the livestock-wildlife interface. Knowledge of the types, frequency and duration of contacts between livestock and wildlife is necessary to identify risk factors for disease transmission and to design possible mitigation strategies. Observing the behaviour of many wildlife species is challenging due to their cryptic nature and avoidance of humans, meaning there are relatively few studies in this area. Further, a consensus on the definition of what constitutes a ‘contact’ between wildlife and livestock is lacking. A systematic review was conducted to investigate which livestock-wildlife contacts have been studied and why, as well as the methods used to observe each species. Over 30,000 publications were screened, of which 122 fulfilled specific criteria for inclusion in the analysis. The majority of studies examined cattle contacts with badgers or with deer; studies involving wild pig contacts with cattle or with domestic pigs were the next most frequent. There was a range of observational methods including motion-activated cameras and global positioning system collars. As a result of the wide variation and lack of consensus in the definitions of direct and indirect contacts, we developed a unified framework to define livestock-wildlife contacts that is sufficiently flexible to be applied to most wildlife and livestock species for non-vector-borne diseases. We hope this framework will help standardise the collection and reporting of contact data; a valuable step towards being able to compare the efficacy of wildlife-livestock observation methods. In doing so, it may aid the development of better disease transmission models and improve the design and effectiveness of interventions to reduce or prevent disease transmission. creator: Sonny A. Bacigalupo creator: Linda K. Dixon creator: Simon Gubbins creator: Adam J. Kucharski creator: Julian A. Drewe uri: https://doi.org/10.7717/peerj.10221 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Bacigalupo et al. title: ZFPM2-AS1 promotes the proliferation, migration, and invasion of human non-small cell lung cancer cells involving the JAK-STAT and AKT pathways link: https://peerj.com/articles/10225 last-modified: 2020-10-26 description: PurposeRecent studies have determined that long non-coding RNAs (lncRNAs) are potential prognostic biomarkers for non-small cell lung cancers (NSCLCs). The purpose of this study was to analyze the function and associated pathways of zinc finger protein multitype 2 antisense RNA 1 (ZFPM2-AS1) in NSCLC cells.MethodsWe used qRT-PCR to analyze ZFPM2-AS1’s transcription level. Its proliferation, migration, and invasion capacities were determined using MTT, colony forming, wound healing, and transwell assays. We additionally analyzed the correlation between ZFPM2 and immune infiltration using the Tumor Immune Estimation Resource (TIMER) database, and the protein expression levels using Western blots.ResultsWe found that ZFPM2-AS1 expression in NSCLC specimens and cell lines was elevated compared to the control group. ZFPM2-AS1 is an oncogene and independent prognostic predictor of poor survival in NSCLCs, and its expression had a positive correlation with tumor size and lymph node metastasis in our clinical data. MTT, colony forming, wound healing, and transwell assays showed a positive correlation between ZFPM2-AS1 expression and the proliferation, migration, and invasion of NSCLC cells in the presence and absence of interferon- (IFN-γ). Using the TIMER database, we hypothesized that ZFPM2 was negatively correlated with ZFPM2-AS1 expression, as well as the immune infiltration levels in lung adenocarcinoma (LUAD). Finally, we found that ZFPM2-AS1 negatively regulated ZFPM2 expression, and had a positive correlation with PD-L1 expression through the JAK-STAT and AKT pathways.ConclusionOur study confirmed that ZFPM2-AS1 promotes the proliferation, migration, and invasion of NSCLC cells via the JAK-STAT and AKT pathways. Further research on the ZFPM2-AS1 pathway regulation mechanism is needed. creator: Xiwen Wang creator: Jun Tang creator: Jungang Zhao creator: Bin Lou creator: Li Li uri: https://doi.org/10.7717/peerj.10225 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: CRUNC: a cryopreservation method for unencapsulated gemmae of Marchantia polymorpha link: https://peerj.com/articles/10174 last-modified: 2020-10-23 description: Genetic modifications such as mutation and transformation are powerful tools to study the function of genes and proteins in the model liverwort Marchantia polymorpha, but maintaining the resulting germplasm requires a practical, reliable method. Cryopreservation methods allow researchers to maintain mutant and transgenic lines of M. polymorpha. To date, two methods have been developed for cryopreservation of M. polymorpha gemmae: in the first method, unencapsulated gemmae are stored in liquid nitrogen at −­196 °C, and in the second method, encapsulated gemmae are stored in liquid nitrogen at −­196 °C or a deep freezer at −80 °C. In the present study, we developed a simple method named CRUNC (cr yopreservation of un en c apsulated gemmae), which can be used to store unencapsulated, dried gemmae of wild-type and transgenic M. polymorpha lines in liquid nitrogen and in freezers at −80 °C and −20 °C. Using the CRUNC method, we observed a high recovery rate (as high as 100%) and successful long-term (5 months) storage of the gemmae. Therefore, the CRUNC method is practical for maintaining valuable M. polymorpha germplasm. creator: Hitomi Takahashi creator: Yutaka Kodama uri: https://doi.org/10.7717/peerj.10174 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Takahashi and Kodama title: Involvement of autophagy in realgar quantum dots (RQDs) inhibition of human endometrial cancer JEC cells link: https://peerj.com/articles/9754 last-modified: 2020-10-23 description: Realgar (As4S4) has been used in traditional Chinese medicines for treatment of malignancies. The poor solubility of As4S4 hampered its clinical applications. Realgar quantum dots (RQDs) were developed to overcome these problems. Previous studies revealed that the RQDs were effective against endometrial cancer JEC cells and hepatocarcinoma HepG2 cells via inducing apoptosis.Apoptosis and autophagy are important programmed cell death pathways leading to anticancer effects. This study further examined effects of RQDs on autophagy, focusing on the formation of the autophagosome in JEC cells. CCK8 assay was used to examine cell proliferation. Flow cytometry was used to analyze cell cycle. Transmission electron microscopy (TEM) was used to examine the autophagy, cells were transfected with pEGFP-C3-MAP1LC3B plasmid to examine effects of RQDs on autophagosome via confocal microscope. Autophagy-related proteins were examined by Western blot. RQDs exhibited cytotoxicity in JEC cells in a concentration- and time- dependent manner. RQDs induced G2 and S phase arrest in JEC cells. RQDs significantly induced autophagy, with the double-membrane and autophagosome-like structures by TEM. The diffused distribution of pEGFP-C3-MAP1LC3B green fluorescence were become the punctuate pattern fluorescence after treatment with RQDs in cells transfected with pEGFP-C3-MAP1LC3B plasmid RQDs increased the expression of autophagyregulatory proteins LC3 I/II, Beclin-1, p62 and Atg12 in a concentration-dependent manner, similar to autophagy induced by serum starvation, except for p62, as induction of p62 is a characteristic of arsenic compounds. Taken together, the present study clearly demonstrated that RQDs can induce autophagy in JEC cells as one of mechanisms of anticancer effects, and indicated that RQDs may be developed as an autophagy inducer. creator: Zhengyun Liu creator: Ke Xu creator: Yan Xu creator: Wanling Zhang creator: Nian Jiang creator: Shengyu Wang creator: Guo Luo creator: Jie Liu creator: Jinzhu Wu creator: Huan Wang uri: https://doi.org/10.7717/peerj.9754 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liu et al. title: Molecular Detection Mapping and Analysis Platform for R (MDMAPR) facilitating the standardization, analysis, visualization, and sharing of qPCR data and metadata link: https://peerj.com/articles/9974 last-modified: 2020-10-23 description: Quantitative polymerase chain reaction (qPCR) has been used as a standard molecular detection tool in many scientific fields. Unfortunately, there is no standard method for managing published qPCR data, and those currently used generally focus on only managing raw fluorescence data. However, associated with qPCR experiments are extensive sample and assay metadata, often under-examined and under-reported. Here, we present the Molecular Detection Mapping and Analysis Platform for R (MDMAPR), an open-source and fully scalable informatics tool for researchers to merge raw qPCR fluorescence data with associated metadata into a standard format, while geospatially visualizing the distribution of the data and relative intensity of the qPCR results. The advance of this approach is in the ability to use MDMAPR to store varied qPCR data. This includes pathogen and environmental qPCR species detection studies ideally suited to geographical visualization. However, it also goes beyond these and can be utilized with other qPCR data including gene expression studies, quantification studies used in identifying health dangers associated with food and water bacteria, and the identification of unknown samples. In addition, MDMAPR’s novel centralized management and geospatial visualization of qPCR data can further enable cross-discipline large-scale qPCR data standardization and accessibility to support research spanning multiple fields of science and qPCR applications. creator: Jiaojia Yu creator: Robert G. Young creator: Lorna E. Deeth creator: Robert H. Hanner uri: https://doi.org/10.7717/peerj.9974 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yu et al. title: Genome-wide identification of Hsp70/110 genes in rainbow trout and their regulated expression in response to heat stress link: https://peerj.com/articles/10022 last-modified: 2020-10-23 description: Heat shock proteins (Hsps) play an important role in many biological processes. However, as a typical cold water fish, the systematic identification of Hsp70/110 gene family of rainbow trout (Oncorhynchus mykiss) has not been reported, and the role of Hsp70/110 gene in the evolution of rainbow trout has not been described systematically. In this study, bioinformatics methods were used to analyze the Hsp70/110 gene family of rainbow trout. A total of 16 hsp70/110 genes were identified and classified into ten subgroups. The 16 Hsp70/110 genes were all distributed on chromosomes 2, 4, 8 and 13. The molecular weight is ranged from 78.93 to 91.39 kD. Gene structure and motif composition are relatively conserved in each subgroup. According to RNA-seq analysis of rainbow trout liver and head kidney, a total of four out of 16 genes were significantly upregulated in liver under heat stress, and a total of seven out of 16 genes were significantly upregulated in head kidney. RT-qPCR was carried out on these gene, and the result were consistent with those of RNA-seq. The significantly regulated expressions of Hsp70/110 genes under heat stress indicats that Hsp70/110 genes are involved in heat stress response in rainbow trout. This systematic analysis provided valuable information about the diverse roles of Hsp70/110 in the evolution of teleost, which will contribute to the functional characterization of Hsp70/110 genes in further research. creator: Fang Ma creator: Lintong Luo uri: https://doi.org/10.7717/peerj.10022 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Ma and Luo title: On the genus Mesopontonia Bruce, 1967 (Crustacea: Decapoda: Palaemonidae) in Korea, with the description of a new species link: https://peerj.com/articles/10190 last-modified: 2020-10-23 description: Mesopontonia verrucimanus and Mesopontonia kimwoni sp. nov. are recorded from high-latitude temperate waters in Munseom Islet, Jejudo Island, Republic of Korea, with both species collected on gorgonians and sponges by trimix diving between 50 and 75 m depth. Mesopontonia kimwoni sp. nov. is morphologically allied to M. brevicarpus, but can be distinguished by the cutting edges of the fingers of the first chela being entire. Significant morphological variation in the rostrum as well as the second pereiopods is documented in M. verrucimanus, although cytochrome c oxidase subunit I (COI) barcode analysis proves this to be infra-specific variation. A key to species of the genus Mesopontonia is provided. creator: Jin-Ho Park creator: Sammy De Grave creator: Taeseo Park uri: https://doi.org/10.7717/peerj.10190 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Park et al. title: The past century of coral bleaching in the Saudi Arabian central Red Sea link: https://peerj.com/articles/10200 last-modified: 2020-10-23 description: Accurate knowledge of the spatial and temporal patterns of coral bleaching is essential both for understanding how coral reef ecosystems are changing today and forecasting their future states. Yet, in many regions of the world, the history of bleaching is poorly known, especially prior to the late 20th century. Here, I use the information preserved within skeleton cores of long-lived Porites corals to reconstruct the past century of bleaching events in the Saudi Arabian central Red Sea. In these cores, skeletal “stress bands”—indicative of past bleaching—captured known bleaching events that occurred in 1998 and 2010, but also revealed evidence of previously unknown bleaching events in 1931, 1978, and 1982. However, these earlier events affected a significantly lesser proportion of corals than 1998 and 2010. Therefore, coral bleaching may have occurred in the central Red Sea earlier than previously recognized, but the frequency and severity of bleaching events since 1998 on nearshore reefs is unprecedented over the past century. Conversely, corals living on mid- to outer-shelf reefs have not been equally susceptible to bleaching as their nearshore counterparts, which was evident in that stress bands were five times more prevalent nearshore. Whether this pattern of susceptible nearshore reefs and resistant outer-shelf reefs continues in the future remains a key question in forecasting coral reef futures in this region. creator: Thomas M. DeCarlo uri: https://doi.org/10.7717/peerj.10200 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 DeCarlo title: Flower structure and development in Vietnamese Sciaphila (Triuridaceae: Pandanales): refined knowledge of the morphological misfit family and implications for taxonomy link: https://peerj.com/articles/10205 last-modified: 2020-10-23 description: The monocot family Triuridaceae is a morphological misfit with respect to several traits of floral morphology, including the uniformly apocarpous polymerous gynoecium and the famous inside-out flowers of Lacandonia. Although Triuridaceae are crucially important for understanding the floral evolution of Pandanales and angiosperms in general, significant knowledge gaps exist which hamper adequate morphological analysis of flowers in this family. The scarcity of morphological data is also reflected in numerous taxonomic inconsistencies. Here we provide a comprehensive study of reproductive organs of four species of Sciaphila occurring in Vietnam (S. arfakiana, S. densiflora, S. nana and S. stellata) including the first investigation of early floral development and gynoecium phyllotaxis. Our observations are mainly based on SEM images. We confirm the perianth (studied in male flowers) to be two-whorled and report a rare sequence of initiation of perianth parts: the outer tepals show a late congenital fusion, as their free lobes appear before the common perianth tube, whereas the inner tepals show an early congenital fusion, with their free lobes initiating on the tube rim. We interpret the stamen appendages as basal adaxial outgrowths of the stamen filaments. We discuss the number of thecae and locules in anthers of Sciaphila, and conclude that 3- and 4-, but not 2-locular anthers are characteristic of this genus. We describe the carpels as consisting of both ascidiate and plicate zones, the former being extremely short and completely obscured by anthesis. The single ovule is attached in the cross-zone. The style is non-plicate. We analyze gynoecium phyllotaxis by estimating its contact parastichies, and by investigating the number and arrangement of the outermost carpels. The carpel arrangement in a given gynoecium is a result of the balance between whorled and irregular (but not spiral) phyllotaxis. We recognize the following figures of gynoecium merism: 6, 9, 10, 10.5, 11 and 12, with the prevalence of those divisible by three. We discuss our results in the light of general diversity of floral structure of monocots. Our data allow to clarify several issues in taxonomy of Asian Sciaphila and indicate directions of further studies. We report a significant range extension for S. densiflora, which is newly recorded for the flora of Vietnam. We describe for the first time staminodes in female flowers of this species. We reveal two distinct morphs of S. nana in Vietnam. We highlight the need of a thorough revision of S. secundiflora species complex in order to verify the species boundaries and, in particular, to test the identity of the Vietnamese S. stellata. creator: Maxim S. Nuraliev creator: Margarita V. Remizowa creator: Dmitry D. Sokoloff uri: https://doi.org/10.7717/peerj.10205 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Nuraliev et al. title: Computerized monitoring of COVID-19 trials, studies and registries in ClinicalTrials.gov registry link: https://peerj.com/articles/10261 last-modified: 2020-10-23 description: Clinical trial registries can provide important information about relevant studies for a given condition to other researchers and the public. We developed a computerized informatics based approach to provide an overview and analysis of COVID-19 studies registered on ClinicalTrials.gov registry. Using the perspective of analyzing active or completed COVID-19 studies, we identified 401 interventional clinical trials, 287 observational studies and 64 registries. We analyzed features of each study type separately such as location, design, interventions and update history. Our results show that the United States had the most COVID-19 interventional trials, France had the most COVID-19 observational studies and France and the United States tied for the most COVID-19 registries on ClinicalTrials.gov. The majority of studies in all three study types had a single study site. For update history “Study Status” is the most updated information and we found that studies located in Canada (2.70 updates per study) and the United States (1.76 updates per study) update their studies more often than studies in any other country. Using normalization and mapping techniques, we identified Hydroxychloroquine (92 studies) as the most common drug intervention, while convalescent plasma (20 studies) is the most common biological intervention. The primary purpose of most interventional trials is for treatment with 298 studies (74.3%). For COVID-19 registries we found the most common proposed follow-up time is 1 year (15 studies). Of specific importance and interest is COVID-19 vaccine trials, of which 12 were identified. Our informatics based approach allows for constant monitoring and updating as well as multiple applications to other conditions and interests. creator: Craig S. Mayer creator: Vojtech Huser uri: https://doi.org/10.7717/peerj.10261 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Mayer and Huser title: Gene flow relates to evolutionary divergence among populations at the range margin link: https://peerj.com/articles/10036 last-modified: 2020-10-22 description: BackgroundMorphological differentiation between populations resulting from local adaptations to environmental conditions is likely to be more pronounced in populations with increasing genetic isolation. In a previous study a positive clinal variation in body size was observed in isolated Roesel’s bush-cricket, Metrioptera roeselii, populations, but were absent from populations within a continuous distribution at the same latitudinal range. This observational study inferred that there was a phenotypic effect of gene flow on climate-induced selection in this species.MethodsTo disentangle genetic versus environmental drivers of population differences in morphology, we measured the size of four different body traits in wild-caught individuals from the two most distinct latitudinally-matched pairs of populations occurring at about 60°N latitude in northern Europe, characterised by either restricted or continuous gene flow, and corresponding individuals raised under laboratory conditions.ResultsIndividuals that originated from the genetically isolated populations were always bigger (femur, pronotum and genital appendages) when compared to individuals from latitudinally-matched areas characterised by continuous gene flow between populations. The magnitude of this effect was similar for wild-caught and laboratory-reared individuals. We found that previously observed size cline variation in both male and female crickets was likely to be the result of local genetic adaptation rather than phenotypic plasticity.ConclusionsThis strongly suggests that restricted gene flow is of major importance for frequencies of alleles that participate in climate-induced selection acting to favour larger phenotypes in isolated populations towards colder latitudes. creator: Peter Kaňuch creator: Berrit Kiehl creator: Anna Cassel-Lundhagen creator: Ane T. Laugen creator: Matthew Low creator: Åsa Berggren uri: https://doi.org/10.7717/peerj.10036 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Kaňuch et al. title: The role of geomorphic zonation in long-term changes in coral-community structure on a Caribbean fringing reef link: https://peerj.com/articles/10103 last-modified: 2020-10-22 description: Ecological processes on coral reefs commonly have limited spatial and temporal scales and may not be recorded in their long-term geological history. The widespread degradation of Caribbean coral reefs over the last 40 years therefore provides an opportunity to assess the impact of more significant ecological changes on the geological and geomorphic structure of reefs. Here, we document the changing ecology of communities in a coral reef seascape within the context of its geomorphic zonation. By comparing basic ecological indices between historical and modern data we show that in 35 years the reef-front zone was transformed from a complex coral assemblage with a three-dimensional structure, to a size-homogenized and flattened one that is quasi indistinguishable from the adjacent non-accretional coral-ground zone. Today coral assemblages at Punta Maroma are characterized by the dominance of opportunistic species which are either tolerant to adverse environmental conditions, including sedimentation, or are known to be the first scleractinian species to recruit on disturbed reefs, implying they reflect a post-hurricane stage of adjustment. Despite an increase in similarity in ecological indices, the reef-front and coral-ground geomorphic zones still retain significant differences in coral assemblages and benthic habitat and are not homogeneous. The partial convergence of coral assemblages certainly has important consequences for the ecology and geological viability of the reef and its role in coastal protection, but environmental physical drivers continue to exert a fundamental role in the character and zonation of benthic communities of this reef seascape. creator: Alexis Enrique Medina-Valmaseda creator: Rosa E. Rodríguez-Martínez creator: Lorenzo Alvarez-Filip creator: Eric Jordan-Dahlgren creator: Paul Blanchon uri: https://doi.org/10.7717/peerj.10103 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Medina-Valmaseda et al. title: Constitution of a comprehensive phytochemical profile and network pharmacology based investigation to decipher molecular mechanisms of Teucrium polium L. in the treatment of type 2 diabetes mellitus link: https://peerj.com/articles/10111 last-modified: 2020-10-22 description: BackgroundType 2 diabetes mellitus (T2DM) is a metabolic disease affecting a huge population worldwide. Teucrium polium L. has been used as a folk medicine for the treatment of T2DM in Anatolia, Turkey. The antihyperglycemic effect of the plant was reported previously. However, there was no detailed study on the underlying molecular mechanisms. In this study, we generated a research plan to clarify the active constituents of the extract and uncover the molecular mechanisms using network pharmacology analysis.MethodsFor this purpose, we composed a dataset of 126 compounds for the phytochemical profile of the aerial parts of T. polium. Drug-likeness of the compounds was evaluated, and 52 compounds were selected for further investigation. A total of 252 T2DM related targets hit by selected compounds were subjected to DAVID database.ResultsThe KEGG pathway analysis showed enrichment for the TNF signaling pathway, insulin resistance, the HIF-1 signaling pathway, apoptosis, the PI3K-AKT signaling pathway, the FOXO signaling pathway, the insulin signaling pathway, and type 2 diabetes mellitus which are related to T2DM . AKT1, IL6, STAT3, TP53, INS, and VEGFA were found to be key targets in protein-protein interaction. Besides these key targets, with this study the role of GSK3β, GLUT4, and PDX1 were also discussed through literature and considered as important targets in the antidiabetic effect of T. polium. Various compounds of T. polium were shown to interact with the key targets activating PI3K-AKT and insulin signaling pathways.ConclusionsAccording to these findings, mainly phenolic compounds were identified as the active components and IRS1/PI3K/AKT signaling and insulin resistance were identified as the main pathways regulated by T. polium. This study reveals the relationship of the compounds in T. polium with the targets of T2DM in human. Our findings suggested the use of T. polium as an effective herbal drug in the treatment of T2DM and provides new insights for further research on the antidiabetic effect of T. polium. creator: Vahap Murat Kutluay creator: Neziha Yagmur Diker uri: https://doi.org/10.7717/peerj.10111 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kutluay and Diker title: Pseudodiaptomus marinus Sato, 1913 in the Black Sea: morphology, genetic analysis, and variability in seasonal and interannual abundance link: https://peerj.com/articles/10153 last-modified: 2020-10-22 description: Calanoid copepod Peudodiaptomus marinus Sato, 1913 was first recorded in Sevastopol Bay in the northern Black Sea in September 2016. We performed regular observations of this new invasive species between October 2016 and December 2018. We conducted bi-weekly plankton sampling at three stations located within or adjacent to Sevastopol Bay. This is the first paper to combine (i) a detailed morphological study, (ii) molecular genetic analysis, and (iii) an investigation of P. marinus seasonal dynamics and interannual abundance variability in the coastal Black Sea. Our morphological research showed similarities between Pseudodiaptomus specimens and existing P. marinus illustrations and descriptions. Our morphological analysis results were confirmed using molecular genetic studies. Based on the genetic variability of ITS2 and cytb, we found that all investigated specimens from Sevastopol Bay belonged to P. marinus. Investigations of P. marinus seasonal and interannual abundance variability showed the same seasonal patterns throughout the studied period, with a higher seasonal abundance from October to November and one pronounced density peak in autumn. The highest abundances (2,000 ind m–3 at the mouth of the bay and more than 5,000 ind m–3at its centre) were recorded in November 2018. In the samples, we found adults of both sexes, including ovigerous females, copepodites I–V, and nauplii, suggesting that the species reproduce in Sevastopol Bay. Our research indicated that P. marinus is a new non-indigenous species (NIS) in the Black Sea, and we will discuss a possible vector of its introduction into this basin. creator: Alexandra Gubanova creator: Inna Drapun creator: Oksana Garbazey creator: Olga Krivenko creator: Ekaterina Vodiasova uri: https://doi.org/10.7717/peerj.10153 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Gubanova et al. title: Using ForeStereo and LIDAR data to assess fire and canopy structure-related risks in relict Abies pinsapo Boiss. forests link: https://peerj.com/articles/10158 last-modified: 2020-10-22 description: In this study we combine information from aerial LIDAR and hemispherical images taken in the field with ForeStereo—a forest inventory device—to assess the vulnerability and to design conservation strategies for endangered Mediterranean fir forests based on the mapping of fire risk and canopy structure spatial variability. We focused on the largest continuous remnant population of the endangered tree species Abies pinsapo Boiss. spanning 252 ha in Sierra de las Nieves National Park (South Andalusia, Spain). We established 49 sampling plots over the study area. Stand structure variables were derived from ForeStereo device, a proximal sensing technology for tree diameter, height and crown dimensions and stand crown cover and basal area retrieval from stereoscopic hemispherical images photogrammetry. With this information, we developed regression models with airborne LIDAR data (spatial resolution of 0.5 points∙m−2). Thereafter, six fuel models were fitted to the plots according to the UCO40 classification criteria, and then the entire area was classified using the Nearest Neighbor algorithm on Sentinel imagery (overall accuracy of 0.56 and a KIA-Kappa Coefficient of 0.46). FlamMap software was used for fire simulation scenarios based on fuel models, stand structure, and terrain data. Besides the fire simulation, we analyzed canopy structure to assess the status and vulnerability of this fir population. The assessment shows a secondary growth forest that has an increasing presence of fuel models with the potential for high fire spread rate fire and burn probability. Our methodological approach has the potential to be integrated as a support tool for the adaptive management and conservation of A. pinsapo across its whole distribution area (<4,000 ha), as well as for other endangered circum-Mediterranean fir forests, as A. numidica de Lannoy and A. pinsapo marocana Trab. in North Africa. creator: Álvaro Cortés-Molino creator: Isabel Aulló-Maestro creator: Ismael Fernandez-Luque creator: Antonio Flores-Moya creator: José A. Carreira creator: A. Enrique Salvo uri: https://doi.org/10.7717/peerj.10158 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Cortés-Molino et al. title: Older adults at high risk of HIV infection in China: a systematic review and meta-analysis of observational studies link: https://peerj.com/articles/9731 last-modified: 2020-10-21 description: There is an increasing prevalence of human immunodeficiency virus (HIV) infection in older adults in China, but the findings across prevalence studies have been mixed. This is the first meta-analysis of the prevalence of HIV infection and its moderating factors in older adults in China. Two investigators systematically and independently searched both international (PubMed, PsycINFO, Web of Sciences and EMBASE) and Chinese (WanFang, CNKI, and CQVIP) databases. HIV infection rates in older adults were analyzed using the random-effects model. Altogether 46 studies were included in the analysis. The pooled prevalence of HIV infection in older adults was 2.1% (95% CI [1.9%–2.3%], I2 = 99.4%). Subgroup analyses revealed that men who have sex with men (MSM), hospital population samples, publications after 2014, studies conducted in the western region of China, and higher study quality were significantly associated with higher HIV infection rate. This meta-analysis found that the HIV infection prevalence in older adults is significantly higher than the general population in China. Attention should be given to this urgent public health issue, and effective HIV/AIDS preventive, screening and treatment measures are warranted in this population. PROSPERO: CRD42019124286. creator: Yuan Yuan Wang creator: Yuan Yang creator: Chang Chen creator: Ling Zhang creator: Chee H. Ng creator: Gabor S. Ungvari creator: Xiaohua Douglas Zhang creator: Yu-Tao Xiang uri: https://doi.org/10.7717/peerj.9731 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: De novo species identification using 16S rRNA gene nanopore sequencing link: https://peerj.com/articles/10029 last-modified: 2020-10-21 description: Nanopore sequencing is rapidly becoming more popular for use in various microbiota-based applications. Major limitations of current approaches are that they do not enable de novo species identification and that they cannot be used to verify species assignments. This severely limits applicability of the nanopore sequencing technology in taxonomic applications. Here, we demonstrate the possibility of de novo species identification and verification using hexamer frequencies in combination with k-means clustering for nanopore sequencing data. The approach was tested on the human infant gut microbiota of 3-month-old infants. Using the hexamer k-means approach we identified two new low abundant species associated with vaginal delivery. In addition, we confirmed both the vaginal delivery association for two previously identified species and the overall high levels of bifidobacteria. Taxonomic assignments were further verified by mock community analyses. Therefore, we believe our de novo species identification approach will have widespread application in analyzing microbial communities in the future. creator: Inga Leena Angell creator: Morten Nilsen creator: Karin C. Lødrup Carlsen creator: Kai-Håkon Carlsen creator: Gunilla Hedlin creator: Christine M. Jonassen creator: Benjamin Marsland creator: Björn Nordlund creator: Eva Maria Rehbinder creator: Carina Saunders creator: Håvard Ove Skjerven creator: Anne Cathrine Staff creator: Cilla Söderhäll creator: Riyas Vettukattil creator: Knut Rudi uri: https://doi.org/10.7717/peerj.10029 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Angell et al. title: Small-scale distribution of microbes and biogeochemistry in the Great Barrier Reef link: https://peerj.com/articles/10049 last-modified: 2020-10-21 description: Microbial communities distribute heterogeneously at small-scales (mm-cm) due to physical, chemical and biological processes. To understand microbial processes and functions it is necessary to appreciate microbes and matter at small scales, however, few studies have determined microbial, viral, and biogeochemical distribution over space and time at these scales. In this study, the small-scale spatial and temporal distribution of microbes (bacteria and chlorophyll a), viruses, dissolved inorganic nutrients and dissolved organic carbon were determined at five locations (spatial) along the Great Barrier Reef (Australia), and over 4 consecutive days (temporal) at a coastal location. Our results show that: (1) the parameters show high small-scale heterogeneity; (2) none of the parameters measured explained the bacterial abundance distributions at these scales spatially or temporally; (3) chemical (ammonium, nitrate/nitrite, phosphate, dissolved organic carbon, and total dissolved nitrogen) and biological (chl a, and bacterial and viral abundances) measurements did not reveal significant relationships at the small scale; and (4) statistically significant differences were found between sites/days for all parameter measured but without a clear pattern. creator: Cátia Carreira creator: Júlia Porto Silva Carvalho creator: Samantha Talbot creator: Isabel Pereira creator: Christian Lønborg uri: https://doi.org/10.7717/peerj.10049 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Carreira et al. title: Comparison of the diversity of cultured and total bacterial communities in marine sediment using culture-dependent and sequencing methods link: https://peerj.com/articles/10060 last-modified: 2020-10-21 description: Despite recent great advances in microbial culture, most microbes have not yet been cultured, and the impact of medium composition on the isolation of microbes from natural systems has not been elucidated. To optimize media for culturing marine microbes, microbial communities in three sediment samples were described using high-throughput sequencing (HTS) and culture-dependent techniques. HTS revealed communities dominated by Gammaproteobacteria, and culture-based methods revealed communities dominated by Actinobacteria. Among the total operational taxonomic units (OTUs) from the HTS dataset, 6% were recovered in the culture collection. Four potentially novel bacterial strains belonging to Oceaniovalibus, Psychrobacter and Salegentibacter were isolated. The combination of media cultured more taxa than any single medium. Nutrient-rich and single-carbon/nitrogen-source media supported the growth of relatively few taxa, and the quality of nitrogen strongly influenced the types of bacteria isolated. creator: Meng Wang creator: Samina Noor creator: Ran Huan creator: Congling Liu creator: JiaYi Li creator: Qingxin Shi creator: Yan-Jiao Zhang creator: Cuiling Wu creator: Hailun He uri: https://doi.org/10.7717/peerj.10060 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: Functional characterization of quorum sensing LuxR-type transcriptional regulator, EasR in Enterobacter asburiae strain L1 link: https://peerj.com/articles/10068 last-modified: 2020-10-21 description: Over the past decades, Enterobacter spp. have been identified as challenging and important pathogens. The emergence of multidrug-resistant Enterobacteria especially those that produce Klebsiella pneumoniae carbapenemase has been a very worrying health crisis. Although efforts have been made to unravel the complex mechanisms that contribute to the pathogenicity of different Enterobacter spp., there is very little information associated with AHL-type QS mechanism in Enterobacter spp. Signaling via N-acyl homoserine lactone (AHL) is the most common quorum sensing (QS) mechanism utilized by Proteobacteria. A typical AHL-based QS system involves two key players: a luxI gene homolog to synthesize AHLs and a luxR gene homolog, an AHL-dependent transcriptional regulator. These signaling molecules enable inter-species and intra-species interaction in response to external stimuli according to population density. In our recent study, we reported the genome of AHL-producing bacterium, Enterobacter asburiae strain L1. Whole genome sequencing and in silico analysis revealed the presence of a pair of luxI/R genes responsible for AHL-type QS, designated as easI/R, in strain L1. In a QS system, a LuxR transcriptional protein detects and responds to the concentration of a specific AHL controlling gene expression. In E. asburiae strain L1, EasR protein binds to its cognate AHLs, N-butanoyl homoserine lactone (C4-HSL) and N–hexanoyl homoserine lactone (C6-HSL), modulating the expression of targeted genes. In this current work, we have cloned the 693 bp luxR homolog of strain L1 for further characterization. The functionality and specificity of EasR protein in response to different AHL signaling molecules to activate gene transcription were tested and validated with β-galactosidase assays. Higher β-galactosidase activities were detected for cells harboring EasR, indicating EasR is a functional transcriptional regulator. This is the first report documenting the cloning and characterization of transcriptional regulator, luxR homolog of E. asburiae. creator: Yin Yin Lau creator: Kah Yan How creator: Wai-Fong Yin creator: Kok-Gan Chan uri: https://doi.org/10.7717/peerj.10068 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Lau et al. title: Morphological and molecular systematic review of Marphysa Quatrefages, 1865 (Annelida: Eunicidae) species from South Africa link: https://peerj.com/articles/10076 last-modified: 2020-10-21 description: A vast polychaete fauna is hidden behind complexes of cryptic and pseudo-cryptic species, which has greatly hindered our understanding of species diversity in several regions worldwide. Among the eunicids, Marphysa sanguineaMontagu, 1813 is a typical example, recorded in three oceans and with various species considered its junior synonyms. In South Africa, specimens previously misidentified as M. sanguinea are now known as Marphysa elityeniLewis & Karageorgopoulos, 2008. Of the six Marphysa Quatrefages, 1865a species recorded from the same region, three have their distributions restricted to South Africa while the others are considered to have worldwide distributions. Here, we evaluated the taxonomic status of the indigenous M. elityeni and investigated the presence of the widespread species Marphysa macintoshiCrossland, 1903 and Marphysa depressaSchmarda, 1861 in South Africa using morphological and molecular data. Our results reveal that M. elityeni is a junior synonym of Marphysa haemasoma, a species previously described from South Africa which is herein reinstated as a valid species. Both M. macintoshi and M. depressa are not present in South Africa and their status as being distributed worldwide deserves further investigation. Marphysa durbanensisDay, 1934 and the new species described here, M. sherlockae n. sp., had been misidentified as M. macintoshi and M. depressa respectively. Thus, the number of Marphysa species with distributions restricted to South Africa increased from three to five. This study reiterates the importance of implementing an integrated taxonomic framework to unravel local biodiversity. creator: Jyothi Kara creator: Isabel C. Molina-Acevedo creator: Joana Zanol creator: Carol Simon creator: Izwandy Idris uri: https://doi.org/10.7717/peerj.10076 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kara et al. title: Vegetation drives the structure of active microbial communities on an acidogenic mine tailings deposit link: https://peerj.com/articles/10109 last-modified: 2020-10-21 description: Plant-microbe associations are increasingly recognized as an inextricable part of plant biology and biogeochemistry. Microbes play an essential role in the survival and development of plants, allowing them to thrive in diverse environments. The composition of the rhizosphere soil microbial communities is largely influenced by edaphic conditions and plant species. In order to decipher how environmental conditions on a mine site can influence the dynamics of microbial communities, we characterized the rhizosphere soil microbial communities associated with paper birch, speckled alder, and spruce that had naturally colonized an acidogenic mine tailings deposit containing heavy metals. The study site, which had been largely undisturbed for five decades, had highly variable vegetation density; with some areas remaining almost barren, and others having a few stands or large thickets of mature trees. Using Illumina sequencing and ordination analyses (redundancy analysis and principal coordinate analysis), our study showed that soil bacterial and fungal community structures correlated mainly with vegetation density, and plant species. Tailings without any vegetation were the most different in bacterial community structure, compared to all other areas on the mine site, as well as an adjacent natural forest (comparison plot). The bacterial genera Acidiferrobacter and Leptospirillum were more abundant in tailings without vegetation than in any of the other sites, while Bradyrhizobium sp. were more abundant in areas of the tailings deposit having higher vegetation density. Frankia sp. is equally represented in each of the vegetation densities and Pseudomonas sp. present a greater relative abundance in boreal forest. Furthermore, alder rhizosphere showed a greater relative abundance of Bradyrhizobium sp. (in comparison with birch and spruce) as well as Haliangium sp. (in comparison with birch). In contrast, fungal community structures were similar across the tailings deposit regardless of vegetation density, showing a greater relative abundance of Hypocrea sp. Tailings deposit fungal communities were distinct from those found in boreal forest soils. Alder rhizosphere had greater relative abundances of Hypocrea sp. and Thelephora sp., while birch rhizosphere were more often associated with Mollisia sp. Our results indicate that, with increasing vegetation density on the mine site, the bacterial communities associated with the individual deciduous or coniferous species studied were increasingly similar to the bacterial communities found in the adjacent forest. In order to properly assess and restore disturbed sites, it is important to characterize and understand the plant-microbe associations that occur since they likely improve plant fitness in these harsh environments. creator: Vanessa Gagnon creator: Michaël Rodrigue-Morin creator: Julien Tremblay creator: Jessica Wasserscheid creator: Julie Champagne creator: Jean-Philippe Bellenger creator: Charles W. Greer creator: Sébastien Roy uri: https://doi.org/10.7717/peerj.10109 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Gagnon et al. title: Immunoglobulin superfamily member 10 is a novel prognostic biomarker for breast cancer link: https://peerj.com/articles/10128 last-modified: 2020-10-21 description: BackgroundImmunoglobulin superfamily member 10 (IGSF10) is a member of the immunoglobulin superfamily that is expressed at high levels in both the gallbladder and ovary. Currently, the role and possible mechanism of IGSF10 in breast cancer remain unclear.MethodBy applying real-time quantitative polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC), the expression of IGSF10 in breast cancer cells and tissues was detected. We collected the clinical information from 700 patients with breast cancer in The Cancer Genome Atlas (TCGA), and analyzed the relationship between IGSF10 expression and the clinicopathological features and survival outcomes of these patients. The potential mechanisms and pathways associated with IGSF10 in breast cancer were explored by performing a gene set enrichment analysis (GSEA).ResultsAccording to TCGA data, qRT-PCR and IHC experiments, levels of the IGSF10 mRNA and protein were significantly decreased in breast cancer tissues. IGSF10 expression was significantly correlated with age, tumor size, and tumor stage. Moreover, shorter overall survival (OS) and relapse-free survival (RFS) correlated with lower IGSF10 expression, according to the survival analysis. The multivariate analysis identified that IGSF10 as an independent prognostic factor for the OS (hazard ratio (HR) = 1.793, 95% confidence interval (CI) [1.141–2.815], P = 0.011) and RFS (HR = 2.298, 95% CI [1.317–4.010], P = 0.003) of patients with breast cancer. Based on the GSEA, IGSF10 was involved in DNA repair, cell cycle, and glycolysis. IGSF10 was also associated with the PI3K/Akt/mTOR and mTORC1 signaling pathways.ConclusionsThis study revealed a clear relationship between IGSF10 expression and the tumorigenesis of breast cancer for the first time. Therefore, further studies are needed to understand the mechanism of IGSF10 in breast cancer. creator: Mengxue Wang creator: Meng Dai creator: Yu-shen Wu creator: Ziying Yi creator: Yunhai Li creator: Guosheng Ren uri: https://doi.org/10.7717/peerj.10128 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: Genomics versus mtDNA for resolving stock structure in the silky shark (Carcharhinus falciformis) link: https://peerj.com/articles/10186 last-modified: 2020-10-21 description: Conservation genetic approaches for elasmobranchs have focused on regions of the mitochondrial genome or a handful of nuclear microsatellites. High-throughput sequencing offers a powerful alternative for examining population structure using many loci distributed across the nuclear and mitochondrial genomes. These single nucleotide polymorphisms are expected to provide finer scale and more accurate population level data; however, there have been few genomic studies applied to elasmobranch species. The desire to apply next-generation sequencing approaches is often tempered by the costs, which can be offset by pooling specimens prior to sequencing (pool-seq). In this study, we assess the utility of pool-seq by applying this method to the same individual silky sharks, Carcharhinus falciformis, previously surveyed with the mtDNA control region in the Atlantic and Indian Oceans. Pool-seq methods were able to recover the entire mitochondrial genome as well as thousands of nuclear markers. This volume of sequence data enabled the detection of population structure between regions of the Atlantic Ocean populations, undetected in the previous study (inter-Atlantic mitochondrial SNPs FST values comparison ranging from 0.029 to 0.135 and nuclear SNPs from 0.015 to 0.025). Our results reinforce the conclusion that sampling the mitochondrial control region alone may fail to detect fine-scale population structure, and additional sampling across the genome may increase resolution for some species. Additionally, this study shows that the costs of analyzing 4,988 loci using pool-seq methods are equivalent to the standard Sanger-sequenced markers and become less expensive when large numbers of individuals (>300) are analyzed. creator: Derek W. Kraft creator: Emily E. Conklin creator: Evan W. Barba creator: Melanie Hutchinson creator: Robert J. Toonen creator: Zac H. Forsman creator: Brian W. Bowen uri: https://doi.org/10.7717/peerj.10186 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Kraft et al. title: DNA barcoding for identification of anuran species in the central region of South America link: https://peerj.com/articles/10189 last-modified: 2020-10-21 description: The use of COI barcodes for specimen identification and species discovery has been a useful molecular approach for the study of Anura. Here, we establish a comprehensive amphibian barcode reference database in a central area of South America, in particular for specimens collected in Mato Grosso do Sul state (Brazil), and to evaluate the applicability of the COI gene for species-level identification. Both distance- and tree-based methods were applied for assessing species boundaries and the accuracy of specimen identification was evaluated. A total of 204 mitochondrial COI barcode sequences were evaluated from 22 genera and 59 species (19 newly barcoded species). Our results indicate that morphological and molecular identifications converge for most species, however, some species may present cryptic species due to high intraspecific variation, and there is a high efficiency of specimen identification. Thus, we show that COI sequencing can be used to identify anuran species present in this region. creator: Ricardo Koroiva creator: Luís Reginaldo Ribeiro Rodrigues creator: Diego José Santana uri: https://doi.org/10.7717/peerj.10189 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Koroiva et al. title: Structure and functions of Yellow-breasted Boubou (Laniarius atroflavus) solos and duets link: https://peerj.com/articles/10214 last-modified: 2020-10-21 description: BackgroundBirds have extremely well-developed acoustic communication and have become popular in bioacoustics research. The majority of studies on bird song have been conducted in the temperate zones where usually males of birds sing to attract females and defend territories. In over 360 bird species mostly inhabiting the tropics both males and females sing together in duets. Avian duets are usually formed when a male and female coordinate their songs. We focused on a species with relatively weakly coordinated duets, with male solo as the prevailing vocalisation type.MethodsInstead of analysing a set of recordings spread over a long time, we analysed whole day microphone-array recordings of the Yellow-breasted Boubou (Laniarius atroflavus), a species endemic to West African montane rainforests. We described the structure of the solo and duet vocalisations and temporal characteristics of daily activity based on 5,934 vocal bouts of 18 focal pairs and their neighbours.ResultsBirds had small, sex specific repertoires. All males shared three types of loud whistles functioning as song type repertoires in both solos and duets. Females vocalised with five types of harsh, atonal notes with a more variable and usually lower amplitude. Three of them were produced both as solos and in duets, while two seem to function as alarm and excitement calls given almost exclusively as a solo. Solos were the most common vocalisation mode (75.4%), with males being more vocally active than females. Duets accounted for 24.6% of all vocalisations and in most cases were initiated by males (81%). The majority of duets were simple (85.1%) consisting of a single male and female song type, but altogether 38 unique duet combinations were described. Males usually initiated singing at dawn and for this used one particular song type more often than expected by chance. Male solo and duet activities peaked around dawn, while female solos were produced evenly throughout the day.DiscussionYellow-breasted Boubou is a duetting species in which males are much more vocal than females and duetting is not a dominating type of vocal activity. Duet structure, context and timing of daily production support the joint resource defence hypothesis and mate guarding/prevention hypotheses, however maintaining pair contact also seems to be important. This study provides for the first time the basic quantitative data describing calls, solos and duet songs in the Yellow-breasted Boubou. creator: Amie Wheeldon creator: Paweł Szymański creator: Michał Budka creator: Tomasz S. Osiejuk uri: https://doi.org/10.7717/peerj.10214 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wheeldon et al. title: DNMSO; an ontology for representing de novo sequencing results from Tandem-MS data link: https://peerj.com/articles/10216 last-modified: 2020-10-21 description: For the identification and sequencing of proteins, mass spectrometry (MS) has become the tool of choice and, as such, drives proteomics. MS/MS spectra need to be assigned a peptide sequence for which two strategies exist. Either database search or de novo sequencing can be employed to establish peptide spectrum matches. For database search, mzIdentML is the current community standard for data representation. There is no community standard for representing de novo sequencing results, but we previously proposed the de novo markup language (DNML). At the moment, each de novo sequencing solution uses different data representation, complicating downstream data integration, which is crucial since ensemble predictions may be more useful than predictions of a single tool. We here propose the de novo MS Ontology (DNMSO), which can, for example, provide many-to-many mappings between spectra and peptide predictions. Additionally, an application programming interface (API) that supports any file operation necessary for de novo sequencing from spectra input to reading, writing, creating, of the DNMSO format, as well as conversion from many other file formats, has been implemented. This API removes all overhead from the production of de novo sequencing tools and allows developers to concentrate on algorithm development completely. We make the API and formal descriptions of the format freely available at https://github.com/savastakan/dnmso. creator: Savaş Takan creator: Jens Allmer uri: https://doi.org/10.7717/peerj.10216 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Takan and Allmer title: High microphone signal-to-noise ratio enhances acoustic sampling of wildlife link: https://peerj.com/articles/9955 last-modified: 2020-10-20 description: BackgroundAutomated sound recorders are a popular sampling tool in ecology. However, the microphones themselves received little attention so far, and specifications that determine the recordings’ sound quality are seldom mentioned. Here, we demonstrate the importance of microphone signal-to-noise ratio for sampling sonant animals.MethodsWe tested 12 different microphone models in the field and measured their signal-to-noise ratios and detection ranges. We also measured the vocalisation activity of birds and bats that they recorded, the bird species richness, the bat call types richness, as well as the performance of automated detection of bird and bat calls. We tested the relationship of each one of these measures with signal-to-noise ratio in statistical models.ResultsMicrophone signal-to-noise ratio positively affects the sound detection space areas, which increased by a factor of 1.7 for audible sound, and 10 for ultrasound, from the lowest to the highest signal-to-noise ratio microphone. Consequently, the sampled vocalisation activity increased by a factor of 1.6 for birds, and 9.7 for bats. Correspondingly, the species pool of birds and bats could not be completely detected by the microphones with lower signal-to-noise ratio. The performance of automated detection of bird and bat calls, as measured by its precision and recall, increased significantly with microphone signal-to-noise ratio.DiscussionMicrophone signal-to-noise ratio is a crucial characteristic of a sound recording system, positively affecting the acoustic sampling performance of birds and bats. It should be maximised by choosing appropriate microphones, and be quantified independently, especially in the ultrasound range. creator: Kevin F.A. Darras creator: Franziska Deppe creator: Yvonne Fabian creator: Agus P. Kartono creator: Andres Angulo creator: Bjørn Kolbrek creator: Yeni A. Mulyani creator: Dewi M. Prawiradilaga uri: https://doi.org/10.7717/peerj.9955 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Darras et al. title: Insect pollination is important in a smallholder bean farming system link: https://peerj.com/articles/10102 last-modified: 2020-10-20 description: BackgroundMany crops are dependent on pollination by insects. Habitat management in agricultural landscapes can support pollinator services and even augment crop production. Common bean (Phaseolus vulgaris L.) is an important legume for the livelihoods of smallholder farmers in many low-income countries, particularly so in East Africa. While this crop is autogamous, it is frequently visited by pollinating insects that could improve yields. However, the value of pollination services to common beans (Kariasii) yield is not known.MethodsWe carried out pollinator-exclusion experiments to determine the contribution of insect pollinators to bean yields. We also carried out a fluorescent-dye experiment to evaluate the role of field margins as refuge for flower-visitors.ResultsSignificantly higher yields, based on pods per plant and seeds per pod, were recorded from open-pollinated and hand-pollinated flowers compared to plants from which pollinators had been excluded indicating that flower visitors contribute significantly to bean yields. Similarly, open and hand-pollinated plants recorded the highest mean seed weight. Extrapolation of yield data to field scale indicated a potential increase per hectare from 681 kg in self-pollinated beans to 1,478 kg in open-pollinated beans indicating that flower visitors contributed significantly to crop yield of beans. Our marking study indicated that flower-visiting insects including bees, flies and lepidopterans moved from the field margin flowers into the bean crop. Overall, these results show that insect pollinators are important for optimising bean yields and an important food security consideration on smallholder farms. Field margin vegetation also provides habitat for flower-visiting insects that pollinate beans. Hence, non-crop habitats merit further research focusing on establishing which field margin species are most important and their capacity to support other ecosystem services such as natural pest regulation or even pests. creator: Filemon Elisante creator: Patrick Ndakidemi creator: Sarah E.J. Arnold creator: Steven R. Belmain creator: Geoff M. Gurr creator: Iain Darbyshire creator: Gang Xie creator: Philip C. Stevenson uri: https://doi.org/10.7717/peerj.10102 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Elisante et al. title: Isolation and evolutionary analyses of porcine epidemic diarrhea virus in Asia link: https://peerj.com/articles/10114 last-modified: 2020-10-20 description: Porcine epidemic diarrhea virus (PEDV) is a leading cause of diarrhea in pigs worldwide. Virus isolation and genetic evolutionary analysis allow investigations into the prevalence of epidemic strains and provide data for the clinical diagnosis and vaccine development. In this study, we investigated the genetic characteristics of PEDV circulation in Asia through virus isolation and comparative genomics analysis. APEDV strain designated HB2018 was isolated from a pig in a farm experiencing a diarrhea outbreak. The complete genome sequence of HB2018 was 28,138 bp in length. Phylogenetic analysis of HB2018 and 207 PEDVs in Asia showed that most PEDV strains circulating in Asia after 2010 belong to genotype GII, particularly GII-a. The PEDV vaccine strain CV777 belonged to GI, and thus, unmatched genotypes between CV777 and GII-a variants might partially explain incomplete protection by the CV777-derived vaccine against PEDV variants in China. In addition, we found the S protein of variant strains contained numerous mutations compared to the S protein of CV777, and these mutations occurred in the N-terminal domain of the S protein. These mutations may influence the antigenicity, pathogenicity, and neutralization properties of the variant strains. creator: Wan Liang creator: Danna Zhou creator: Chao Geng creator: Keli Yang creator: Zhengying Duan creator: Rui Guo creator: Wei Liu creator: Fangyan Yuan creator: Zewen Liu creator: Ting Gao creator: Ling Zhao creator: Dongwan Yoo creator: Yongxiang Tian uri: https://doi.org/10.7717/peerj.10114 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liang et al. title: Visual duration bisection in profoundly deaf individuals link: https://peerj.com/articles/10133 last-modified: 2020-10-20 description: BackgroundPrevious research has been designed to study the effect of hearing loss on supra-second duration estimation in the visual channel and position effect of visual abilities among deaf populations. The current study aimed to investigate the sub-second duration perception of different visual fields in profoundly deaf individuals.MethodsA total of 16 profoundly deaf undergraduates and 16 hearing undergraduates completed a visual duration bisection task in which participants made judgments about whether a series of probe durations that were linearly spaced from 200 ms to 800 ms at 100 ms intervals were more similar to a standard short duration (200 ms) or a standard long duration (800 ms). The probe stimuli were presented in the center, left, or right of the screen. A repeated measure analysis of variance (ANOVA) with a between-participants factor of group and a within-participants factor of position, and a one-sample t-test were conducted.ResultsThe Weber ratio (WR) values of deaf participants were significantly higher than those of hearing participants, regardless of the presented positions of the visual stimulus. The bisection point (BP) value of deaf participants was significantly lower than 500 ms (average mean of 200/800 ms) and the BP value of hearing participants did not significantly differ from 500 ms, although the overall difference of BP values between the deaf group and hearing group did not reach significance. For deaf participants, the BP value in the center condition was significantly lower than 500 ms; however, the difference between the BP value in the left condition and 500 ms did not reach significance, indicating that their duration discrimination accuracy in the left visual field was better than that in the center visual field.ConclusionsHearing loss impaired visual sub-second duration perception, and deaf individuals showed a left visual field advantage of duration discrimination accuracy during the visual duration bisection task. creator: Feng Zhang creator: Kaige Jin creator: Sainan Zhang uri: https://doi.org/10.7717/peerj.10133 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zhang et al. title: Polarity and epithelial-mesenchymal transition of retinal pigment epithelial cells in proliferative vitreoretinopathy link: https://peerj.com/articles/10136 last-modified: 2020-10-20 description: Under physiological conditions, retinal pigment epithelium (RPE) is a cellular monolayer composed of mitotically quiescent cells. Tight junctions and adherens junctions maintain the polarity of RPE cells, and are required for cellular functions. In proliferative vitreoretinopathy (PVR), upon retinal tear, RPE cells lose cell-cell contact, undergo epithelial-mesenchymal transition (EMT), and ultimately transform into myofibroblasts, leading to the formation of fibrocellular membranes on both surfaces of the detached retina and on the posterior hyaloids, which causes tractional retinal detachment. In PVR, RPE cells are crucial contributors, and multiple signaling pathways, including the SMAD-dependent pathway, Rho pathway, MAPK pathways, Jagged/Notch pathway, and the Wnt/β-catenin pathway are activated. These pathways mediate the EMT of RPE cells, which play a key role in the pathogenesis of PVR. This review summarizes the current body of knowledge on the polarized phenotype of RPE, the role of cell-cell contact, and the molecular mechanisms underlying the RPE EMT in PVR, emphasizing key insights into potential approaches to prevent PVR. creator: Hui Zou creator: Chenli Shan creator: Linlin Ma creator: Jia Liu creator: Ning Yang creator: Jinsong Zhao uri: https://doi.org/10.7717/peerj.10136 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zou et al. title: Mining the potential prognostic value of synaptosomal-associated protein 25 (SNAP25) in colon cancer based on stromal-immune score link: https://peerj.com/articles/10142 last-modified: 2020-10-20 description: BackgroundColon cancer is one of the deadliest tumors worldwide. Stromal cells and immune cells play important roles in cancer biology and microenvironment across different types of cancer. This study aimed to identify the prognostic value of stromal/immune cell-associated genes for colon cancer in The Cancer Genome Atlas (TCGA) database using bioinformatic technology.MethodsThe gene expression data and corresponding clinical information of colon cancer were downloaded from TCGA database. Stromal and immune scores were estimated based on the ESTIMATE algorithm. Sanger software was used to identify the differentially expressed genes (DEGs) and prognostic DEGs based on stromal and immune scores. External validation of prognostic biomarkers was conducted in Gene Expression Omnibus (GEO) database. Gene ontology (GO) analysis, pathway enrichment analysis, and gene set enrichment analysis (GSEA) were used for functional analysis. STRING and Cytoscape were used to assess the protein-protein interaction (PPI) network and screen hub genes. Quantitative real-time PCR (qRT-PCR) was used to validate the expression of hub genes in clinical tissues. Synaptosomal-associated protein 25 (SNAP25) was selected for analyzing its correlations with tumor-immune system in the TISIDB database.ResultsWorse overall survivals of colon cancer patients were found in high stromal score group (2963 vs. 1930 days, log-rank test P = 0.038) and high immune score group (2894 vs. 2230 days, log-rank test P = 0.076). 563 up-regulated and 9 down-regulated genes were identified as stromal-immune score-related DEGs. 70 up-regulated DEGs associated with poor outcomes were identified by COX proportional hazard regression model, and 15 hub genes were selected later. Then, we verified aquaporin 4 (AQP4) and SNAP25 as prognostic biomarkers in GEO database. qRT-PCR results revealed that AQP4 and SNAP25 were significantly elevated in colon cancer tissues compared with adjacent normal tissues (P = 0.003, 0.001). GSEA and TISIDB suggested that SNAP25 involved in cancer-related signaling pathway, immunity and metabolism progresses.ConclusionSNAP25 is a microenvironment-related and immune-related gene that can predict poor outcomes in colon cancer. creator: Jinyan Zou creator: Darong Duan creator: Changfa Yu creator: Jie Pan creator: Jinwei Xia creator: Zaixing Yang creator: Shasha Cai uri: https://doi.org/10.7717/peerj.10142 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zou et al. title: Craniofacial ontogeny in Tylosaurinae link: https://peerj.com/articles/10145 last-modified: 2020-10-20 description: Mosasaurs were large, globally distributed aquatic lizards that lived during the Late Cretaceous. Despite numerous specimens of varying maturity, a detailed growth series has not been proposed for any mosasaur taxon. Two taxa—Tylosaurus proriger and T. kansasensis/nepaeolicus—have robust fossil records with specimens spanning a wide range of sizes and are thus ideal for studying mosasaur ontogeny. Tylosaurus is a genus of particularly large mosasaurs with long, edentulous anterior extensions of the premaxilla and dentary that lived in Europe and North America during the Late Cretaceous. An analysis of growth in Tylosaurus provides an opportunity to test hypotheses of the synonymy of T. kansasensis with T. nepaeolicus, sexual dimorphism, anagenesis, and heterochrony. Fifty-nine hypothetical growth characters were identified, including size-dependent, size-independent, and phylogenetic characters, and quantitative cladistic analysis was used to recover growth series for the two taxa. The results supported the synonymy of T. kansasensis with T. nepaeolicus and that T. kansasensis represent juveniles of T. nepaeolicus. A Spearman rank-order correlation test resulted in a significant correlation between two measures of size (total skull length and quadrate height) and maturity. Eleven growth changes were shared across both species, neither of the ontogram topologies showed evidence of skeletal sexual dimorphism, and a previous hypothesis of paedomorphy in T. proriger was not rejected. Finally, a novel hypothesis of anagenesis in Western Interior Seaway Tylosaurus species, driven by peramorphy, is proposed here. creator: Amelia R. Zietlow uri: https://doi.org/10.7717/peerj.10145 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zietlow title: miR-29a-3p directly targets Smad nuclear interacting protein 1 and inhibits the migration and proliferation of cervical cancer HeLa cells link: https://peerj.com/articles/10148 last-modified: 2020-10-20 description: Smad nuclear interacting protein 1 (SNIP1) is a nuclear protein and involved in essential biological processes. MicroRNAs are effective regulators of tumorigenesis and cancer progression via targeting multiple genes. In present study, we aimed to investigate the function of SNIP1 and identify novel miRNA-SNIP1 axis in the development of cervical cancer. The results showed for the first time that silencing of the SNIP1 gene inhibited the migration and proliferation in HeLa cells significantly. Bioinformatics analysis and dual luciferase reporter assay demonstrated that miR-29a-3p could target 3′ UTR of SNIP1 directly. The mRNA and protein expression levels of SNIP1 were negative regulated by miR-29a-3p according to the RT-qPCR and Western blot analysis, respectively. Furthermore, functional studies showed that over-expression of miR-29a-3p restrained HeLa cells migration and proliferation, and the mRNA expression of SNIP1 downstream genes (HSP27, c-Myc, and cyclin D1) were down-regulated by miR-29a-3p. Together, we concluded that miR-29a-3p suppressed the migration and proliferation in HeLa cells by directly targeting SNIP1. The newly identified miR-29a-3p/SNIP1 axis could provide new insight into the development of cervical cancer. creator: Ying Chen creator: Weiji Zhang creator: Lijun Yan creator: Peng Zheng creator: Jin Li uri: https://doi.org/10.7717/peerj.10148 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: Spatiotemporal accumulation of fatal pharyngeal entrapment of flatfish in harbour porpoises (Phocoena phocoena) in the German North Sea link: https://peerj.com/articles/10160 last-modified: 2020-10-20 description: The evolution of a permanent separation of the upper respiratory and digestive tract is one of the adaptions cetaceans evolved for their aquatic life. Generally, it prevents odontocetes from choking on either saltwater or foreign bodies during ingestion under water. Nevertheless, several sporadic single case reports from different parts of the world show that this separation can be reversed especially by overly large items of prey. This incident can have a fatal outcome for the odontocetes. The German federal state of Schleswig-Holstein has a year-round, permanent and systematic stranding network that retrieves stranded marine mammals from its shorelines and constantly enables post-mortem examinations. In 2016, with nine affected animals, a high incidence of fatal pharyngeal entrapment of flatfish in harbour porpoises (Phocoena phocoena) occurred during spring and early summer on the German North Sea island of Sylt. All flatfish were identified as common sole (Solea solea). A retrospective post-mortem data analysis over a 30-year period from the North and Baltic Sea revealed similar yearly and seasonally case accumulations on the same island in the 1990s as well as several single case events over the whole timespan. All cases except one were caused by flatfish. When flatfish speciation was performed, only common sole was identified. From 1990 to 2019, of all examined harbour porpoises, 0.3% (2/713) from the Baltic Sea and 5.5% (45/820) from the North Sea died due to fish entrapped in the pharynx. On the North Sea coast, the occurrence of fatal obstruction shows high yearly variations from 0 to 33.3%. Years that stand out are especially 1990 to 1992, 1995, as well as 2016. The majority of all cases generally occurred between April and July, indicating also a seasonality of cases. This study evaluates the occurrence of fatal pharyngeal entrapment of fish in two geographically separated harbour porpoise populations. Additionally, common sole is clearly identified as a potentially risky item of prey for these small odontocetes. creator: Stephanie Gross creator: Marco Roller creator: Holger Haslob creator: Miguel Grilo creator: Jan Lakemeyer creator: Anja Reckendorf creator: Peter Wohlsein creator: Ursula Siebert uri: https://doi.org/10.7717/peerj.10160 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Gross et al. title: Isolation and sequence-based characterization of a koala symbiont: Lonepinella koalarum link: https://peerj.com/articles/10177 last-modified: 2020-10-20 description: Koalas (Phascolarctos cinereus) are highly specialized herbivorous marsupials that feed almost exclusively on Eucalyptus leaves, which are known to contain varying concentrations of many different toxic chemical compounds. The literature suggests that Lonepinella koalarum, a bacterium in the Pasteurellaceae family, can break down some of these toxic chemical compounds. Furthermore, in a previous study, we identified L. koalarum as the most predictive taxon of koala survival during antibiotic treatment. Therefore, we believe that this bacterium may be important for koala health. Here, we isolated a strain of L. koalarum from a healthy koala female and sequenced its genome using a combination of short-read and long-read sequencing. We placed the genome assembly into a phylogenetic tree based on 120 genome markers using the Genome Taxonomy Database (GTDB), which currently does not include any L. koalarum assemblies. Our genome assembly fell in the middle of a group of Haemophilus, Pasteurella and Basfia species. According to average nucleotide identity and a 16S rRNA gene tree, the closest relative of our isolate is L. koalarum strain Y17189. Then, we annotated the gene sequences and compared them to 55 closely related, publicly available genomes. Several genes that are known to be involved in carbohydrate metabolism could exclusively be found in L. koalarum relative to the other taxa in the pangenome, including glycoside hydrolase families GH2, GH31, GH32, GH43 and GH77. Among the predicted genes of L. koalarum were 79 candidates putatively involved in the degradation of plant secondary metabolites. Additionally, several genes coding for amino acid variants were found that had been shown to confer antibiotic resistance in other bacterial species against pulvomycin, beta-lactam antibiotics and the antibiotic efflux pump KpnH. In summary, this genetic characterization allows us to build hypotheses to explore the potentially beneficial role that L. koalarum might play in the koala intestinal microbiome. Characterizing and understanding beneficial symbionts at the whole genome level is important for the development of anti- and probiotic treatments for koalas, a highly threatened species due to habitat loss, wildfires, and high prevalence of Chlamydia infections. creator: Katherine E. Dahlhausen creator: Guillaume Jospin creator: David A. Coil creator: Jonathan A. Eisen creator: Laetitia G.E. Wilkins uri: https://doi.org/10.7717/peerj.10177 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Dahlhausen et al. title: Human papillomavirus infection and follow-up on positive results in 7222 female samples obtained from 2016 to 2019 in Hefei, China link: https://peerj.com/articles/10179 last-modified: 2020-10-20 description: BackgroundHuman papillomavirus (HPV) infection rates in women vary regionally. This study analyzed HPV infection in women of different age groups in Hefei, China, performed follow-up on positive cases, and discussed infection prognoses.MethodsSamples (7,222) of exfoliated cervical cells were collected in Hefei and tested with an HPV assay kit against 27 HPV genotypes. Statistical software was used to analyze the data.ResultsThe total positive rate of infection was 17.13% (1,068 women), and the 51–60-year age group had the highest HPV infection rate (19.82%). There were statistically significant differences between rates in the 21–30 and 31–40 (P = 0.002), 21–30 and 41–50 (P = 0.0003), 21–30 and 51–60 (P = 0.00003), and 51–60 and >60 age groups (P = 0.046). High-risk infection (15.67%) and single infection (13.01%) were the main types of HPV infection. The dominant genotypes of high-risk infection were HPV 52 (2.42%), HPV 16 (2.01%), HPV 53 (1.43%), HPV 58 (1.32%) and HPV 66 (1.01%). We conducted follow-up on cases in 69 of 94 women who had a history of 1–4 years of positive infection, and in 18 (seven treated, 11 untreated) patients, infection status turned negative (26.09%). Seventeen of the fifty-one women whose infections did not turn negative received treatment. Persistent infection was predominantly observed in high-risk genotypes (56 of 69).ConclusionsThe results recommend that women in Hefei improve health awareness and receive a 9-valent vaccine. Additionally, women with persistent infections should consult a gynecologist to prevent cervical lesions. creator: Liduo Peng creator: Liping Yin creator: Yaqian Dai creator: Yuanjing Peng creator: Yuanhong Xu creator: Huaqing Hu creator: Jinping Qiao uri: https://doi.org/10.7717/peerj.10179 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Peng et al. title: What families really think about the quality of early intervention centers: a perspective from mixed methods link: https://peerj.com/articles/10193 last-modified: 2020-10-20 description: BackgroundFamilies are a fundamental aspect in the current perspective of Early Intervention, and knowing their opinion with quantitative and qualitative research is necessary for its improvement. The objective of this research was to evaluate the quality of the service perceived in Early Intervention Centers and its relationship with satisfaction and future intention, as well as to identify factors that are associated with the perception of users.MethodsA measurement model of 50 items and an open question to gather qualitative information was used in a sample of 233 participants. A confirmatory factor analysis and a regression analysis were conducted. Regarding the qualitative data, the information was subjected to a thematic content analysis in order to delve into the perception of the participants.ResultsThe model showed a satisfactory fit and the regression analysis indicated that treatment rooms (β =  − 0.28) and adaptation of activities (β = 0.27) have greater weight with respect to satisfaction, whereas for future intention, the factors of greater weight were adaptation of activities (β = 0.23) and location (β = 0.20). The qualitative analysis showed three themes: facilitators, barriers and suggestions for improvement. Within facilitators, the participants were satisfied with the Early Intervention professionals, and they made improvement suggestions for the detected barriers to improve the facilities and the follow-up of the child.ConclusionsThe study offers a wide perspective of the perception of the service with an active participation of families in the treatment within the Early Intervention service. This will allow professionals in Early Intervention, service providers and researchers to consider the families as intervention agents capable of providing their opinion and making decisions, and not only as passive elements. creator: Rita Pilar Romero-Galisteo creator: Pablo Gálvez Ruiz creator: Angel Blanco Villaseñor creator: Maria Rodríguez-Bailón creator: Manuel González-Sánchez uri: https://doi.org/10.7717/peerj.10193 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Romero-Galisteo et al. title: Construction and analysis of macrophage infiltration related circRNA-miRNA-mRNA regulatory networks in hepatocellular carcinoma link: https://peerj.com/articles/10198 last-modified: 2020-10-20 description: BackgroundMacrophage play a crucial role in regulating tumor progression. This study intended to investigate the circular RNA (circRNA) regulatory network associated with macrophage infiltration in hepatocellular carcinoma (HCC).MethodsThe immune cell fractions of HCC from The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium were calculated by Estimation of the Proportion of Immune and Cancer cells algorithm. The differentially expressed mRNAs (DEmRNAs), microRNAs (DEmiRNAs) and circRNAs (DEcircRNAs) were identified from HCC and adjacent non-tumor cases of TCGA or Gene Expression Omnibus database. The DEmRNAs related to macrophage were selected by weighted gene co-expression network analysis and then utilized to generate the circRNA-miRNA-mRNA network. A hub circRNA regulatory network was established based on the co-expressed DEmiRNAs and DEmRNAs owning contrary correlation with the clinical characteristics, survival and macrophage infiltration level. A gene signature based on the DEmRNAs in hub network was also generated for further evaluation. The circRNA binding bite for miRNA was detected by luciferase assay.ResultsHigh macrophage fraction predicted good survival for HCC. A circRNA-miRNA-mRNA network was constructed by 27 macrophage related DEmRNAs, 21 DEmiRNAs, and 15 DEcircRNAs. Among this network, the expression of hsa-miR-139-5p was negatively correlated with CDCA8, KPNA2, PRC1 or TOP2A. Hsa-miR-139-5p low or targeted DEmRNA high expression was associated with low macrophage infiltration, high grade, advanced stage and poor prognosis of HCC. Additionally, the risk score generated by 4-DEmRNA signature could reflect the macrophage infiltration status and function as an independent prognostic factor for HCC. Finally, hsa_circ_0007456 acting on hsa-miR-139-5p related network was viewed as the hub circRNA regulatory network. Taken together, some circRNA regulatory networks may be associated with macrophage infiltration, which provides clues for mechanism study and therapeutic strategies of HCC. creator: Yuhan Chen creator: Yalin Li creator: Guanglei Zheng creator: Peitao Zhou uri: https://doi.org/10.7717/peerj.10198 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: Soil organic matter dynamics in semiarid agroecosystems transitioning to dryland link: https://peerj.com/articles/10199 last-modified: 2020-10-20 description: Recent interest in improving soil health and agricultural sustainability recognizes the value of soil organic carbon (SOC) sequestration and nutrient cycling. The main goal of this study was to evaluate the response of various SOC and nitrogen (N) components in semiarid cropping systems transitioning from limited-irrigation to dryland and a restored grassland in the Southern High Plains of USA. Cropping systems evaluated include dryland winter wheat (Triticum aestivum L.)–sorghum (Sorghum bicolor L.)–fallow with conventional tillage (DLCTF) and no-tillage (DLNTF), limited-irrigation winter wheat–sorghum–fallow with no-tillage and cover cropping (LINTC) and no-tillage fallow (LINTF), and an undisturbed grassland (NG). Soil samples were collected from 0–15 cm and 15–30 cm depths and analyzed for SOC, total N, inorganic N, and soil microbial biomass carbon (SMBC) contents. The CO2 and N2O release during a eight-weeks long laboratory incubation were also analyzed. Results show 14% and 13% reduction in SOC and total N from 0–30 cm depth with the transition from limited-irrigation to dryland cropping systems while 51% more SOC and 41% more total N with the transition to grassland. The SMBC was 42% less in dryland cropping systems and 100% more in NG than the limited-irrigation cropping systems. However, the grassland was N limited, with 93% less inorganic N in NG compared to only 11% less in dryland cropping systems than in limited-irrigation cropping systems. The microbial respiration measured as CO2-C was highest in NG, followed by limited-irrigation and dryland cropping systems. The N2O-N release showed the lowest rate of N loss from dryland cropping systems, followed by NG and limited-irrigation cropping systems. This study demonstrated loss of SOC and N in agroecosystems transitioned to dryland crop-fallow systems, with greater magnitude of change observed in the biologically active fraction of soil organic matter. Grassland restoration could be an important strategy to increase SOC and nutrients in hot, dry, semiarid agroecosystems transitioning to dryland. creator: Rajan Ghimire creator: Babu Ram Khanal uri: https://doi.org/10.7717/peerj.10199 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Ghimire and Khanal title: Structure and expression analysis of seven salt-related ERF genes of Populus link: https://peerj.com/articles/10206 last-modified: 2020-10-20 description: Ethylene response factors (ERFs) are plant-specific transcription factors (TFs) that play important roles in plant growth and stress defense and have received a great amount of attention in recent years. In this study, seven ERF genes related to abiotic stress tolerance and response were identified in plants of the Populus genus. Systematic bioinformatics, including sequence phylogeny, genome organisation, gene structure, gene ontology (GO) annotation, etc. were detected. Expression-pattern of these seven ERF genes were analyzed using RT-qPCR and cross validated using RNA-Seq. Data from a phylogenetic tree and multiple alignment of protein sequences indicated that these seven ERF TFs belong to three subfamilies and contain AP2, YRG, and RAYD conserved domains, which may interact with downstream target genes to regulate the plant stress response. An analysis of the structure and promoter region of these seven ERF genes showed that they have multiple stress-related motifs and cis-elements, which may play roles in the plant stress-tolerance process through a transcriptional regulation mechanism; moreover, the cellular_component and molecular_function terms associated with these ERFs determined by GO annotation supported this hypothesis. In addition, the spatio-temporal expression pattern of these seven ERFs, as detected using RT-qPCR and RNA-seq, suggested that they play a critical role in mediating the salt response and tolerance in a dynamic and tissue-specific manner. The results of this study provide a solid basis to explore the functions of the stress-related ERF TFs in Populus abiotic stress tolerance and development process. creator: Juanjuan Huang creator: Shengji Wang creator: Xingdou Wang creator: Yan Fan creator: Youzhi Han uri: https://doi.org/10.7717/peerj.10206 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Huang et al. title: Feroxichthys yunnanensis gen. et sp. nov. (Colobodontidae, Neopterygii), a large durophagous predator from the Middle Triassic (Anisian) Luoping Biota, eastern Yunnan, China link: https://peerj.com/articles/10229 last-modified: 2020-10-20 description: Neopterygii is a large group of ray-finned fishes which underwent a rapid radiation in the Middle Triassic. Until recently, 11 stem neopterygians have been recovered from the early Middle Triassic Luoping Biota in eastern Yunnan, China, and they are small to medium-sized fishes. Here, I report the discovery of a new stem neopterygian, Feroxichthys yunnanensis gen. et sp. nov. from the Luoping Biota, which represents the first evidence of large-sized stem neopteygians in this biota with a total length of ~340 mm (290 mm in standard length). The skull of the new taxon is exceptionally well-preserved, showing some peculiar features rarely known in other stem neopterygians, for example fusion of paired premaxillae, fusion of lacrimal with maxilla, and a fused parieto-dermopterotic with a strong posterior process. Phylogenetic studies recover Feroxichthys as a basal colobodontid, and a revised diagnosis of this family is presented. The feeding apparatus indicates that Feroxichthys might have been predominantly durophagous, resembling other colobodontids. However, the anterior peg-like teeth in the jaws of Feroxichthys are much longer and stronger than other colobodontids, enabling a more powerful initial prey capture before food was passed posteriorly to molariform teeth for crushing in the oral cavity. As a mysterious large durophagous predator previously unknown from the Luoping Biota, the new finding is important not only for understanding the early diversification of neopterygians during this age but also for investigating the trophic structure in this marine ecosystem. creator: Guang-Hui Xu uri: https://doi.org/10.7717/peerj.10229 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Xu title: Spatial differentiation of gully clusters based on the regional scale: an example from northeastern China link: https://peerj.com/articles/9907 last-modified: 2020-10-19 description: Gully erosion represents a serious environmental threat around the world, but their spatial distribution law are unclear at the regional scale. To quantitatively characterize the spatial distribution model of gullies and determine the regularity of regional differentiation, this paper used spatial analyst and statistics method to study the spatial distribution of gullies in 34 sample areas of northeastern China based on interpretations of high-resolution remote sensing images. The results showed that the kernel density could quantitatively describe the continuous spatial clustering of gullies. Gullies in northeastern China had the characteristics of a spatially unbalanced distribution at the scale of the sample area. The average kernel density of the 34 sample areas (Moran’s I was 0.43, P¡0.01P < 0.01) also indicated clustering distribution at the regional scale. The horizontal clustering characteristics of gullies exhibited an azonal distribution of being low values in the middle plain and high values on the three mountainous areas. The average kernel density in the southeastern part of the study area was highest (maximum value of 2.38). In the vertical direction, gullies were relatively undeveloped in low- and high-altitude areas, while middle-altitude areas were beneficial to the development of gullies. The effect of height differences on gully development was more significant than altitude. As the height difference increased, gullies tended to be more clustered, which can be expressed by a power function. The results of this study will not only help to understand the regional differentiation characteristics of gullies but will also provide a scientific reference for the study of spatial distribution of gullies in future. creator: Ying Zhao creator: Bin Zhang creator: Wei Qin creator: Jun Luo creator: Hui Liu creator: Qingchun Deng creator: Wei Lv creator: Yuli He creator: Houling Wang uri: https://doi.org/10.7717/peerj.9907 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhao et al. title: Generation of human liver organoids from pluripotent stem cell-derived hepatic endoderms link: https://peerj.com/articles/9968 last-modified: 2020-10-19 description: BackgroundThe use of a personalized liver organoid derived from human-induced pluripotent stem cells (HuiPSCs) is advancing the use of in vitro disease models for the design of specific, effective therapies for individuals. Collecting patient peripheral blood cells for HuiPSC generation is preferable because it is less invasive; however, the capability of blood cell-derived HuiPSCs for hepatic differentiation and liver organoid formation remains uncertain. Moreover, the currently available methods for liver organoid formation require a multistep process of cell differentiation or a combination of hepatic endodermal, endothelial and mesenchymal cells, which is a major hurdle for the application of personalized liver organoids in high-throughput testing of drug toxicity and safety. To demonstrate the capability of blood cell-derived HuiPSCs for liver organoid formation without support from endothelial and mesenchymal cells.MethodsThe peripheral blood-derived HuiPSCs first differentiated into hepatic endoderm (HE) in two-dimensional (2D) culture on Matrigel-coated plates under hypoxia for 10 days. The HE was then collected and cultured in 3D culture using 50% Matrigel under ambient oxygen. The maturation of hepatocytes was further induced by adding hepatocyte growth medium containing HGF and oncostatin M on top of the 3D culture and incubating the culture for an additional 12–17 days. The function of the liver organoids was assessed using expression analysis of hepatocyte-specific gene and proteins. Albumin (ALB) synthesis, glycogen and lipid storage, and metabolism of indocyanine were evaluated. The spatial distribution of albumin was examined using immunofluorescence and confocal microscopy.ResultsCD34+ hematopoietic cell-derived HuiPSCs were capable of differentiating into definitive endoderm expressing SOX17 and FOXA2, hepatic endoderm expressing FOXA2, hepatoblasts expressing AFP and hepatocytes expressing ALB. On day 25 of the 2D culture, cells expressed SOX17, FOXA2, AFP and ALB, indicating the presence of cellular heterogeneity. In contrast, the hepatic endoderm spontaneously formed a spherical, hollow structure in a 3D culture of 50% Matrigel, whereas hepatoblasts and hepatocytes could not form. Microscopic observation showed a single layer of polygonal-shaped cells arranged in a 3D structure. The hepatic endoderm-derived organoid synthesis ALB at a higher level than the 2D culture but did not express definitive endoderm-specific SOX17, indicating the greater maturity of the hepatocytes in the liver organoids. Confocal microscopic images and quantitative ELISA confirmed albumin synthesis in the cytoplasm of the liver organoid and its secretion. Overall, 3D culture of the hepatic endoderm is a relatively fast, simple, and less laborious way to generate liver organoids from HuiPSCs that is more physiologically relevant than 2D culture. creator: Kasem Kulkeaw creator: Alisa Tubsuwan creator: Nongnat Tongkrajang creator: Narisara Whangviboonkij uri: https://doi.org/10.7717/peerj.9968 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kulkeaw et al. title: Using trace elements to identify the geographic origin of migratory bats link: https://peerj.com/articles/10082 last-modified: 2020-10-19 description: The expansion of the wind energy industry has had benefits in terms of increased renewable energy production but has also led to increased mortality of migratory bats due to interactions with wind turbines. A key question that could guide bat-related management activities is identifying the geographic origin of bats killed at wind-energy facilities. Generating this information requires developing new methods for identifying the geographic sources of individual bats. Here we explore the viability of assigning geographic origin using trace element analyses of fur to infer the summer molting location of eastern red bats (Lasiurus borealis). Our approach is based on the idea that the concentration of trace elements in bat fur is related through the food chain to the amount of trace elements present in the soil, which varies across large geographic scales. Specifically, we used inductively coupled plasma–mass spectrometry to determine the concentration of fourteen trace elements in fur of 126 known-origin eastern red bats to generate a basemap for assignment throughout the range of this species in eastern North America. We then compared this map to publicly available soil trace element concentrations for the U.S. and Canada, used a probabilistic framework to generate likelihood-of-origin maps for each bat, and assessed how well trace element profiles predicted the origins of these individuals. Overall, our results suggest that trace elements allow successful assignment of individual bats 80% of the time while reducing probable locations in half. Our study supports the use of trace elements to identify the geographic origin of eastern red and perhaps other migratory bats, particularly when combined with data from other biomarkers such as genetic and stable isotope data. creator: Jamin G. Wieringa creator: Juliet Nagel creator: David M. Nelson creator: Bryan C. Carstens creator: H. Lisle Gibbs uri: https://doi.org/10.7717/peerj.10082 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wieringa et al. title: Spatiotemporal expansion of human brucellosis in Shaanxi Province, Northwestern China and model for risk prediction link: https://peerj.com/articles/10113 last-modified: 2020-10-19 description: BackgroundHuman brucellosis imposes a heavy burden on the health and economy of endemic regions. Since 2011, China has reported at least 35,000 human brucellosis cases annually, with more than 90% of these cases reported in the northern. Given the alarmingly high incidence and variation in the geographical distribution of human brucellosis cases, there is an urgent need to decipher the causes of such variation in geographical distribution.MethodWe conducted a retrospective epidemiological study in Shaanxi Province from January 1, 2005 to December 31, 2018 to investigate the association between meteorological factors and transmission of human brucellosis according to differences in geographical distribution and seasonal fluctuation in northwestern China for the first time.ResultsHuman brucellosis cases were mainly distributed in the Shaanbei upland plateau before 2008 and then slowly extended towards the southern region with significant seasonal fluctuation. The results of quasi-Poisson generalized additive mixed model (GAMM) indicated that air temperature, sunshine duration, rainfall, relative humidity, and evaporation with maximum lag time within 7 months played crucial roles in the transmission of human brucellosis with seasonal fluctuation. Compared with the Shaanbei upland plateau, Guanzhong basin had more obvious fluctuations in the occurrence of human brucellosis due to changes in meteorological factors. Additionally, the established GAMM model showed high accuracy in predicting the occurrence of human brucellosis based on the meteorological factors.ConclusionThese findings may be used to predict the seasonal fluctuations of human brucellosis and to develop reliable and cost-effective prevention strategies in Shaanxi Province and other areas with similar environmental conditions. creator: Zurong Yang creator: Miaomiao Pang creator: Qingyang Zhou creator: Shuxuan Song creator: Weifeng Liang creator: Junjiang Chen creator: Tianci Guo creator: Zhongjun Shao creator: Kun Liu uri: https://doi.org/10.7717/peerj.10113 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yang et al. title: Chemical patterns of colony membership and mother-offspring similarity in Antarctic fur seals are reproducible link: https://peerj.com/articles/10131 last-modified: 2020-10-19 description: Replication studies are essential for evaluating the validity of previous research findings. However, it has proven challenging to reproduce the results of ecological and evolutionary studies, partly because of the complexity and lability of many of the phenomena being investigated, but also due to small sample sizes, low statistical power and publication bias. Additionally, replication is often considered too difficult in field settings where many factors are beyond the investigator’s control and where spatial and temporal dependencies may be strong. We investigated the feasibility of reproducing original research findings in the field of chemical ecology by performing an exact replication of a previous study of Antarctic fur seals (Arctocephalus gazella). In the original study, skin swabs from 41 mother-offspring pairs from two adjacent breeding colonies on Bird Island, South Georgia, were analyzed using gas chromatography-mass spectrometry. Seals from the two colonies differed significantly in their chemical fingerprints, suggesting that colony membership may be chemically encoded, and mothers were also chemically similar to their pups, hinting at the possible involvement of phenotype matching in mother-offspring recognition. In the current study, we generated and analyzed chemical data from a non-overlapping sample of 50 mother-offspring pairs from the same two colonies 5 years later. The original results were corroborated in both hypothesis testing and estimation contexts, with p-values remaining highly significant and effect sizes, standardized between studies by bootstrapping the chemical data over individuals, being of comparable magnitude. However, exact replication studies are only capable of showing whether a given effect can be replicated in a specific setting. We therefore investigated whether chemical signatures are colony-specific in general by expanding the geographic coverage of our study to include pups from a total of six colonies around Bird Island. We detected significant chemical differences in all but a handful of pairwise comparisons between colonies. This finding adds weight to our original conclusion that colony membership is chemically encoded, and suggests that chemical patterns of colony membership not only persist over time but can also be generalized over space. Our study systematically confirms and extends our previous findings, while also implying more broadly that spatial and temporal heterogeneity need not necessarily negate the reproduction and generalization of ecological research findings. creator: Jonas Tebbe creator: Emily Humble creator: Martin Adam Stoffel creator: Lisa Johanna Tewes creator: Caroline Müller creator: Jaume Forcada creator: Barbara Caspers creator: Joseph Ivan Hoffman uri: https://doi.org/10.7717/peerj.10131 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Tebbe et al. title: Photosynthetic performance and stevioside concentration are improved by the arbuscular mycorrhizal symbiosis in Stevia rebaudiana under different phosphate concentrations link: https://peerj.com/articles/10173 last-modified: 2020-10-19 description: In plants, phosphorus (P) uptake occurs via arbuscular mycorrhizal (AM) symbiosis and through plant roots. The phosphate concentration is known to affect colonization by AM fungi, and the effect depends on the plant species. Stevia rebaudiana plants are valuable sources of sweetener compounds called steviol glycosides (SGs), and the principal components of SGs are stevioside and rebaudioside A. However, a detailed analysis describing the effect of the phosphate concentration on the colonization of AM fungi in the roots and the relationship of these factors to the accumulation of SGs and photochemical performance has not been performed; such an analysis was the aim of this study. The results indicated that low phosphate concentrations (20 and 200 µM KH2PO4) induced a high percentage of colonization by Rhizophagus irregularis in the roots of S. rebaudiana, while high phosphate concentrations (500 and 1,000 µM KH2PO4) reduced colonization. The morphology of the colonization structure is a typical Arum-type mycorrhiza, and a mycorrhiza-specific phosphate transporter was identified. Colonization with low phosphate concentrations improved plant growth, chlorophyll and carotenoid concentration, and photochemical performance. The transcription of the genes that encode kaurene oxidase and glucosyltransferase (UGT74G1) was upregulated in colonized plants at 200 µM KH2PO4, which was consistent with the observed patterns of stevioside accumulation. In contrast, at 200 µM KH2PO4, the transcription of UGT76G1 and the accumulation of rebaudioside A were higher in noncolonized plants than in colonized plants. These results indicate that a low phosphate concentration improves mycorrhizal colonization and modulates the stevioside and rebaudioside A concentration by regulating the transcription of the genes that encode kaurene oxidase and glucosyltransferases, which are involved in stevioside and rebaudioside A synthesis in S. rebaudiana. creator: Luis G. Sarmiento-López creator: Melina López-Meyer creator: Gabriela Sepúlveda-Jiménez creator: Luis Cárdenas creator: Mario Rodríguez-Monroy uri: https://doi.org/10.7717/peerj.10173 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Sarmiento-López et al. title: Nursing home residents’ ADL status, institution-dwelling and association with outdoor activity: a cross-sectional study link: https://peerj.com/articles/10202 last-modified: 2020-10-19 description: IntroductionThe Norwegian regulations for nursing homes consider access to meaningful activities to be an indicator for the quality of nursing homes. Activities of daily living (ADL) provide important basic self-care skills for nursing home residents. Due to the physical changes caused by ageing and comorbidities, nursing home residents may experience functional decline over time, which may affect their ability to perform meaningful ADL, such as outdoor activity, which is considered a valuable and meaningful activity in Norwegian culture. This study aimed to investigate the association between ADL status, institution-dwelling and outdoor activity among nursing home residents.MethodsThis cross-sectional study included 784 residents aged >67 years living in 21 nursing homes in 15 Norwegian municipalities between November 2016 and May 2018. The Barthel Index was used to assess the nursing home residents’ ADL status. Other variables collected were age, gender, body weight and height, visits per month, institution, ward, and participation in weekly outdoor activities. Descriptive statistics were used to provide an overview of the residents’ characteristics. A Poisson regression model was used to test the association between the outdoor activity level as the dependent variable and ADL score, institution, and other control variables as independent variables.ResultsMore than half (57%) of the nursing home residents in this sample did not go outdoors. More than 50% of the residents had an ADL score <10, which indicates low performance status. Further, we found that residents’ ADL status, institution, ward, and number of visits had an impact on how often the residents went outdoors.DiscussionThe nursing home residents in this study rarely went outdoors, which is interesting because Norwegians appreciate this activity. Differences in the number of visits might explain why some residents went outdoors more often than other residents did. Our findings also highlight that the institutions impact the outdoor activity. How the institutions are organized and how important this activity is considered to be in the institutions determine how often the activity is performed.ConclusionThe low frequency of the outdoor activities might be explained by a low ADL score. More than 50% of the residents had an ADL score <10, which indicates low performance status. Despite regulations for nursing home quality in Norway, this result suggests that organizational differences matter, which is an important implication for further research, health policy and practice. creator: Anne Marie Sandvoll creator: Ellen Karine Grov creator: Morten Simonsen uri: https://doi.org/10.7717/peerj.10202 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Sandvoll et al. title: A study of soil seed banks across one complete chronosequence of secondary succession in a karst landscape link: https://peerj.com/articles/10226 last-modified: 2020-10-19 description: Anthropogenic disturbance and distinctive geochemistry have resulted in rocky desertification in many karst regions of the world. Seed banks are crucial to vegetation regeneration in degraded karst ecosystems characterized by a discontinuous distribution of soil and seasonal drought stress. However, the dynamics of seed banks across one complete series of secondary succession and the underlying mechanisms remain unclear. We selected eight typical stages during secondary succession, conducted aboveground vegetation survey and collected 960 soil samples in the Guiyang karst landscape of China. Seed density, species richness and plant life forms in seed banks were determined via the germination method. The results indicated that the seed density in seed banks before and after field seed germination was significantly different among most succession stages. Community succession had impacts on the seed density of seed banks before and after field seed germination. Seed density ranged from 1,042 seedlings.m−2 in evergreen broadleaf forests to 3,755 seedlings.m−2 in the herb community, which was a relatively high density. The seed density and similar species composition between the seed banks and vegetation declined with succession from early to later stages. Species richness in seed banks was highest in middle succession stages and increased with increasing species richness of aboveground vegetation. The species richness of the five life forms in the seed banks showed different variations across these succession stages. The conservation of diverse aboveground vegetation can maintain the diversity of seed banks for restoration. creator: Xiaole He creator: Li Yuan creator: Zhen Hong Wang creator: Zizong Zhou creator: Li Wan uri: https://doi.org/10.7717/peerj.10226 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 He et al. title: Unique myological changes associated with ossified fabellae: a femorofabellar ligament and systematic review of the double-headed popliteus link: https://peerj.com/articles/10028 last-modified: 2020-10-16 description: IntroductionThe fabella is a sesamoid bone embedded in the tendon of the lateral head of the gastrocnemius. It is the only bone in the human body to increase in prevalence in the last 100 years. As the fabella can serve as an origin/insertion for muscles, tendons, and/or ligaments (e.g., the oblique popliteal and fabellofibular ligaments), temporal changes in fabella prevalence could lead to temporal changes in “standard” knee anatomy. The aim of this study was to investigate unique myological changes to the posterolateral corner knee associated with ossified fabella presence and perform a systematic review to contextualize our results.MethodsThirty-three fresh frozen cadaveric knees were considered. As the knees were all used for previous experimentation, the knees were in variable levels of preservation. Those with adequate preservation were used to determine ossified fabella presence/absence. When ossified fabellae were present, unique myologies associated with the fabella were recorded. A systematic review was performed on the double-headed popliteus to investigate possible correlations between this anatomical variant and the fabella.ResultsOf the 33 knees, 30 preserved enough soft tissue to determine fabella presence/absence: 16/30 knees had fabellae (five cartilaginous and 11 ossified). Eight of the eleven knees with ossified fabellae retained enough soft tissue to investigate the posterolateral knee anatomy. Of these, 4/8 exhibited unique myological changes. One knee had a double-headed popliteus muscle where one head originated from the medial side of a large, bulbous fabella. A systematic review revealed double-headed popliteus muscles are rare, but individuals are 3.7 times more likely to have a fabella if they have a double-headed popliteus. Another knee had a large, thick ligament stretching from the lateral edge of the fabella to the inferoposterior edge of the lateral femoral epicondyle, deep to the lateral collateral ligament (LCL) and near the popliteal sulcus. We found no mention of such a ligament in the literature and refer to it here as the “femorofabellar ligament”. In all four knees, the plantaris and lateral gastrocnemius appeared to share a common tendinous origin, and the fabella was located at/near the junction of these muscles. In the case of the double-headed popliteus, the fabella clearly served as an origin for the plantaris.ConclusionsDespite being found in an average of 36.80% of human knees, most standard anatomical models fail to account for the fabella and/or the unique myological changes associated with fabella presence. Although our sample is small, these data highlight aspects of human biological variability generally not considered when creating generalized anatomical models. Further work is needed to identify additional changes associated with ossified fabellae and the functional consequences of omitting these changes from models. creator: Michael A. Berthaume creator: Spencer Barnes creator: Kiron K. Athwal creator: Lukas Willinger uri: https://doi.org/10.7717/peerj.10028 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Berthaume et al. title: A robust semi-supervised NMF model for single cell RNA-seq data link: https://peerj.com/articles/10091 last-modified: 2020-10-16 description: BackgroundSingle-cell RNA-sequencing (scRNA-seq) technology is a powerful tool to study organism from a single cell perspective and explore the heterogeneity between cells. Clustering is a fundamental step in scRNA-seq data analysis and it is the key to understand cell function and constitutes the basis of other advanced analysis. Nonnegative Matrix Factorization (NMF) has been widely used in clustering analysis of transcriptome data and achieved good performance. However, the existing NMF model is unsupervised and ignores known gene functions in the process of clustering. Knowledges of cell markers genes (genes that only express in specific cells) in human and model organisms have been accumulated a lot, such as the Molecular Signatures Database (MSigDB), which can be used as prior information in the clustering analysis of scRNA-seq data. Because the same kind of cells is likely to have similar biological functions and specific gene expression patterns, the marker genes of cells can be utilized as prior knowledge in the clustering analysis.MethodsWe propose a robust and semi-supervised NMF (rssNMF) model, which introduces a new variable to absorb noises of data and incorporates marker genes as prior information into a graph regularization term. We use rssNMF to solve the clustering problem of scRNA-seq data.ResultsTwelve scRNA-seq datasets with true labels are used to test the model performance and the results illustrate that our model outperforms original NMF and other common methods such as KMeans and Hierarchical Clustering. Biological significance analysis shows that rssNMF can identify key subclasses and latent biological processes. To our knowledge, this study is the first method that incorporates prior knowledge into the clustering analysis of scRNA-seq data. creator: Peng Wu creator: Mo An creator: Hai-Ren Zou creator: Cai-Ying Zhong creator: Wei Wang creator: Chang-Peng Wu uri: https://doi.org/10.7717/peerj.10091 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wu et al. title: Broad spectrum antimicrobial activities from spore-forming bacteria isolated from the Vietnam Sea link: https://peerj.com/articles/10117 last-modified: 2020-10-16 description: The widespread occurrence of pathogenic bacteria resistant to last-line antibiotics has resulted in significant challenges in human and veterinary medicine. There is an urgent need for new antimicrobial agents that can be used to control these life threating pathogens. We report the identification of antimicrobial activities, against a broad range of bacterial pathogens, from a collection of marine-derived spore-forming bacteria. Although marine environments have been previously investigated as sources of novel antibiotics, studies on such environments are still limited and there remain opportunities for further discoveries and this study has used resources derived from an under-exploited region, the Vietnam Sea. Antimicrobial activity was assessed against a panel of Gram-positive and Gram-negative bacteria, including several multi-drug resistant pathogens. From a total of 489 isolates, 16.4% had antimicrobial activity. Of 23 shortlisted isolates with the greatest antimicrobial activity, 22 were Bacillus spp. isolates and one was a Paenibacillus polymyxa isolate. Most of the antimicrobial compounds were sensitive to proteases, indicating that they were proteins rather than secondary metabolites. The study demonstrated that marine bacteria derived from the Vietnam Sea represent a rich resource, producing antimicrobial compounds with activity against a broad range of clinically relevant bacterial pathogens, including important antibiotic resistant pathogens. Several isolates were identified that have particularly broad range activities and produce antimicrobial compounds that may have value for future drug development. creator: Khanh Minh Chau creator: Dong Van Quyen creator: Joshua M. Fraser creator: Andrew T. Smith creator: Thi Thu Hao Van creator: Robert J. Moore uri: https://doi.org/10.7717/peerj.10117 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chau et al. title: High-throughput analysis of adaptation using barcoded strains of Saccharomyces cerevisiae link: https://peerj.com/articles/10118 last-modified: 2020-10-16 description: BackgroundExperimental evolution of microbes can be used to empirically address a wide range of questions about evolution and is increasingly employed to study complex phenomena ranging from genetic evolution to evolutionary rescue. Regardless of experimental aims, fitness assays are a central component of this type of research, and low-throughput often limits the scope and complexity of experimental evolution studies. We created an experimental evolution system in Saccharomyces cerevisiae that utilizes genetic barcoding to overcome this challenge.ResultsWe first confirm that barcode insertions do not alter fitness and that barcode sequencing can be used to efficiently detect fitness differences via pooled competition-based fitness assays. Next, we examine the effects of ploidy, chemical stress, and population bottleneck size on the evolutionary dynamics and fitness gains (adaptation) in a total of 76 experimentally evolving, asexual populations by conducting 1,216 fitness assays and analyzing 532 longitudinal-evolutionary samples collected from the evolving populations. In our analysis of these data we describe the strengths of this experimental evolution system and explore sources of error in our measurements of fitness and evolutionary dynamics.ConclusionsOur experimental treatments generated distinct fitness effects and evolutionary dynamics, respectively quantified via multiplexed fitness assays and barcode lineage tracking. These findings demonstrate the utility of this new resource for designing and improving high-throughput studies of experimental evolution. The approach described here provides a framework for future studies employing experimental designs that require high-throughput multiplexed fitness measurements. creator: Vincent J. Fasanello creator: Ping Liu creator: Carlos A. Botero creator: Justin C. Fay uri: https://doi.org/10.7717/peerj.10118 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Fasanello et al. title: Impact of COVID-19 outbreak on the mental health status of undergraduate medical students in a COVID-19 treating medical college: a prospective longitudinal study link: https://peerj.com/articles/10164 last-modified: 2020-10-16 description: BackgroundThe COVID-19 pandemic is found to affect the mental health of the population. Undergraduate medical students are especially prone to mental health disorders and hence could be more vulnerable to the impact of the pandemic.MethodsA prospective longitudinal study was conducted on 217 undergraduate medical students in a medical college at Chennai, India. Depression, anxiety, and stress levels were recorded using Depression Anxiety Stress Scale 21 Items (DASS21) before and during the COVID-19 outbreak in India in December 2019 and June 2020, respectively. In the follow-up survey, in addition to DASS21, the Pittsburgh Sleep Quality Index to assess sleep quality and a self-administered questionnaire to assess the impact of COVID-19 related stressors were used. The self-administered questionnaire assessed the status of COVID-19 testing, interactions with COVID-19 patients, self-perceived levels of concerns and worries related to academics (COVID-19-AA (academic apprehensions)) and those pertaining to the self and family/friends (COVID-19-GA (general apprehensions)). Cross-sectional and longitudinal comparison of overall scores of depression, anxiety, and stress and scores stratified by gender, year of study, place of residence and monthly family income were performed. Predictors for depression, anxiety, and stress during COVID-19 were investigated using adjusted binary logistic regression analysis and results were expressed as adjusted odds ratio with 95% confidence interval (CI). A P value < 0.05 was considered statistically significant.ResultsThe average scores of depression, anxiety, and stress during the baseline survey were 7.55 ± 7.86, 4.6 ± 6.19 and 7.31 ± 7.34 with the prevalence (95% Cl) of 33.2% [27–39.9%], 21.2% [16–27.2%] and 20.7% [15.5–26.7%]; in follow-up survey, the mean scores were 8.16 ± 8.9, 6.11 ± 7.13 and 9.31 ± 8.18 with the prevalence being 35.5% [29.1–42.2%], 33.2% [27–39.9%] and 24.9% [19.3–31.2%] for depression, anxiety, and stress respectively. There was a significant increase in both the prevalence and levels of anxiety and stress (P < 0.001), with depression remaining unchanged during COVID-19, irrespective of gender, year of study, place of residence and family’s monthly income. Poor sleep quality, higher levels of baseline depression, anxiety, and stress, higher COVID-19-GA, COVID-19 patients in family/friends and direct interactions with COVID-19 patients were found to be significant predictors of negative mental health in undergraduate medical students. COVID-19-AA was not significantly associated with depression, anxiety, and stress.ConclusionThe COVID-19 pandemic appears to negatively affect the mental health of the undergraduate medical students with the prevalence and levels of anxiety and stress being increased, and depression symptoms remaining unaltered. Addressing and mitigating the negative effect of COVID-19 on the mental health of this population is crucial. creator: Ilango Saraswathi creator: Jayakumar Saikarthik creator: K. Senthil Kumar creator: Kumar Madhan Srinivasan creator: M. Ardhanaari creator: Raghunath Gunapriya uri: https://doi.org/10.7717/peerj.10164 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Saraswathi et al. title: Chrysophyllum cainito stem bark extract induces apoptosis in Human hepatocarcinoma HepG2 cells through ROS-mediated mitochondrial pathway link: https://peerj.com/articles/10168 last-modified: 2020-10-16 description: Hepatocellular carcinoma is the most common type of primary liver cancer in humans. This study aimed to demonstrate anticancer properties of an aqueous extract from Chrysophyllum cainito stem bark (CE) and its underlying mechanisms. Our MTT assay results showed that CE significantly reduced human hepatocellular carcinoma (HepG2) cell viability with the IC50of 100 µg/mL, while human dermal primary fibroblast (HDFa) cells showed less susceptibility in every concentration tested. Determined by Annexin V staining, the proportion of apoptotic HepG2 cells increased in a dose-dependent fashion after 24 hour-exposure of CE. The results from Western blot analysis confirmed that CE reduced procaspase-3, suggesting apoptosis by activating caspase-3 cleavage. Using the DCFH-DA and DiOC6 fluorescent probes, it was found that CE significantly stimulated the generation of reactive oxygen species (ROS) and reduced mitochondrial membrane potential (Δψm), respectively. According to cell cycle analysis, CE (100 µg/mL) profoundly increased the percentage of cells in the sub-G1 phase, indicating cell apoptosis. These data suggest that CE induces apoptosis and cell death in human hepatocellular carcinoma via generation of intracellular ROS and disruption of Δψm. This is the first demonstration of the anticancer activity with proposed underlying mechanism of CE in liver cancer cells. creator: Hau V. Doan creator: Pishyaporn Sritangos creator: Roongtip Iyara creator: Nuannoi Chudapongse uri: https://doi.org/10.7717/peerj.10168 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Doan et al. title: Testing the short-and long-term effects of elevated prenatal exposure to different forms of thyroid hormones link: https://peerj.com/articles/10175 last-modified: 2020-10-16 description: Maternal thyroid hormones (THs) are known to be crucial in embryonic development in humans, but their influence on other, especially wild, animals remains poorly understood. So far, the studies that experimentally investigated the consequences of maternal THs focused on short-term effects, while early organisational effects with long-term consequences, as shown for other prenatal hormones, could also be expected. In this study, we aimed at investigating both the short- and long-term effects of prenatal THs in a bird species, the Japanese quail Coturnix japonica. We experimentally elevated yolk TH content (the prohormone T4, and its active metabolite T3, as well as a combination of both hormones). We analysed hatching success, embryonic development, offspring growth and oxidative stress as well as their potential organisational effects on reproduction, moult and oxidative stress in adulthood. We found that eggs injected with T4 had a higher hatching success compared with control eggs, suggesting conversion of T4 into T3 by the embryo. We detected no evidence for other short-term or long-term effects of yolk THs. These results suggest that yolk THs are important in the embryonic stage of precocial birds, but other short- and long-term consequences remain unclear. Research on maternal THs will greatly benefit from studies investigating how embryos use and respond to this maternal signalling. Long-term studies on prenatal THs in other taxa in the wild are needed for a better understanding of this hormone-mediated maternal pathway. creator: Tom Sarraude creator: Bin-Yan Hsu creator: Ton Groothuis creator: Suvi Ruuskanen uri: https://doi.org/10.7717/peerj.10175 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Sarraude et al. title: Positive selection within the genomes of SARS-CoV-2 and other Coronaviruses independent of impact on protein function link: https://peerj.com/articles/10234 last-modified: 2020-10-16 description: BackgroundThe emergence of a novel coronavirus (SARS-CoV-2) associated with severe acute respiratory disease (COVID-19) has prompted efforts to understand the genetic basis for its unique characteristics and its jump from non-primate hosts to humans. Tests for positive selection can identify apparently nonrandom patterns of mutation accumulation within genomes, highlighting regions where molecular function may have changed during the origin of a species. Several recent studies of the SARS-CoV-2 genome have identified signals of conservation and positive selection within the gene encoding Spike protein based on the ratio of synonymous to nonsynonymous substitution. Such tests cannot, however, detect changes in the function of RNA molecules.MethodsHere we apply a test for branch-specific oversubstitution of mutations within narrow windows of the genome without reference to the genetic code.ResultsWe recapitulate the finding that the gene encoding Spike protein has been a target of both purifying and positive selection. In addition, we find other likely targets of positive selection within the genome of SARS-CoV-2, specifically within the genes encoding Nsp4 and Nsp16. Homology-directed modeling indicates no change in either Nsp4 or Nsp16 protein structure relative to the most recent common ancestor. These SARS-CoV-2-specific mutations may affect molecular processes mediated by the positive or negative RNA molecules, including transcription, translation, RNA stability, and evasion of the host innate immune system. Our results highlight the importance of considering mutations in viral genomes not only from the perspective of their impact on protein structure, but also how they may impact other molecular processes critical to the viral life cycle. creator: Alejandro Berrio creator: Valerie Gartner creator: Gregory A. Wray uri: https://doi.org/10.7717/peerj.10234 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Berrio et al. title: The fungal community in non-rhizosphere soil of Panax ginseng are driven by different cultivation modes and increased cultivation periods link: https://peerj.com/articles/9930 last-modified: 2020-10-15 description: Continuous cropping obstacles severely hindered the sustained development of the ginseng industry. Among the obstacles, an imbalance of soil microbiome community was considered one of the major culprits. The fungal community is an essential part of the soil microbiome community. Extensive characterization of the fungal community composition and variation during ginseng cultivation will help us understand the mechanism underlying continuous cropping obstacles. By using a high-throughput amplicon sequencing method, the non-rhizospheric fungal community of farmland cultivated ginseng of 2 years old (C2) and 5 years old (C5), understory wild ginseng of 15 years old (W15) and 35 years old (W35), fallow fields which have been abandoned for 10 (F10) years were characterized. Farmland cultivated ginseng and understory wild ginseng harbored distinct non-rhizospheric fungal communities, and extension of cultivation periods enlarged the fungal community difference between two cultivation modes. Extended cultivation periods significantly decreased the OTU richness and PD whole tree indices, and OTU number and cultivation periods were negatively correlated. Extension of cultivation periods led to an increased abundance of pathotrophs. Still, the increased abundance of pathotrophs may not be the leading cause of severe continuous cropping obstacles in farmland cultivated ginseng. Compared with understory wild ginseng, farmland cultivated ginseng had a lower abundance of symbiotrophs and a higher abundance of saprotrophs. This changed symbiotrophs/saprotrophs ratio may have some correlation with the severe continuous cropping obstacles that occurred in farmland cultivated ginseng. Fallowing on the fungal community of the non-rhizosphere soil was generally opposite of that of extension of ginseng cultivation periods. The impacts of farmland cultivation on the fungal community of the non-rhizosphere soil can last for decades, even if the following is practiced. creator: Yu Bao creator: Bao Qi creator: Wei Huang creator: Bao Liu creator: Yu Li uri: https://doi.org/10.7717/peerj.9930 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Bao et al. title: Molecular epidemiology and clinical characteristics of herpangina children in Beijing, China: a surveillance study link: https://peerj.com/articles/9991 last-modified: 2020-10-15 description: BackgroundHerpangina is a highly infectious disease, which is usually prevalent in preschool children.MethodsThis study analyzed the clinical and pathogenic characteristics of herpangina children to demonstrate the epidemiology of herpangina. Clinical manifestations, laboratory indicators and pharyngeal swabs were collected from children with herpangina who were monitored by Tongzhou Center for Disease Control and Prevention in Beijing, 2008. Utilizing pharyngeal swabs, virus extraction and amplification were performed for nucleotide identification and sequencing. The phylogenetic analysis was conducted based on all sequences amplified in this study and strains retrieved from GenBank.ResultsAmong 190 children with herpangina, 69.0% (131/190) were positive for enterovirus. Eight genotypes were identified, mainly including CV-A6 (39/131), CV-A4 (25/131), CV-A10 (24/131). The phylogenetic analysis showed one CV-A6 strain of Tongzhou was imported from Japan. CV-A10 strains were clustered into five groups (A-E). The dominant cluster of CV-A10 was Group E6 between 2009 and 2013, and converted to Group E5 after 2013. CV-A6 was the predominant pathogen causing herpangina in Tongzhou in 2018, followed by CV-A4 and CV-A10.ConclusionsThe circulation of coxsackievirus had spatiotemporal cluster. In controlling the transmission of herpangina, the surveillance and reporting system should be enhanced. creator: Tian-Shuo Zhao creator: Juan Du creator: Hong-Jun Li creator: Yan Cui creator: Yaqiong Liu creator: Yanna Yang creator: Fuqiang Cui creator: Qing-Bin Lu uri: https://doi.org/10.7717/peerj.9991 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhao et al. title: Injuries among children and adolescents in a rapidly growing urban African metropolis: a cross-sectional survey of 1,968 households in Dar es Salaam, Tanzania link: https://peerj.com/articles/10048 last-modified: 2020-10-15 description: ObjectivesTo assess the patterns and incidence of child and adolescent injury and explore associations with household deprivation and child characteristics in a low-income urban setting.Study DesignCross-sectional household survey in Dar es Salaam, Tanzania.MethodsData collection took place during July 2009. Injuries requiring medical attention were recorded with a one month period of recall. A total of 1,968 households representing 3,927 children and adolescents were visited by health workers. Gender-, age-, and type-specific injury incidence was compiled. Odds ratios were calculated to measure associations with child injury, perceived deprivation, household characteristics and child characteristics.ResultsOne household in five reported injuries. The estimated incidence was 3.2 per 10,000 child-years. The most common identifiable injuries were falls (41%), cuts (22%) and burns (16%). Male and younger children aged 1–4 years were at higher risk (respectively OR = 1.36; p = 0.004; OR = 1.47; p ≤ 0.001).ConclusionsIn Dar e Salaam injuries are common. Future investigations should take into account both subjective and objective measurements of relative household deprivation and a clear criteria for the assessment of injury severity in community-based survey contexts. creator: Mónica Alejandra Pérez Méndez creator: Hamisi A. Kigwangalla creator: Till Bärnighausen creator: Michael Lowery Wilson uri: https://doi.org/10.7717/peerj.10048 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Pérez Méndez et al. title: Multiple factors influence local perceptions of snow leopards and Himalayan wolves in the central Himalayas, Nepal link: https://peerj.com/articles/10108 last-modified: 2020-10-15 description: An understanding of local perceptions of carnivores is important for conservation and management planning. In the central Himalayas, Nepal, we interviewed 428 individuals from 85 settlements using a semi-structured questionnaire to quantitatively assess local perceptions and tolerance of snow leopards and wolves. We used generalized linear mixed effect models to assess influential factors, and found that tolerance of snow leopards was much higher than of wolves. Interestingly, having experienced livestock losses had a minor impact on perceptions of the carnivores. Occupation of the respondents had a strong effect on perceptions of snow leopards but not of wolves. Literacy and age had weak impacts on snow leopard perceptions, but the interaction among these terms showed a marked effect, that is, being illiterate had a more marked negative impact among older respondents. Among the various factors affecting perceptions of wolves, numbers of livestock owned and gender were the most important predictors. People with larger livestock herds were more negative towards wolves. In terms of gender, males were more positive to wolves than females, but no such pattern was observed for snow leopards. People’s negative perceptions towards wolves were also related to the remoteness of the villages. Factors affecting people’s perceptions could not be generalized for the two species, and thus need to be addressed separately. We suggest future conservation projects and programs should prioritize remote settlements. creator: Madhu Chetri creator: Morten Odden creator: Olivier Devineau creator: Thomas McCarthy creator: Per Wegge uri: https://doi.org/10.7717/peerj.10108 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Chetri et al. title: Conflicting phylogenetic signals in plastomes of the tribe Laureae (Lauraceae) link: https://peerj.com/articles/10155 last-modified: 2020-10-15 description: BackgroundGene tree discordance is common in phylogenetic analyses. Many phylogenetic studies have excluded non-coding regions of the plastome without evaluating their impact on tree topology. In general, plastid loci have often been treated as a single unit, and tree discordance among these loci has seldom been examined. Using samples of Laureae (Lauraceae) plastomes, we explored plastome variation among the tribe, examined the influence of non-coding regions on tree topology, and quantified intra-plastome conflict.ResultsWe found that the plastomes of Laureae have low inter-specific variation and are highly similar in structure, size, and gene content. Laureae was divided into three groups, subclades I, II and III. The inclusion of non-coding regions changed the phylogenetic relationship among the three subclades. Topologies based on coding and non-coding regions were largely congruent except for the relationship among subclades I, II and III. By measuring the distribution of phylogenetic signal across loci that supported different topologies, we found that nine loci (two coding regions, two introns and five intergenic spacers) played a critical role at the contentious node.ConclusionsOur results suggest that subclade III and subclade II are successively sister to subclade I. Conflicting phylogenetic signals exist between coding and non-coding regions of Laureae plastomes. Our study highlights the importance of evaluating the influence of non-coding regions on tree topology and emphasizes the necessity of examining discordance among different plastid loci in phylogenetic studies. creator: Tian-Wen Xiao creator: Yong Xu creator: Lu Jin creator: Tong-Jian Liu creator: Hai-Fei Yan creator: Xue-Jun Ge uri: https://doi.org/10.7717/peerj.10155 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Xiao et al. title: Colonization of novel algal habitats by juveniles of a marine tube-dwelling amphipod link: https://peerj.com/articles/10188 last-modified: 2020-10-15 description: BackgroundDispersal is an important process affecting population dynamics and connectivity. For marine direct developers, both adults and juveniles may disperse. Although the distribution of juveniles can be initially constrained by their mothers’ choice, they may be able to leave the parental habitat and colonize other habitats. We investigated the effect of habitat quality, patch size and presence of conspecific adults on the colonization of novel habitats by juveniles of the tube-dwelling amphipod Cymadusa filosa associated with the macroalgal host Sargassum filipendula.MethodsWe tested the factors listed above on the colonization of juveniles by manipulating natural and artificial plants in both the field and laboratory.ResultsIn the laboratory, juveniles selected high-quality habitats (i.e., natural alga), where both food and shelter are provided, when low-quality resources (i.e., artificial alga) were also available. In contrast, habitat quality and algal patch size did not affect the colonization by juveniles in the field. Finally, the presence of conspecific adults did not affect the colonization of juveniles under laboratory condition but had a weak effect in the field experiment. Our results suggest that C. filosa juveniles can select and colonize novel habitats, and that such process can be partially affected by habitat quality, but not by patch size. Also, the presence of conspecifics may affect the colonization by juveniles. Successful colonization by this specific developmental stage under different scenarios indicates that juveniles may act as a dispersal agent in this species. creator: Marilia Bueno creator: Glauco B.O. Machado creator: Fosca P.P. Leite uri: https://doi.org/10.7717/peerj.10188 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Bueno et al. title: Physical inactivity as a risk factor to mortality by ischemic heart disease during economic and political crisis in Brazil link: https://peerj.com/articles/10192 last-modified: 2020-10-15 description: BackgroundTo investigate the burden of mortality due to ischemic heart disease (IHD) attributable to low levels of physical activity in the Brazilian population (aged ≥ 25 years) before, during and after economic and political crises (2007–2017).MethodsThis study was focused on IHD as a cause of death. The International Statistical Classification of Diseases (10th revision) codes related to IHD have been mapped. The data used for the physical activity estimates of the present study refer to surveys with random sampling carried out in the Brazilian territory that considers all domains of physical activity. The contribution of physical activity for mortality due to IHD was estimated using a comparative risk assessment approach. In addition, we verified the association between mortality due to IHD attributable to low levels of physical activity according to the Socio-demographic Index of the Brazilian states.ResultsFor males it was estimated that in 2007 and 2017 there were, respectively, 9,585 and 11,821 deaths due to IHD as a result low physical activity. For females there were 8,689 deaths in 2007 and 10,779 deaths in 2017 due to IHD attributable to low physical activity. From 2007 to 2017, there was 12.0% (for males) and 16.0% (for females) of reduction in age-adjusted mortality rate due to IHD attributable to low physical activity. This reduction was not observed in the Northern and Northeastern regions of Brazil for the male population. Brazilian states with better socioeconomic conditions showed greater reductions in age-adjusted mortality rate due to IHD attributable to low physical activity (male: ρ = −0.74; female: ρ = −0.54)ConclusionThe fiscal austerity policies implemented and the lower investment in social programs in the period of economic and political crisis highlighted the social inequalities between Brazilian geographic regions for the burden of mortality due to IHD attributable to low levels of physical activity. creator: Diego Augusto Santos Silva uri: https://doi.org/10.7717/peerj.10192 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Silva title: Trypanosoma cruzi-infected triatomines and rodents co-occur in a coastal island of northern Chile link: https://peerj.com/articles/9967 last-modified: 2020-10-14 description: Trypanosoma cruzi, the cause agent of Chagas disease, is transmitted mainly by blood-feeding insects of the subfamily Triatominae. The T. cruzi life cycle alternates between triatomines and mammalian hosts, excluding birds and reptiles. Triatomines of Mepraia genus are wild vectors of T. cruzi in Chile. Mepraia specimens infected with T. cruzi have been detected in Pan de Azúcar and Santa María islands. The most common vertebrates that inhabit these islands are birds and reptiles, and it is unknown whether small mammals are present. Consequently, it is relevant to know whether there are any T. cruzi-infected small mammals on those islands to elucidate the T. cruzi cycle. To clarify this crossroads, islands of northern Chile were explored to determine if T. cruzi-infected triatomines and rodents co-occur in islands of northern Chile. T. cruzi DNA was detected by conventional and real-time PCR in three islands: on Santa María and Pan de Azúcar islands T. cruzi was detected in Mepraia sp samples, while on Pan de Azúcar (6.1%) and Damas islands (15%) was detected in the rodent Abrothrix olivacea. We show for the first time in Chile the occurrence of insular rodents infected with T. cruzi, and a complete T. cruzi life cycle in a coastal island. Our results provide new insights to understand the T. cruzi infection in the wild cycle. creator: Ricardo Campos-Soto creator: Gabriel Díaz-Campusano creator: Nicol Quiroga creator: Catalina Muñoz-San Martín creator: Ninette Rives-Blanchard creator: Fernando Torres-Pérez uri: https://doi.org/10.7717/peerj.9967 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Campos-Soto et al. title: Spatial benthic community analysis of shallow coral reefs to support coastal management in Culebra Island, Puerto Rico link: https://peerj.com/articles/10080 last-modified: 2020-10-14 description: Caribbean coral reefs provide essential ecosystem services to society, including fisheries, tourism and shoreline protection from coastal erosion. However, these reefs are also exhibiting major declining trends, leading to the evolution of novel ecosystems dominated by non-reef building taxa, with potentially altered ecological functions. In the search for effective management strategies, this study characterized coral reefs in front of a touristic beach which provides economic benefits to the surrounding coastal communities yet faces increasing anthropogenic pressures and conservation challenges. Haphazard photo-transects were used to address spatial variation patterns in the reef’s benthic community structure in eight locations. Statistically significant differences were found with increasing distance from the shoreline, reef rugosity, Diadema antillarum density, among reef locations, and as a function of recreational use. Nearshore reefs reflected higher percent macroalgal cover, likely due to increased exposure from both recreational activities and nearby unsustainable land-use practices. However, nearshore reefs still support a high abundance of the endangered reef-building coral Orbicella annularis, highlighting the need to conserve these natural shoreline protectors. There is an opportunity for local stakeholders and regulatory institutions to collaboratively implement sea-urchin propagation, restoration of endangered Acroporid coral populations, and zoning of recreational densities across reefs. Our results illustrate vulnerable reef hotspots where these management interventions are needed and recommend guidelines to address them. creator: Nicolás X. Gómez-Andújar creator: Edwin A. Hernandez-Delgado uri: https://doi.org/10.7717/peerj.10080 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Gómez-Andújar and Hernandez-Delgado title: Ecological stoichiometry of plant leaves, litter and soils in a secondary forest on China’s Loess Plateau link: https://peerj.com/articles/10084 last-modified: 2020-10-14 description: Ecological stoichiometry can reveal nutrient cycles in soil and plant ecosystems and their interactions. However, the ecological stoichiometry characteristics of leaf-litter-soil system of dominant grasses, shrubs and trees are still unclear as are their intrinsic relationship during vegetation restoration. This study selected three dominant plant types of grasses (Imperata cylindrica (I. cylindrica) and Artemisiasacrorum (A.sacrorum)), shrubs (Sophora viciifolia (S. viciifolia) and Hippophae rhamnoides (H. rhamnoides)) and trees (Quercus liaotungensis (Q. liaotungensis) and Betula platyphylla (B. platyphylla)) in secondary forest areas of the Chinese Loess Plateau to investigate ecological stoichiometric characteristics and their intrinsic relationships in leaf-litter-soil systems. The results indicated that N concentration and N:P ratios in leaf and litter were highest in shrubland; leaf P concentration in grassland was highest and litter in forestland had the highest P concentration. Soil C, N and P concentrations were highest in forestland (P < 0.05) and declined with soil depth. Based on the theory that leaf N:P ratio indicates nutritional limitation of plant growth, this study concluded that grass and shrub growth was limited by N and P element, respectively, and forest growth was limited by both of N and P elements. The relationships between the N concentration in soil, leaf and litter was not significant (P >0.5), but the soil P concentration was significantly correlated with litter P concentration (P < 0.05). These finding enhance understanding of nutrient limitations in different plant communities during vegetation restoration and provide insights for better management of vegetation restoration. creator: Zongfei Wang creator: Fenli Zheng uri: https://doi.org/10.7717/peerj.10084 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang and Zheng title: Quantile-dependent expressivity of plasma adiponectin concentrations may explain its sex-specific heritability, gene-environment interactions, and genotype-specific response to postprandial lipemia link: https://peerj.com/articles/10099 last-modified: 2020-10-14 description: Background“Quantile-dependent expressivity” occurs when the effect size of a genetic variant depends upon whether the phenotype (e.g. adiponectin) is high or low relative to its distribution. We have previously shown that the heritability (h2) of adiposity, lipoproteins, postprandial lipemia, pulmonary function, and coffee and alcohol consumption are quantile-specific. Whether adiponectin heritability is quantile specific remains to be determined.MethodsPlasma adiponectin concentrations from 4,182 offspring-parent pairs and 1,662 sibships from the Framingham Heart Study were analyzed. Quantile-specific heritability from offspring-parent (βOP,h2 = 2βOP/(1 + rspouse)) and full-sib regression slopes (βFS, h2 = {(1 + 8rspouseβFS)0.05-1}/(2rspouse)) were robustly estimated by quantile regression with nonparametric significance assigned from 1,000 bootstrap samples.ResultsQuantile-specific h2 (± SE) increased with increasing percentiles of the offspring’s age- and sex-adjusted adiponectin distribution when estimated from βOP (Ptrend = 2.2 × 10−6): 0.30 ± 0.03 at the 10th, 0.33 ± 0.04 at the 25th, 0.43 ± 0.04 at the 50th, 0.55 ± 0.05 at the 75th, and 0.57 ± 0.08 at the 90th percentile, and when estimated from βFS (Ptrend = 7.6 × 10−7): 0.42 ± 0.03 at the 10th, 0.44 ± 0.04 at the 25th, 0.56 ± 0.05 at the 50th, 0.73 ± 0.08 at the 75th, and 0.79 ± 0.11 at the 90th percentile. Consistent with quantile-dependent expressivity, adiponectin’s: (1) heritability was greater in women in accordance with their higher adiponection concentrations; (2) relationships to ADIPOQ polymorphisms were modified by adiposity in accordance with its adiponectin-lowering effect; (3) response to rosiglitazone was predicted by the 45T> G ADIPOQ polymorphism; (4) difference by ADIPOQ haplotypes increased linearly with increasing postprandial adiponectin concentrations.ConclusionAdiponectin heritability is quantile dependent, which may explain sex-specific heritability, gene-environment and gene-drug interactions, and postprandial response by haplotypes. creator: Paul T. Williams uri: https://doi.org/10.7717/peerj.10099 license: https://creativecommons.org/publicdomain/zero/1.0/ rights: ©2020 Williams title: Heparin-binding protein as a novel biomarker for sepsis-related acute kidney injury link: https://peerj.com/articles/10122 last-modified: 2020-10-14 description: BackgroundSepsis-related acute kidney injury (AKI) is associated with high morbidity and mortality among patients. Underlying pathomechanisms include capillary leakage and fluid loss into the interstitial tissue and constant exposure to pathogens results in activation of inflammatory cascades, organ dysfunction and subsequently organ damage.MethodsTo identify novel factors that trigger sepsis-related acute kidney injury, plasma levels of Granzyme A, as representative of a lymphocyte-derived protease, and heparin-binding protein as indicator for neutrophil-derived mediators, were investigated retrospectively in 60 sepsis patients.ResultsWhile no association was found between plasma levels of lymphocyte-derived Granzyme A and the incidence of sepsis-related AKI, sepsis patients with AKI had significantly higher plasma levels of heparin-binding protein compared to those without AKI. This applies both to heparin-binding protein peak values (43.30 ±  23.34 vs. 30.25 ±  15.63 pg/mL; p = 0.005) as well as mean values (27.93 ±  14.39 vs. 22.02 ±  7.65 pg/mL; p = 0.021). Furthermore, a heparin-binding protein cut-off value of 23.89 pg/mL was established for AKI diagnosis.ConclusionThis study identifies the neutrophil-derived heparin-binding protein as a valuable new biomarker for AKI in sepsis. Beyond the diagnostic perspective, this offers prospect for further research on pathogenesis of AKI and novel therapeutic approaches. creator: Sahra Pajenda creator: Andreja Figurek creator: Ludwig Wagner creator: Daniela Gerges creator: Alice Schmidt creator: Harald Herkner creator: Wolfgang Winnicki uri: https://doi.org/10.7717/peerj.10122 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Pajenda et al. title: Association of pre-pregnancy body mass index with adverse pregnancy outcome among first-time mothers link: https://peerj.com/articles/10123 last-modified: 2020-10-14 description: BackgroundStudies have reported an increased risk of adverse pregnancy outcome associated with pre-pregnancy body mass index (BMI). However, the data on such associations in urban areas of southern Chinese women is limited, which drive us to clarify the associations of pre-pregnancy BMI and the risks of adverse pregnancy outcomes (preterm birth (PTB) and low birth weight (LBW)) and maternal health outcomes (gestational hypertension and cesarean delivery).MethodsWe performed a hospital-based case-control study including 3,864 Southern Chinese women who gave first birth to a live singleton infant from January 2015 to December 2015. PTB was stratified into three subgroups according to gestational age (extremely PTB, very PTB and moderate PTB). Besides, we combined birth weight and gestational age to dichotomise as being small for gestational age (SGA, less than the tenth percentile of weight for gestation) and non-small for gestational age (NSGA, large than the tenth percentile of weight for gestation), gestational week was also classified into categories of term, 34-36 week and below 34 week.. We then divided newborns into six groups: (1) term and NSGA; (2) 34–36 week gestation and NSGA; (3) below 34 week gestation and NSGA; (4) term and SAG; (5) 34–36 week gestation and SAG; (6) below 34 week gestation and SAG. Adjusted logistic regression models was used to estimate the odds ratios of adverse outcomes.ResultsUnderweight women were more likely to give LBW (AOR = 1.44, 95% CI [1.11–1.89]), the similar result was seen in term and SAG as compared with term and NSAG (AOR = 1.78, 95% CI [1.45–2.17]), whereas underweight was significantly associated with a lower risk of gestational hypertension (AOR = 0.45, 95% CI [0.25–0.82) and caesarean delivery (AOR = 0.74, 95% CI [0.62–0.90]). The risk of extremely PTB is relatively higher among overweight and obese mothers in a subgroup analysis of PTB (AOR = 8.12, 95% CI [1.11–59.44]; AOR = 15.06, 95% CI [1.32–172.13], respectively). Both maternal overweight and obesity were associated with a greater risk of gestational hypertension (AOR = 1.71, 95% CI [1.06–2.77]; AOR = 5.54, 95% CI [3.02–10.17], respectively) and caesarean delivery (AOR = 1.91, 95% CI [1.53–2.38]; AOR = 1.85, 95% CI [1.21–2.82], respectively).ConclusionsOur study suggested that maternal overweight and obesity were associated with a significantly higher risk of gestational hypertension, caesarean delivery and extremely PTB. Underweight was correlated with an increased risk of LBW and conferred a protective effect regarding the risk for gestational hypertension and caesarean delivery for the first-time mothers among Southern Chinese. creator: Li Li creator: Yanhong Chen creator: Zhifeng Lin creator: Weiyan Lin creator: Yangqi Liu creator: Weilin Ou creator: Chengli Zeng creator: Li Ke uri: https://doi.org/10.7717/peerj.10123 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Li et al. title: Long non-coding RNA LINC01234 regulates proliferation, migration and invasion via HIF-2α pathways in clear cell renal cell carcinoma cells link: https://peerj.com/articles/10149 last-modified: 2020-10-14 description: Long non-coding RNAs (lncRNAs) have been proved to have an important role in different malignancies including clear cell renal cell carcinoma (ccRCC). However, their role in disease progression is still not clear. The objective of the study was to identify lncRNA-based prognostic biomarkers and further to investigate the role of one lncRNA LINC01234 in progression of ccRCC cells. We found that six adverse prognostic lncRNA biomarkers including LINC01234 were identified in ccRCC patients by bioinformatic analysis using The Cancer Genome Atlas database. LINC01234 knockdown impaired cell proliferation, migration and invasion in vitro as compared to negative control. Furthermore, the epithelial-mesenchymal transition was inhibited after LINC01234 knockdown. Additionally, LINC01234 knockdown impaired hypoxia-inducible factor-2a (HIF-2α) pathways, including a suppression of the expression of HIF-2α, vascular endothelial growth factor A, epidermal growth factor receptor, c-Myc, Cyclin D1 and MET. Together, these datas showed that LINC01234 was likely to regulate the progression of ccRCC by HIF-2α pathways, and LINC01234 was both a promising prognostic biomarker and a potential therapeutic target for ccRCC. creator: Feilong Yang creator: Cheng Liu creator: Guojiang Zhao creator: Liyuan Ge creator: Yimeng Song creator: Zhigang Chen creator: Zhuo Liu creator: Kai Hong creator: Lulin Ma uri: https://doi.org/10.7717/peerj.10149 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Yang et al. title: Belowground fungal community diversity, composition and ecological functionality associated with winter wheat in conventional and organic agricultural systems link: https://peerj.com/articles/9732 last-modified: 2020-10-13 description: Understanding the impacts of agricultural practices on belowground fungal communities is crucial in order to preserve biological diversity in agricultural soils and enhance their role in agroecosystem functioning. Although fungal communities are widely distributed, relatively few studies have correlated agricultural production practices. We investigated the diversity, composition and ecological functionality of fungal communities in roots of winter wheat (Triticum aestivum) growing in conventional and organic farming systems. Direct and nested polymerase chain reaction (PCR) amplifications spanning the internal transcribed spacer (ITS) region of the rDNA from pooled fine root samples were performed with two different sets of fungal specific primers. Fungal identification was carried out through similarity searches against validated reference sequences (RefSeq). The R package ‘picante’ and FUNGuild were used to analyse fungal community composition and trophic mode, respectively. Either by direct or cloning sequencing, 130 complete ITS sequences were clustered into 39 operational taxonomic units (OTUs) (25 singletons), belonging to the Ascomycota (24), the Basidiomycota (14) and to the Glomeromycota (1). Fungal communities from conventional farming sites are phylogenetically more related than expected by chance. Constrained ordination analysis identified total N, total S and Pcal that had a significant effect on the OTU’s abundance and distribution, and a further correlation with the diversity of the co-occurring vegetation could be hypothesised. The functional predictions based on FUNGuild suggested that conventional farming increased the presence of plant pathogenic fungi compared with organic farming. Based on diversity, OTU distribution, nutrition mode and the significant phylogenetic clustering of fungal communities, this study shows that fungal communities differ across sampling sites, depending on agricultural practices. Although it is not fully clear which factors determine the fungal communities, our findings suggest that organic farming systems have a positive effect on fungal communities in winter wheat crops. creator: Sigisfredo Garnica creator: Ronja Rosenstein creator: Max Emil Schön uri: https://doi.org/10.7717/peerj.9732 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Garnica et al. title: Differentially expressed transcripts and associated protein pathways in basilar artery smooth muscle cells of the high-salt intake–induced hypertensive rat link: https://peerj.com/articles/9849 last-modified: 2020-10-13 description: The pathology of cerebrovascular disorders, such as hypertension, is associated with genetic changes and dysfunction of basilar artery smooth muscle cells (BASMCs). Long-term high-salt diets have been associated with the development of hypertension. However, the molecular mechanisms underlying salt-sensitive hypertension-induced BASMC modifications have not been well defined, especially at the level of variations in gene transcription. Here, we utilized high-throughput sequencing and subsequent signaling pathway analyses to find a two–fold change or greater upregulated expression of 203 transcripts and downregulated expression of 165 transcripts in BASMCs derived from rats fed a high-salt diet compared with those from control rats. These differentially expressed transcripts were enriched in pathways involved in cellular, morphological, and structural plasticity, autophagy, and endocrine regulation. These transcripts changes in the BASMCs derived from high-salt intake–induced hypertensive rats may provide critical information about multiple cellular processes and biological functions that occur during the development of cerebrovascular disorders and provide potential new targets to help control or block the development of hypertension. creator: Junhao Huang creator: Lesha Zhang creator: Yang Fang creator: Wan Jiang creator: Juan Du creator: Jinhang Zhu creator: Min Hu creator: Bing Shen uri: https://doi.org/10.7717/peerj.9849 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Huang et al. title: Data sharing policies of journals in life, health, and physical sciences indexed in Journal Citation Reports link: https://peerj.com/articles/9924 last-modified: 2020-10-13 description: BackgroundMany scholarly journals have established their own data-related policies, which specify their enforcement of data sharing, the types of data to be submitted, and their procedures for making data available. However, except for the journal impact factor and the subject area, the factors associated with the overall strength of the data sharing policies of scholarly journals remain unknown. This study examines how factors, including impact factor, subject area, type of journal publisher, and geographical location of the publisher are related to the strength of the data sharing policy.MethodsFrom each of the 178 categories of the Web of Science’s 2017 edition of Journal Citation Reports, the top journals in each quartile (Q1, Q2, Q3, and Q4) were selected in December 2018. Of the resulting 709 journals (5%), 700 in the fields of life, health, and physical sciences were selected for analysis. Four of the authors independently reviewed the results of the journal website searches, categorized the journals’ data sharing policies, and extracted the characteristics of individual journals. Univariable multinomial logistic regression analyses were initially conducted to determine whether there was a relationship between each factor and the strength of the data sharing policy. Based on the univariable analyses, a multivariable model was performed to further investigate the factors related to the presence and/or strength of the policy.ResultsOf the 700 journals, 308 (44.0%) had no data sharing policy, 125 (17.9%) had a weak policy, and 267 (38.1%) had a strong policy (expecting or mandating data sharing). The impact factor quartile was positively associated with the strength of the data sharing policies. Physical science journals were less likely to have a strong policy relative to a weak policy than Life science journals (relative risk ratio [RRR], 0.36; 95% CI [0.17–0.78]). Life science journals had a greater probability of having a weak policy relative to no policy than health science journals (RRR, 2.73; 95% CI [1.05–7.14]). Commercial publishers were more likely to have a weak policy relative to no policy than non-commercial publishers (RRR, 7.87; 95% CI, [3.98–15.57]). Journals by publishers in Europe, including the majority of those located in the United Kingdom and the Netherlands, were more likely to have a strong data sharing policy than a weak policy (RRR, 2.99; 95% CI [1.85–4.81]).ConclusionsThese findings may account for the increase in commercial publishers’ engagement in data sharing and indicate that European national initiatives that encourage and mandate data sharing may influence the presence of a strong policy in the associated journals. Future research needs to explore the factors associated with varied degrees in the strength of a data sharing policy as well as more diverse characteristics of journals related to the policy strength. creator: Jihyun Kim creator: Soon Kim creator: Hye-Min Cho creator: Jae Hwa Chang creator: Soo Young Kim uri: https://doi.org/10.7717/peerj.9924 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kim et al. title: Plasma miRNA profiles associated with stable warfarin dosage in Chinese patients link: https://peerj.com/articles/9995 last-modified: 2020-10-13 description: BackgroundWe used bioinformatic analysis and quantitative reverse transcription polymerase chain reaction (RT-qPCR) assays to investigate the association between plasma microRNAs (miRNAs) and stable warfarin dosage in a Chinese Han population.MethodsBioinformatics analysis was used to screen out potential warfarin dose-associated miRNAs. Three plasma miRNAs were validated in 99 samples by RT-qPCR. Kruskal–Wallis test and multivariate logistic regression were used to compare differences in plasma miRNAs expression levels between three warfarin dosage groups.ResultsThere were significant between-group differences among the three dose groups for hsa-miR-133b expression (p = 0.005), but we observed an “n-shaped” dose-dependent curve rather than a linear relationship. Expression levels of hsa-miR-24-3p (p = 0.475) and hsa-miR-1276 (p = 0.558) were not significantly different in the multivariate logistic regression.ConclusionmiRNAs have received extensive attention as ideal biomarkers and possible therapeutic targets for various diseases. However, they are not yet widely used in precision medicine. Our results indicate that hsa-miR-133b may be a possible reference factor for the warfarin dosage algorithm. These findings emphasize the importance of a comprehensive evaluation of complex relationships in warfarin dose prediction models and provide new avenues for future pharmacogenomics studies. creator: Li Zhao creator: Jin Wang creator: Shaoxin Shi creator: Yuan Wu creator: Jumei Liu creator: Shiwei He creator: Yue Zou creator: Huabin Xie creator: Shengxiang Ge creator: Huiming Ye uri: https://doi.org/10.7717/peerj.9995 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhao et al. title: Mitochondrial genomes of twelve species of hyperdiverse Trigonopterus weevils link: https://peerj.com/articles/10017 last-modified: 2020-10-13 description: Mitochondrial genomes of twelve species of Trigonopterus weevils are presented, ten of them complete. We describe their gene order and molecular features and test their potential for reconstructing the phylogeny of this hyperdiverse genus comprising > 1,000 species. The complete mitochondrial genomes examined herein ranged from 16,501 bp to 21,007 bp in length, with an average AT content of 64.2% to 69.7%. Composition frequencies and skews were generally lower across species for atp6, cox1-3, and cob genes, while atp8 and genes coded on the minus strand showed much higher divergence at both nucleotide and amino acid levels. Most variation within genes was found at the codon level with high variation at third codon sites across species, and with lesser degree at the coding strand level. Two large non-coding regions were found, CR1 (between rrnS and trnI genes) and CR2 (between trnI and trnQ), but both with large variability in length; this peculiar structure of the non-coding region may be a derived character of Curculionoidea. The nad1 and cob genes exhibited an unusually high interspecific length variation of up to 24 bp near the 3′ end. This pattern was probably caused by a single evolutionary event since both genes are only separated by trnS2 and length variation is extremely rare in mitochondrial protein coding genes. We inferred phylogenetic trees using protein coding gene sequences implementing both maximum likelihood and Bayesian approaches, each for both nucleotide and amino acid sequences. While some clades could be retrieved from all reconstructions with high confidence, there were also a number of differences and relatively low support for some basal nodes. The best partition scheme of the 13 protein coding sequences obtained by IQTREE suggested that phylogenetic signal is more accurate by splitting sequence variation at the codon site level as well as coding strand, rather than at the gene level. This result corroborated the different patterns found in Trigonopterus regarding to A+T frequencies and AT and GC skews that also greatly diverge at the codon site and coding strand levels. creator: Raden Pramesa Narakusumo creator: Alexander Riedel creator: Joan Pons uri: https://doi.org/10.7717/peerj.10017 license: https://creativecommons.org/licenses/by-nc/4.0/ rights: ©2020 Narakusumo et al. title: Utilising one minute and four minute recovery when employing the resistance training contrast method does not negatively affect subsequent jump performance in the presence of concurrent training link: https://peerj.com/articles/10031 last-modified: 2020-10-13 description: BackgroundLittle is known about contrast training and post-activation performance enhancement (PAPE) in a same day concurrent training model. The aim of the current study was to examine the use of two short duration (1-min and 4-min) recovery periods on drop jump performance in same day concurrently trained athletes.MethodsTen professional Australian Rules footballers (age, 20.6 ± 1.9 yr; height, 184.8 ± 6.9 cm; body mass, 85.8 ± 8.4 kg) completed two resistance training sessions with different PAPE recovery durations; 1-min and 4-min, 1 h following a field-based endurance session. Baseline (pre) drop jumps were compared to post-test maximal drop jumps, performed after each set of three squats (where each participant was encouraged to lift as heavy as they could), to determine changes between 1-min and 4-min recovery periods. Data were analysed by fitting a mixed model (significance was set at P ≤ 0.05). Corrected Hedges’ g standardised effect sizes ±95% confidence limits were calculated using group means ± SDs.ResultsThere were no significant differences between baseline and experimental sets 1, 2 and 3 for reactive strength index (RSI), flight time, and total and relative impulse for either recovery duration. However, for contact time, 1-min baseline was significantly different from set 2 (mean difference; 95% CI [0.029; 0.000–0.057 s], P = 0.047, ES; 95% CI [−0.27; −1.20 to 0.66]). For RSI and flight time, 1-min was significantly higher than 4-min (RSI: 0.367; 0.091 to 0.642, P = 0.010, ES; 95% CI [0.52; −0.37 to 1.42]; flight time: 0.033; 0.003 to 0.063 s, P = 0.027, ES; 95% CI [0.86; −0.06 to 1.78]).DiscussionShort recovery periods of 1-min may be a time-efficient form of prescribing strength-power exercise in contrast loading schemes. Longer recovery periods do not appear to benefit immediate, subsequent performance. creator: Dean Ritchie creator: Justin W.L. Keogh creator: Peter Reaburn creator: Jonathan D. Bartlett uri: https://doi.org/10.7717/peerj.10031 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Ritchie et al. title: Plastome structure and adaptive evolution of Calanthe s.l. species link: https://peerj.com/articles/10051 last-modified: 2020-10-13 description: Calanthe s.l. is the most diverse group in the tribe Collabieae (Orchidaceae), which are pantropical in distribution. Illumina sequencing followed by de novo assembly was used in this study, and the plastid genetic information of Calanthe s.l. was used to investigate the adaptive evolution of this taxon. Herein, the complete plastome of five Calanthe s.l. species (Calanthe davidii, Styloglossum lyroglossa, Preptanthe rubens, Cephalantheropsis obcordata, and Phaius tankervilliae) were determined, and the two other published plastome sequences of Calanthe s.l. were added for comparative analyses to examine the evolutionary pattern of the plastome in the alliance. The seven plastomes ranged from 150,181 bp (C. delavayi) to 159,014 bp (C. davidii) in length and were all mapped as circular structures. Except for the three ndh genes (ndhC, ndhF, and ndhK) lost in C. delavayi, the remaining six species contain identical gene orders and numbers (115 gene). Nucleotide diversity was detected across the plastomes, and we screened 14 mutational hotspot regions, including 12 non-coding regions and two gene regions. For the adaptive evolution investigation, three species showed positive selected genes compared with others, C. obcordata (cemA), S. lyroglossa (infA, ycf1 and ycf2) and C. delavayi (nad6 and ndhB). Six genes were under site-specific positive selection in Calanthe s.l., namely, accD, ndhB, ndhD, rpoC2, ycf1, and ycf2, most of which are involved in photosynthesis. These results, including the new plastomes, provide resources for the comparative plastome, breeding, and plastid genetic engineering of orchids and flowering plants. creator: Yanqiong Chen creator: Hui Zhong creator: Yating Zhu creator: Yuanzhen Huang creator: Shasha Wu creator: Zhongjian Liu creator: Siren Lan creator: Junwen Zhai uri: https://doi.org/10.7717/peerj.10051 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: An efficient sorghum protoplast assay for transient gene expression and gene editing by CRISPR/Cas9 link: https://peerj.com/articles/10077 last-modified: 2020-10-13 description: Protoplasts are commonly used in genetic and breeding research. In this study, the isolation of sorghum protoplasts was optimized and applied to transient gene expression and editing by CRISPR/Cas9. The protoplast was most viable in 0.5 M mannitol, which was the highest of three concentrations after 48- and 72-hours treatments. Using this method we can derive an average of 1.6×106 cells which vary from 5 to 22 nm in size. The average transfection of the protoplasts was 68.5% using the PEG-mediated method. The subcellular assays located Sobic.002G279100-GFP and GFP proteins in the cell compartments as predicted bioinformatically. Two CRISPR/Cas9 plasmids were transfected into sorghum protoplasts to screen for an appropriate sgRNA for gene editing. One plasmid can correctly edit the target region using a single protoplast cell as template DNA. Our results indicated that the protoplast assays as optimized are suitable for transient gene expression and sgRNA screening in CRISPR/Cas9 gene editing procedures. creator: Ruirui Meng creator: Chenchen Wang creator: Lihua Wang creator: Yanlong Liu creator: Qiuwen Zhan creator: Jiacheng Zheng creator: Jieqin Li uri: https://doi.org/10.7717/peerj.10077 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Meng et al. title: Integrating multiple microarray dataset analysis and machine learning methods to reveal the key genes and regulatory mechanisms underlying human intervertebral disc degeneration link: https://peerj.com/articles/10120 last-modified: 2020-10-13 description: Intervertebral disc degeneration (IDD), a major cause of lower back pain, has multiple contributing factors including genetics, environment, age, and loading history. Bioinformatics analysis has been extensively used to identify diagnostic biomarkers and therapeutic targets for IDD diagnosis and treatment. However, multiple microarray dataset analysis and machine learning methods have not been integrated. In this study, we downloaded the mRNA, microRNA (miRNA), long noncoding RNA (lncRNA), and circular RNA (circRNA) expression profiles (GSE34095, GSE15227, GSE63492GSE116726, GSE56081 and GSE67566) associated with IDD from the GEO database. Using differential expression analysis and recursive feature elimination, we extracted four optimal feature genes. We then used the support vector machine (SVM) to make a classification model with the four optimal feature genes. The ROC curve was used to evaluate the model’s performance, and the expression profiles (GSE63492, GSE116726, GSE56081, and GSE67566) were used to construct a competitive endogenous RNA (ceRNA) regulatory network and explore the underlying mechanisms of the feature genes. We found that three miRNAs (hsa-miR-4728-5p, hsa-miR-5196-5p, and hsa-miR-185-5p) and three circRNAs (hsa_circRNA_100723, hsa_circRNA_104471, and hsa_circRNA_100750) were important regulators with more interactions than the other RNAs across the whole network. The expression level analysis of the three datasets revealed that BCAS4 and SCRG1 were key genes involved in IDD development. Ultimately, our study proposes a novel approach to determining reliable and effective targets in IDD diagnosis and treatment. creator: Hongze Chang creator: Xiaolong Yang creator: Kemin You creator: Mingwei Jiang creator: Feng Cai creator: Yan Zhang creator: Liang Liu creator: Hui Liu creator: Xiaodong Liu uri: https://doi.org/10.7717/peerj.10120 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chang et al. title: Upper-room ultraviolet air disinfection might help to reduce COVID-19 transmission in buildings: a feasibility study link: https://peerj.com/articles/10196 last-modified: 2020-10-13 description: As the world’s economies come out of the lockdown imposed by the COVID-19 pandemic, there is an urgent need for technologies to mitigate COVID-19 transmission in confined spaces such as buildings. This feasibility study looks at one such technology, upper-room ultraviolet (UV) air disinfection, that can be safely used while humans are present in the room space, and which has already proven its efficacy as an intervention to inhibit the transmission of airborne diseases such as measles and tuberculosis. Using published data from various sources, it is shown that the SARS-CoV-2 virus, the causative agent of COVID-19, is highly likely to be susceptible to UV-C damage when suspended in air, with a UV susceptibility constant likely to be in the region 0.377–0.590 m2/J, similar to that for other aerosolised coronaviruses. As such, the UV-C flux required to disinfect the virus is expected to be acceptable and safe for upper-room applications. Through analysis of expected and worst-case scenarios, the efficacy of the upper-room UV-C approach for reducing COVID-19 transmission in confined spaces (with moderate but sufficient ceiling height) is demonstrated. Furthermore, it is shown that with SARS-CoV-2, it should be possible to achieve high equivalent air change rates using upper-room UV air disinfection, suggesting that the technology might be particularly applicable to poorly ventilated spaces. creator: Clive B. Beggs creator: Eldad J. Avital uri: https://doi.org/10.7717/peerj.10196 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Beggs and Avital title: Soil carbon, nitrogen and phosphorus ecological stoichiometry shifts with tree species in subalpine plantations link: https://peerj.com/articles/9702 last-modified: 2020-10-12 description: Understanding ecological stoichiometric characteristics of soil nutrient elements, such as carbon (C), nitrogen (N) and phosphorus (P) is crucial to guide ecological restoration of plantations in ecologically vulnerable areas, such as alpine and subalpine regions. However, there has been only a few related studies, and thus whether and how different tree species would affect soil C:N:P ecological stoichiometry remains unclear. We compared soil C:N:P ecological stoichiometry of Pinus tabulaeformis, Larix kaempferi and Cercidiphyllum japonicum to primary shrubland in a subalpine region. We observed strong tree-specific and depth-dependent effects on soil C:N:P stoichiometry in subalpine plantations. In general, the C:N, C:P and N:P of topsoil (0–10 cm) are higher than subsoil (>10 cm) layer at 0–30 cm depth profiles. The differences in C:N, N:P and C:P at the topsoil across target tree species were significantly linked to standing litter stock, tree biomass/total aboveground biomass and Margalef’s index of plant community, respectively, whereas the observed variations of C:N, N:P and C:P ratio among soil profiles are closely related to differences in soil bulk density, soil moisture, the quantity and quality of aboveground litter inputs as well as underground fine root across plantations examined. Our results highlight that soil nutrients in plantation depend on litter quantity and quality of selected tree species as well as soil physical attributes. Therefore, matching site with trees is crucial to enhance ecological functioning in degraded regions resulting from human activity. creator: Kaibin Qi creator: Xueyong Pang creator: Bing Yang creator: Weikai Bao uri: https://doi.org/10.7717/peerj.9702 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Qi et al. title: Genetic characterization of feline panleukopenia virus from dogs in Vietnam reveals a unique Thr101 mutation in VP2 link: https://peerj.com/articles/9752 last-modified: 2020-10-12 description: BackgroundCanine parvovirus type 2 (CPV-2) and feline parvovirus (FPV) are known as the main causes of several serious diseases and have a severe impact on puppies and kittens, respectively. FPV and new CPV-2 variants are all able to infect cats, causing diseases indistinguishable from feline panleukopenia. However, FPV only replicates efficiently in feline cells in vitro and replicates in dogs in the thymus and bone marrow without being shed in feces. In our previous study, the genotypes of six parvoviral isolates were unable to be identified using a SimpleProbe® real-time PCR assay.MethodsIn the present study, we characterized previously unidentified FPV-like viruses isolated from dogs in Vietnam. The six isolates were utilized to complete VP2 gene sequencing and to conduct phylogenetic analyses.ResultsSequence analysis of the six parvoviral strains identified the species as being similar to FPV. Phylogenetic analysis demonstrated that the complete VP2 genes of the strains are similar to those of FPV. The FPV-like strains contain a Thr101 mutation in the VP2 protein, which is different from prototype FPV strains.DiscussionOur data provide evidence for the existence of changes in the charge, protein contact potential and molecular surface of the core of the receptor-binding size with an Ile101 to Thr101 mutation. This is also the first study to provide reliable evidence that FPV may be a threat to the Vietnamese dog population. creator: Minh Hoang creator: Cheng-Nan Wu creator: Chuen-Fu Lin creator: Huong Thanh Thi Nguyen creator: Van Phan Le creator: Ming-Tang Chiou creator: Chao-Nan Lin uri: https://doi.org/10.7717/peerj.9752 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Hoang et al. title: Physiological aspects and energetic contribution in 20s:10s high-intensity interval exercise at different intensities link: https://peerj.com/articles/9791 last-modified: 2020-10-12 description: BackgroundOne of the most popular high-intensity interval exercises is the called “Tabata Protocol”. However, most investigations have limitations in describing the work intensity, and this fact appears to be due to the protocol unfeasibility. Furthermore, the physiological demands and energetic contribution during this kind of exercise remain unclear.MethodsEight physically active students (21.8 ± 3.7 years) and eight well-trained cycling athletes (27.8 ± 6.4 years) were enrolled. In the first visit, we collected descriptive data and the peak power output (PPO). On the next three visits, in random order, participants performed interval training with the same time structure (effort:rest 20s:10s) but using different intensities (115%, 130%, and 170% of PPO). We collected the number of sprints, power output, oxygen consumption, blood lactate, and heart rate.ResultsThe analysis of variance for multivariate test (number of sprints, power output, blood lactate, peak heart rate and percentage of maximal heart rate) showed significant differences between groups (F = 9.62; p = 0.001) and intensities (F = 384.05; p < 0.001), with no interactions (F = 0.94; p = 0.57). All three energetic contributions and intensities were different between protocols. The higher contribution was aerobic, followed by alactic and lactic. The aerobic contribution was higher at 115%PPO, while the alactic system showed higher contribution at 130%PPO. In conclusion, the aerobic system was predominant in the three exercise protocols, and we observed a higher contribution at lower intensities. creator: Gabriel V. Protzen creator: Charles Bartel creator: Victor S. Coswig creator: Paulo Gentil creator: Fabricio B. Del Vecchio uri: https://doi.org/10.7717/peerj.9791 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Protzen et al. title: Genomic signatures of globally enhanced gene duplicate accumulation in the megadiverse higher Diptera fueling intralocus sexual conflict resolution link: https://peerj.com/articles/10012 last-modified: 2020-10-12 description: Gene duplication is an important source of evolutionary innovation. To explore the relative impact of gene duplication during the diversification of major insect model system lineages, we performed a comparative analysis of lineage-specific gene duplications in the fruit fly Drosophila melanogaster (Diptera: Brachycera), the mosquito Anopheles gambiae (Diptera: Culicomorpha), the red flour beetle Tribolium castaneum (Coleoptera), and the honeybee Apis mellifera (Hymenoptera). Focusing on close to 6,000 insect core gene families containing maximally six paralogs, we detected a conspicuously higher number of lineage-specific duplications in Drosophila (689) compared to Anopheles (315), Tribolium (386), and Apis (223). Based on analyses of sequence divergence, phylogenetic distribution, and gene ontology information, we present evidence that an increased background rate of gene duplicate accumulation played an exceptional role during the diversification of the higher Diptera (Brachycera), in part by providing enriched opportunities for intralocus sexual conflict resolution, which may have boosted speciation rates during the early radiation of the megadiverse brachyceran subclade Schizophora. creator: Riyue Bao creator: Markus Friedrich uri: https://doi.org/10.7717/peerj.10012 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Bao and Friedrich title: Instrumental validity and intra/inter-rater reliability of a novel low-cost digital pressure algometer link: https://peerj.com/articles/10162 last-modified: 2020-10-12 description: BackgroundPain assessment is a key measure that accompanies treatments in a wide range of clinical settings. A low-cost valid and reliable pressure algometer would allow objective assessment of pressure pain to assist a variety of health professionals. However, the pressure algometer is often expensive, which limits its daily use in both clinical and research settings.ObjectivesThis study aimed to assess the instrumental validity, and the intra- and inter-rater reliability of an inexpensive digital adapted pressure algometer.MethodsA single rater applied 60 random compressions on a force platform. The pressure pain thresholds of 20 volunteers were collected twice (3 days apart) by two raters. The main outcome measurements were as follows: the maximal peak force (in kPa) and the pressure pain threshold (adapted pressure algometer vs. force platform). Cronbach’s α test was used to assess internal consistency. The standard error of measurement provided estimates of measurement error, and the measurement bias was estimated with the Bland–Altman method, with lower and upper limits of agreement.ResultsNo differences were observed when comparing the compression results (P = 0.51). The validity and internal intra-rater consistencies ranged from 0.84 to 0.99, and the standard error of measurement from 0.005 to 0.04 kPa. Very strong (r = 0.73–0.74) to near-perfect (r = 0.99) correlations were found, with a low risk of bias for all measurements. The results demonstrated the validity and intra-rater reliability of the digitally adapted pressure algometer. Inter-rater reliability results were moderate (r = 0.55–0.60; Cronbach’s α = 0.71–0.75).ConclusionThe adapted pressure algometer provide valid and reliable measurements of pressure pain threshold. The results support more widespread use of the pressure pain threshold method among clinicians. creator: Daniel Jerez-Mayorga creator: Carolina Fernanda dos Anjos creator: Maria de Cássia Macedo creator: Ilha Gonçalves Fernandes creator: Esteban Aedo-Muñoz creator: Leonardo Intelangelo creator: Alexandre Carvalho Barbosa uri: https://doi.org/10.7717/peerj.10162 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Jerez-Mayorga et al. title: Mutations of SARS-CoV-2 nsp14 exhibit strong association with increased genome-wide mutation load link: https://peerj.com/articles/10181 last-modified: 2020-10-12 description: SARS-CoV-2 is a betacoronavirus responsible for COVID-19, a pandemic with global impact that first emerged in late 2019. Since then, the viral genome has shown considerable variance as the disease spread across the world, in part due to the zoonotic origins of the virus and the human host adaptation process. As a virus with an RNA genome that codes for its own genomic replication proteins, mutations in these proteins can significantly impact the variance rate of the genome, affecting both the survival and infection rate of the virus, and attempts at combating the disease. In this study, we analyzed the mutation densities of viral isolates carrying frequently observed mutations for four proteins in the RNA synthesis complex over time in comparison to wildtype isolates. Our observations suggest mutations in nsp14, an error-correcting exonuclease protein, have the strongest association with increased mutation load without selective pressure and across the genome, compared to nsp7, nsp8 and nsp12, which form the core polymerase complex. We propose nsp14 as a priority research target for understanding genomic variance rate in SARS-CoV-2 isolates and nsp14 mutations as potential predictors for high mutability strains. creator: Doğa Eskier creator: Aslı Suner creator: Yavuz Oktay creator: Gökhan Karakülah uri: https://doi.org/10.7717/peerj.10181 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Eskier et al. title: Convolutional neural network in upper limb functional motion analysis after stroke link: https://peerj.com/articles/10124 last-modified: 2020-10-09 description: In this work, implementation of Convolutional Neural Network (CNN) for the purpose of analysis of functional upper limb movement pattern was applied. The main aim of the study was to compare motion of selected activities of daily living of participants after stroke with the healthy ones (in similar age). The optical, marker-based motion capture system was applied for the purpose of data acquisition. There were some attempts made in order to find the existing differences in the motion pattern of the upper limb. For this purpose, the motion features of dominant and non-dominant upper limb of healthy participants were compared with motion features of paresis and non-paresis upper limbs of participants after stroke. On the basis of the newly collected data set, a new CNN application was presented to the classification of motion data in two different class label configurations. Analyzing individual segments of the upper body, it turned out that the arm was the most sensitive segment for capturing changes in the trajectory of the lifting movements of objects. creator: Agnieszka Szczęsna creator: Monika Błaszczyszyn creator: Aleksandra Kawala-Sterniuk uri: https://doi.org/10.7717/peerj.10124 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Szczęsna et al. title: The effectiveness of the Peyton’s 4-step teaching approach on skill acquisition of procedures in health professions education: A systematic review and meta-analysis with integrated meta-regression link: https://peerj.com/articles/10129 last-modified: 2020-10-09 description: BackgroundAcquisition of procedures is an important element in health professions education. Traditionally procedures are taught using a “see one - do one” approach. That is a teacher demonstrates and describes a procedure and afterwards the students practice the procedure. A more recent teaching approach for the acquisition of procedural skills was presented by Walker and Peyton. Peyton’s teaching approach is a stepwise teaching approach and consists of the following four steps: demonstration, deconstruction, comprehension and performance. The aims of this study were (i) to systematically evaluate the effectiveness of Peyton’s 4–step teaching approach on the acquisition of procedural skills in health professions education and (ii) to evaluate whether studies with fewer students per teacher showed a larger between group difference than studies with more students per teacher.MethodsWe searched in Medline, PsycInfo, Embase and ERIC for eligible studies. Records were screened by two independent reviewers. A random effects meta-analysis was performed to evaluate skill acquisition and time needed to perform the procedures at post-acquisition and retention tests. A meta-regression was used to explore the effect of the number of students per teacher on the estimated effect of the educational interventions.ResultsAn effect size of 0.45 SMD (95% CI [0.15; 0.75]) at post-acquisition and 0.7 SMD (95% CI [−0.09; 1.49]) at retention testing were in favour of Peyton’s teaching approach for skill acquisition. The groups using Peyton’s teaching approach needed considerably less time to perform the procedure at post-acquisition (SMD: −0.8; 95% [CI −2.13 to 1.62]) and retention (SMD: −2.65; 95% CI [−7.77 to 2.47]) testing. The effectiveness of Peyton’s teaching approach was less clear in subgroup analyses using peer teachers. Meta-regression showed that the number of students per teacher was an important moderator variable.ConclusionPeyton’s teaching approach is an effective teaching approach for skill acquisition of procedural skills in health professions education. When peer students or student tutors are used as teachers the effectiveness of Peyton’s teaching approach is less clear. Peyton’s teaching approach is more effective when small groups with few students per teacher are used. creator: Katia Giacomino creator: Rahel Caliesch creator: Karl Martin Sattelmayer uri: https://doi.org/10.7717/peerj.10129 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Giacomino et al. title: Validity of an inertial system for measuring velocity, force, and power during hamstring exercises performed on a flywheel resistance training device link: https://peerj.com/articles/10169 last-modified: 2020-10-09 description: BackgroundInertial hamstring exercises promote functional changes leading to lower rates of hamstring injuries. However, variable training measurement systems have not been specifically validated for hamstring exercises. Accordingly, this study aimed to evaluate the validity of the Inertial Measurement System (IMS) to measure the velocity, force, and power during the performance of different hamstring exercises on a flywheel resistance training device.MethodsFifteen males (average age: 22.4 ± 2.5 years; body mass: 77.3 ± 9.8 kg; height: 179.5 ± 7.4 cm; weekly physical activity: 434.0 ± 169.2 min; years of strength training: 4.3 ± 2.2 years) performed the bilateral stiff-leg deadlift (SDL), 45° hip extension (HE), and unilateral straight knee bridge (SKB) in two sessions (familiarization and evaluation) with a 1-week interval between them. The velocity, force, and power (average and peak values) in the concentric and eccentric phases for each of the exercises were recorded simultaneously with IMS and MuscleLab.ResultsConsistency between IMS and MuscleLab was good to excellent for all variables, with r ranges from 0.824 to 0.966 in SDL, from 0.822 to 0.971 in HE, and from 0.806 to 0.969 in SKB. Acceptable levels of agreement between devices were observed in general for all exercises, the “bias” ranging from 1.1% to 13.2%. Although MuscleLab showed higher values than IMS for peak velocity, force and power values, the effect size was only relevant for 5 of the 36 parameters. IMS is a new and valid system to monitor inertial hamstring exercises on a new flywheel device. In this way, IMS could have potential practical applications for any professional or athlete who wants to monitor inertial hamstring exercises. creator: Rodrigo Martín-San Agustín creator: Mariana Sánchez-Barbadora creator: José A. García-Vidal uri: https://doi.org/10.7717/peerj.10169 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Martín-San Agustín et al. title: The phylogenetics of Teleosauroidea (Crocodylomorpha, Thalattosuchia) and implications for their ecology and evolution link: https://peerj.com/articles/9808 last-modified: 2020-10-08 description: Teleosauroidea was a clade of ancient crocodylomorphs that were a key element of coastal marine environments during the Jurassic. Despite a 300-year research history and a recent renaissance in the study of their morphology and taxonomy, macroevolutionary studies of teleosauroids are currently limited by our poor understanding of their phylogenetic interrelationships. One major problem is the genus Steneosaurus, a wastebasket taxon recovered as paraphyletic or polyphyletic in phylogenetic analyses. We constructed a newly updated phylogenetic data matrix containing 153 taxa (27 teleosauroids, eight of which were newly added) and 502 characters, which we analysed under maximum parsimony using TNT 1.5 (weighted and unweighted analyses) and Bayesian inference using MrBayes v3.2.6 (standard, gamma and variation). The resulting topologies were then analysed to generate comprehensive higher-level phylogenetic hypotheses of teleosauroids and shed light on species-level interrelationships within the clade. The results from our parsimony and Bayesian analyses are largely consistent. Two large subclades within Teleosauroidea are recovered, and they are morphologically, ecologically and biogeographically distinct from one another. Based on comparative anatomical and phylogenetic results, we propose the following major taxonomic revisions to Teleosauroidea: (1) redefining Teleosauridae; (2) introducing one new family and three new subfamilies; (3) the resurrection of three historical genera; and (4) erecting seven new generic names and one new species name. The phylogeny infers that the Laurasian subclade was more phenotypically plastic overall than the Sub-Boreal-Gondwanan subclade. The proposed phylogeny shows that teleosauroids were more diverse than previously thought, in terms of morphology, ecology, dispersal and abundance, and that they represented some of the most successful crocodylomorphs during the Jurassic. creator: Michela M. Johnson creator: Mark T. Young creator: Stephen L. Brusatte uri: https://doi.org/10.7717/peerj.9808 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Johnson et al. title: Every hill has its leopard: patterns of space use by leopards (Panthera pardus) in a mixed use landscape in India link: https://peerj.com/articles/10072 last-modified: 2020-10-08 description: Understanding abundance and distribution of species is often necessary for wildlife conservation. However, elusive species such as the leopard (Panthera pardus) that have wide geographical distribution and typically low abundance pose a constant challenge to conservationists due to logistical and methodological constraints. Although leopard abundance has been estimated at the scale of protected areas or other smaller regions, reliable information describing leopard distribution over large spatial scales remains largely unavailable. Knowledge about space use by leopards within landscapes could help improve conservation management, reduce human-wildlife conflict, and also facilitate population status monitoring. We carried out occupancy surveys across c. 24,000 km2 in southern India in a landscape that consisted a mosaic of leopards’ natural habitats and highly human-dominated areas. We investigated the effects of key ecological and anthropogenic variables in determining leopard space use patterns. We addressed imperfect detections obtained using sign surveys conducted on spatially replicated transects within sampling units by modeling detection as a function of spatial auto-correlation and covariates. Our results show that the probability of site-use by leopards across the landscape varied between 0.02 (95% CI [0.01–0.09]) and 0.99 (95% CI [0.99–1.0]) across the study area. The best model (AIC weight = 0.97) showed that the probability of leopard space use was affected by the proportion of natural habitats and the presence of large wild prey in the sampling unit. Given that India is undergoing rapid modifications due to economic changes and demand for natural resources, we emphasize the need for landscape-based approach for conserving and monitoring leopards. We argue that leopards are an indicator of functional ecosystems represented by scrub, deciduous forest and rocky outcrops that do not always get prioritized for conservation, unlike densely forested habitats. Similarly, conservation of natural large wild prey, especially outside the protected area system, should assume greater importance, which could also have a positive impact on reducing human-leopard conflict. creator: Sanjay Gubbi creator: Koustubh Sharma creator: Vijaya Kumara uri: https://doi.org/10.7717/peerj.10072 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Gubbi et al. title: The challenge of managing the commercial harvesting of the sea urchin Paracentrotus lividus: advanced approaches are required link: https://peerj.com/articles/10093 last-modified: 2020-10-08 description: Sea urchins act as a keystone herbivore in marine coastal ecosystems, regulating macrophyte density, which offers refuge for multiple species. In the Mediterranean Sea, both the sea urchin Paracentrotus lividus and fish preying on it are highly valuable target species for artisanal fisheries. As a consequence of the interactions between fish, sea urchins and macrophyte, fishing leads to trophic disorders with detrimental consequences for biodiversity and fisheries. In Sardinia (Western Mediterranean Sea), regulations for sea urchin harvesting have been in place since the mid 90s. However, given the important ecological role of P. lividus, the single-species fishery management may fail to take into account important ecosystem interactions. Hence, a deeper understanding of population dynamics, their dependance on environmental constraints and multispecies interactions may help to achieve long-term sustainable use of this resource. This work aims to highlight how sea urchin population structure varies spatially in relation to local environmental constraints and species interactions, with implications for their management. The study area (Sinis Peninsula, West Sardinia, Italy) that includes a Marine Reserve was divided into five sectors. These display combinations of the environmental constraints influencing sea urchin population dynamics, namely type of habitat (calcareous rock, granite, basalt, patchy and continuous meadows of Posidonia oceanica), average bottom current speed and predatory fish abundance. Size-frequency distribution of sea urchins under commercial size (<5 cm diameter size) assessed during the period from 2004 to 2007, before the population collapse in 2010, were compared for sectors and types of habitat. Specific correlations between recruits (0–1 cm diameter size) and bottom current speeds and between middle-sized sea urchins (2–5 cm diameter size) and predatory fish abundance were assessed. Parameters representing habitat spatial configuration (patch density, perimeter-to-area ratio, mean patch size, largest patch index, interspersion/juxtaposition index) were calculated and their influence on sea urchin density assessed. The density of sea urchins under commercial size was significantly higher in calcareous rock and was positively and significantly influenced by the density and average size of the rocky habitat patches. Recruits were significantly abundant in rocky habitats, while they were almost absent in P. oceanica meadows. The density of middle-sized sea urchins was more abundant in calcareous rock than in basalt, granite or P. oceanica. High densities of recruits resulted significantly correlated to low values of average bottom current speed, while a negative trend between the abundance of middle-sized sea urchins and predatory fish was found. Our results point out the need to account for the environmental constraints influencing local sea urchin density in fisheries management. creator: Simone Farina creator: Maura Baroli creator: Roberto Brundu creator: Alessandro Conforti creator: Andrea Cucco creator: Giovanni De Falco creator: Ivan Guala creator: Stefano Guerzoni creator: Giorgio Massaro creator: Giovanni Quattrocchi creator: Giovanni Romagnoni creator: Walter Brambilla uri: https://doi.org/10.7717/peerj.10093 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Farina et al. title: The advanced lung cancer inflammation index predicts short and long-term outcomes in patients with colorectal cancer following surgical resection: a retrospective study link: https://peerj.com/articles/10100 last-modified: 2020-10-08 description: Background and PurposeSeveral studies have proposed that the advanced lung cancer inflammation index (ALI), a new inflammation-related index, can be used for the prognosis assessment of various malignancies. However, few studies have reported its prognostic value in colorectal cancer (CRC). Therefore, this study explored the relationship between ALI and outcomes in CRC patients.MethodsA total of 662 CRC patients who underwent surgery between 2012 and 2014 were included. The ALI was defined as: body mass index × serum albumin/neutrophil to lymphocyte ratio. The X-tile program identified the optimal cut-off value of ALI. Logistic regression analyses determined factors affecting postoperative complications. The Kaplan–Meier method and Cox proportional hazards analyses evaluated potential prognostic factors.ResultsThe optimal cut-off of ALI in males and females were 31.6 and 24.4, respectively. Low-ALI was an independent risk factor for postoperative complications in CRC patients (odds ratio: 1.933, 95% CI [1.283–2.911], p = 0.002). Low-ALI groups also had significantly lower progression-free survival (PFS) and overall survival (OS), when compared with the high-ALI group, especially at advance tumor stages. Using multivariate analysis, ALI was determined as an independent prognostic factor for PFS (hazard ratio: 1.372, 95% CI [1.060–1.777], p = 0.016) and OS (hazard ratio: 1.453, 95% confidence interval: 1.113–1.898, p = 0.006).ConclusionALI is an independent predictor of short and long-term outcomes in CRC patients, especially at advance tumor stages. The ALI-based nomograms can provide accurate and individualized prediction of postoperative complication risk and survival for CRC patients. creator: Hailun Xie creator: Shizhen Huang creator: Guanghui Yuan creator: Jiaan Kuang creator: Ling Yan creator: Lishuang Wei creator: Shuangyi Tang creator: Jialiang Gan uri: https://doi.org/10.7717/peerj.10100 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Xie et al. title: Exploration of ovine milk whey proteome during postnatal development using an iTRAQ approach link: https://peerj.com/articles/10105 last-modified: 2020-10-08 description: BackgroundOvine milk is a rich source of bioactive proteins that supports the early growth and development of the newborn lambs. A large number of researches had targeted to the identification of ovine milk fat globule membrane proteins (MFGMPs), caseins (CNs), mastitis milk proteins in past years, but the dynamic change tendency of milk whey proteins during postnatal development has received limited attention. This research aimed to investigate the dynamic changes of ovine milk whey proteins after delivery, and explore the functions of whey proteins on early development of the newborns.MethodsIn this research, Hu sheep milk samples were collected from six individuals by manual milking manner, at 0 d, 3 d, 7 d, 14 d, 28 d and 56 d after delivery, respectively. The milk whey proteins were identified and quantified by the isobaric tag for relative and absolute quantification (iTRAQ) coupled with liquid chromatography (LC)-electrospray ionization (ESI) tandem MS (MS/MS) methods. In addition, biological functions of differentially expressed proteins (DEPs) were annotated by Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis.ResultsA total of 310 proteins were identified , of which 121 were differentially expressed. In detail, 30 (10 up-regulated and 20 down-regulated), 22 (11 up-regulated and 11 down-regulated), 11 (four up-regulated and seven down-regulated), 11 (eight up-regulated and three down-regulated), 10 (six up-regulated and four down-regulated) DEPs were identified in 3 d vs. 0 d, 7 d vs. 3 d, 14 d vs. 7 d, 28 d vs. 14 d, 56 d vs. 28 d comparison groups, respectively. The GO annotation analysis revealed that biological process principally involved metabolic and biological regulation, the major cellular location were organelle, cell and extracellular region, and the mainly molecular function were binding and catalytic activity. Circadian rhythm, fatty acid biosynthesis and African trypanosomiasis were enriched by KEGG annotation analysis.ConclusionThe study reveals a comprehensive understanding of Hu sheep milk proteome, suggesting whey proteins change dramatically in early development of newborn lambs, which provide a potential guidance for early weaning of lambs. creator: Xueying Zhang creator: Fadi Li creator: Fang Qin creator: Wanhong Li creator: Xiangpeng Yue uri: https://doi.org/10.7717/peerj.10105 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: Isolation, growth, and nitrogen fixation rates of the Hemiaulus-Richelia (diatom-cyanobacterium) symbiosis in culture link: https://peerj.com/articles/10115 last-modified: 2020-10-08 description: Nitrogen fixers (diazotrophs) are often an important nitrogen source to phytoplankton nutrient budgets in N-limited marine environments. Diazotrophic symbioses between cyanobacteria and diatoms can dominate nitrogen-fixation regionally, particularly in major river plumes and in open ocean mesoscale blooms. This study reports the successful isolation and growth in monocultures of multiple strains of a diatom-cyanobacteria symbiosis from the Gulf of Mexico using a modified artificial seawater medium. We document the influence of light and nutrients on nitrogen fixation and growth rates of the host diatom Hemiaulus hauckii Grunow together with its diazotrophic endosymbiont Richelia intracellularis Schmidt, as well as less complete results on the Hemiaulus membranaceus-R. intracellularis symbiosis. The symbioses rates reported here are for the joint diatom-cyanobacteria unit. Symbiont diazotrophy was sufficient to support both the host diatom and cyanobacteria symbionts, and the entire symbiosis replicated and grew without added nitrogen. Maximum growth rates of multiple strains of H. hauckii symbioses in N-free medium with N2 as the sole N source were 0.74–0.93 div d−1. Growth rates followed light saturation kinetics in H. hauckii symbioses with a growth compensation light intensity (EC) of 7–16 µmol m−2s−1and saturation light level (EK) of 84–110 µmol m−2s−1. Nitrogen fixation rates by the symbiont while within the host followed a diel pattern where rates increased from near-zero in the scotophase to a maximum 4–6 h into the photophase. At the onset of the scotophase, nitrogen-fixation rates declined over several hours to near-zero values. Nitrogen fixation also exhibited light saturation kinetics. Maximum N2 fixation rates (84 fmol N2 heterocyst−1h−1) in low light adapted cultures (50 µmol m−2s−1) were approximately 40–50% of rates (144–154 fmol N2 heterocyst−1h−1) in high light (150 and 200 µmol m−2s−1) adapted cultures. Maximum laboratory N2 fixation rates were ~6 to 8-fold higher than literature-derived field rates of the H. hauckii symbiosis. In contrast to published results on the Rhizosolenia-Richelia symbiosis, the H. hauckii symbiosis did not use nitrate when added, although ammonium was consumed by the H. hauckii symbiosis. Symbiont-free host cell cultures could not be established; however, a symbiont-free H. hauckii strain was isolated directly from the field and grown on a nitrate-based medium that would not support DDA growth. Our observations together with literature reports raise the possibility that the asymbiotic H. hauckii are lines distinct from an obligately symbiotic H. hauckii line. While brief descriptions of successful culture isolation have been published, this report provides the first detailed description of the approaches, handling, and methodologies used for successful culture of this marine symbiosis. These techniques should permit a more widespread laboratory availability of these important marine symbioses. creator: Amy E. Pyle creator: Allison M. Johnson creator: Tracy A. Villareal uri: https://doi.org/10.7717/peerj.10115 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Pyle et al. title: Overgrazing-induced legacy effects may permit Leymus chinensis to cope with herbivory link: https://peerj.com/articles/10116 last-modified: 2020-10-08 description: There is growing evidence that herbivory-induced legacy effects permit plants to cope with herbivory. However, herbivory-induced defense strategies in plants against grazing mammals have received little attention. To further understand the grazing-induced legacy effects on plants, we conducted a greenhouse experiment with Leymus chinensis experiencing different grazing histories. We focused on grazing-induced legacy effects on above-ground spatial avoidance and below-ground biomass allocation. Our results showed that L. chinensis collected from the continuous overgrazing plot (OG) exhibited higher performance under simulated grazing in terms of growth, cloning and colonizing ability than those collected from the 35-year no-grazing plot (NG). The enhanced adaptability of OG was attributed to increased above-ground spatial avoidance, which was mediated by larger leaf angle and shorter height (reduced vertical height and increased leaf angle contributed to the above-ground spatial avoidance at a lower herbivory stubble height, while reduced tiller natural height contributed to above-ground spatial avoidance at a higher herbivory stubble height). Contrary to our prediction, OG pre-allocated less biomass to the rhizome, which does not benefit the herbivory tolerance and avoidance of L. chinensis; however, this also may reflect a tolerance strategy where reduced allocation to rhizomes is associated with increased production of ramets. creator: Fenghui Guo creator: Xiliang Li creator: Saheed Olaide Jimoh creator: Yong Ding creator: Yong Zhang creator: Shangli Shi creator: Xiangyang Hou uri: https://doi.org/10.7717/peerj.10116 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Guo et al. title: Investigation for a multi-silique trait in Brassica napus by alternative splicing analysis link: https://peerj.com/articles/10135 last-modified: 2020-10-08 description: BackgroundFlower and fruit development are vital stages of the angiosperm lifecycle. We previously investigated the multi-silique trait in the rapeseed (Brassica napus) line zws-ms on a genomic and transcriptomic level, leading to the identification of two genomic regions and several candidate genes associated with this trait. However, some events on the transcriptome level, like alternative splicing, were poorly understood.MethodsPlants from zws-ms and its near-isogenic line (NIL) zws-217 were both grown in Xindu with normal conditions and a colder area Ma’erkang. Buds from the two lines were sampled and RNA was isolated to perform the transcriptomic sequencing. The numbers and types of alternative splicing (AS) events from the two lines were counted and classified. Genes with AS events and expressed differentially between the two lines, as well as genes with AS events which occurred in only one line were emphasized. Their annotations were further studied.ResultsFrom the plants in Xindu District, an average of 205,496 AS events, which could be sorted into five AS types, were identified. zws-ms and zws-217 shared highly similar ratios of each AS type: The alternative 5′ and 3′ splice site types were the most common, while the exon skipping type was observed least often. Eleven differentially expressed AS genes were identified, of which four were upregulated and seven were downregulated in zws-ms. Their annotations implied that five of these genes were directly associated with the multi-silique trait. While samples from colder area Ma’erkang generated generally reduced number of each type of AS events except for Intron Retention; but the number of differentially expressed AS genes increased significantly. Further analysis found that among the 11 differentially expressed AS genes from Xindu, three of them maintained the same expression models, while the other eight genes did not show significant difference between the two lines in expression level. Additionally, the 205 line-specific expressed AS genes were analyzed, of which 187 could be annotated, and two were considered to be important.DiscussionThis study provides new insights into the molecular mechanism of the agronomically important multi-silique trait in rapeseed on the transcriptome level and screens out some environment-responding candidate genes. creator: Liang Chai creator: Jinfang Zhang creator: Haojie Li creator: Benchuan Zheng creator: Jun Jiang creator: Cheng Cui creator: Liangcai Jiang uri: https://doi.org/10.7717/peerj.10135 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chai et al. title: Adequacy of clinical information in X-ray referrals for traumatic ankle injury with reference to the Ottawa Ankle Rules—a retrospective clinical audit link: https://peerj.com/articles/10152 last-modified: 2020-10-08 description: Study ObjectiveTo assess the adequacy of clinical information with reference to the Ottawa Ankle Rules (OAR) in X-ray referrals for adults with traumatic ankle injury in the ED of a South Australian tertiary hospital and report upon referring trends between emergency department clinicians.MethodsA retrospective clinical audit of adult ankle X-ray referrals in the emergency department was conducted. Eligible referrals were screened for their adherence to the OAR, patient details, clinical history and referrer. A logistic regression was used to determine the influence of these factors on the likelihood of being referred for X-rays despite not meeting the OAR criteria. Sensitivity, specificity, positive and negative likelihood ratios and their associated confidence intervals were calculated to assess the diagnostic accuracy of the OAR for those referred.ResultsOut of the 262 eligible referrals, 163 were deemed to have met the criteria for the OAR. Physiotherapists showed the highest OAR compliance of 77.3% and were the most accurate in their use of the rules, with a sensitivity of 0.86. Medical officers, registrars and interns were 2.5 times more likely to still refer a patient for X-ray if they did not meet the OAR criteria, compared to physiotherapists as the baseline. Patient age, duration of injury etc. were not significantly associated with likelihood of referral (even when they did not meet OAR criteria). The overall sensitivity, specificity, positive and negative likelihood ratios of the OAR were 0.59 (95% CI [0.47–0.71]), 0.37 (95% CI [0.30–0.44]), 0.93 (95% CI [0.76–1.16]) and 1.10 (95% CI [0.82–1.48]) respectively.ConclusionThe results of this audit demonstrated poor sensitivity and moderate compliance by referrers with the rule. Reasonable evidence exists for the implementation of individual and/or institutional-based change strategies to improve clinician compliance and accuracy with use of the OAR. creator: Yolanda E. Gomes creator: Minh Chau creator: Helen A. Banwell creator: Josephine Davies creator: Ryan S. Causby uri: https://doi.org/10.7717/peerj.10152 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Gomes et al. title: Coupled exuviae of the Ordovician Ovalocephalus (Pliomeridae, Trilobita) in South China and its behavioral implications link: https://peerj.com/articles/10166 last-modified: 2020-10-08 description: Ecdysis was a vital process during the lives of trilobites. In addition to preserving the morphological changes in trilobite ontogeny, the preservation of its action often captured interesting behavioral information. Abundant exuviae of Ovalocephalus tetrasulcatus are preserved in the Ordovician strata in central Hubei, China, and some of them are arranged with two or three together end to end or superimposed. The preserved patterns and burial conditions indicate that these specimens were caused by the active behavior of trilobites. It is speculated that these exuvial clusters were formed by two or three trilobites in line to molt; that is, after one trilobite finished molting, other trilobites molted in front of, behind, or overlying the previously molted shells. This ecdysis strategy is interpreted as related to the postulated herding behavior of some trilobites, representing a behavioral response of the trilobites to choose a nearby safe zone during some risky life activities. creator: Ruiwen Zong uri: https://doi.org/10.7717/peerj.10166 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zong title: A taxonomic review of the genus Astrocladus (Echinodermata, Ophiuroidea, Euryalida, Gorgonocephalidae) from Japanese coastal waters link: https://peerj.com/articles/9836 last-modified: 2020-10-07 description: Japanese species of the genus Astrocladus (Echinodermata, Ophiuroidea, Euryalida, Gorgonocephalidae) are reviewed. Astrocladus coniferus recently has two junior synonyms, A. dofleini Döderlein and A. pardalis Döderlein, however, status of these species has long been questioned. These species concepts have not been reviewed in recent years and no molecular phylogenetic analyses have been performed. Observations of the lectotype of A. coniferus, as well as the lectotype and four paralectotypes of A. dofleini and the holotype of A. pardalis have revealed that A. coniferus and A. pardalis are conspecific and morphologically distinguishable from A. dofleini. Astrocladus coniferus and A. dofleini are supported as distinct species by our molecular data. Additionally, we re-describe A. exiguus and A. annulatus, based on recently collected specimens and the holotype. We conclude that four species, A. annulatus, A. coniferus, A. dofleini, and A. exiguus are present in Japanese waters. creator: Masanori Okanishi creator: Hisanori Kohtsuka creator: Toshihiko Fujita uri: https://doi.org/10.7717/peerj.9836 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Okanishi et al. title: Lifestyle choices of Brazilian college students link: https://peerj.com/articles/9830 last-modified: 2020-10-07 description: BackgroundLifestyle choices reflect the beliefs that individuals attribute to aspects of life. This construct can be assessed with the Individual Lifestyle Profile (PEVI) questionnaire, which measures elements of Nutrition, Physical Activity, Preventive Behaviors, Social Relationships and Stress Management.ObjectiveThe objective of this study was to estimate the psychometric properties of the PEVI applied to a sample of Brazilian university students, identifying the prevalence of each lifestyle component according to participants’ age, sex, weight status, course area/field and economic stratum and to estimate the contribution of these characteristics on physical and psychological lifestyle.MethodsThe PEVI data was analyzed by confirmatory factor analysis, using the indexes chi-square per degrees of freedom ratio (χ2/df), Comparative Fit Index (CFI), Tucker-Lewis Index (TLI) and Root Mean Square Error of Approximation (RMSEA). First-order and second-order models (physical and psychological lifestyle) were tested. Prevalences of lifestyle components were calculated and compared by participants’ age, sex, weight status, course area/field and economic stratum. A hypothetical causal structural model was elaborated to investigate the impact of sample characteristics on physical and psychological lifestyles. This model was evaluated considering the global fit to the data (χ2/df, CFI, TLI and RMSEA) and the hypothetical causal trajectories (β) (α = 5%).ResultsA sample of 1,303 students was used. The mean age was 20.9 (standard deviation = 2.8) years, 66.8% of participants were females, 63.4% had weights in the normal range and 73.7% were students of the social and exact sciences. The PEVI data showed an adequate fit for both the first- (χ2/df = 2.03, CFI = 0.98; TLI = 0.97; RMSEA = 0.04) and second-order (χ2/df = 2.25; CFI = 0.97; TLI = 0.97; RMSEA = 0.04) models. There was a higher prevalence of unfavorable physical and psychological lifestyle choices among females, among underweight and obese individuals, in older students and in those with lower economic strata. Moreover, negative behaviors in physical lifestyle were more prevalent in students from human/social/exact sciences and worse psychological lifestyle was observed among health sciences students. These results were confirmed by a structural model.ConclusionThe PEVI data presented validity and reliability. Negative lifestyle choices had high prevalence among students. Moreover, individual characteristics had different impact on physical and psychological lifestyle choices. creator: Bianca G. Martins creator: João Marôco creator: Mauro V.G. Barros creator: Juliana A.D.B. Campos uri: https://doi.org/10.7717/peerj.9830 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Martins et al. title: Effects of taphonomic deformation on geometric morphometric analysis of fossils: a study using the dicynodont Diictodon feliceps (Therapsida, Anomodontia) link: https://peerj.com/articles/9925 last-modified: 2020-10-07 description: Taphonomic deformation, the distortion of fossils as a result of geological processes, poses problems for the use of geometric morphometrics in addressing paleobiological questions. Signal from biological variation, such as ontogenetic trends and sexual dimorphism, may be lost if variation from deformation is too high. Here, we investigate the effects of taphonomic deformation on geometric morphometric analyses of the abundant, well known Permian therapsid Diictodon feliceps. Distorted Diictodon crania can be categorized into seven typical styles of deformation: lateral compression, dorsoventral compression, anteroposterior compression, “saddle-shape” deformation (localized collapse at cranial mid-length), anterodorsal shear, anteroventral shear, and right/left shear. In simulated morphometric datasets incorporating known “biological” signals and subjected to uniform shear, deformation was typically the main source of variance but accurate “biological” information could be recovered in most cases. However, in empirical datasets, not only was deformation the dominant source of variance, but little structure associated with allometry and sexual dimorphism was apparent, suggesting that the more varied deformation styles suffered by actual fossils overprint biological variation. In a principal component analysis of all anomodont therapsids, deformed Diictodon specimens exhibit significant dispersion around the “true” position of this taxon in morphospace based on undistorted specimens. The overall variance associated with deformation for Anomodontia as a whole is minor, and the major axes of variation in the study sample show a strong phylogenetic signal instead. Although extremely problematic for studying variation in fossil taxa at lower taxonomic levels, the cumulative effects of deformation in this study are shown to be random, and inclusion of deformed specimens in higher-level analyses of morphological disparity are warranted. Mean morphologies of distorted specimens are found to approximate the morphology of undistorted specimens, so we recommend use of species-level means in higher-level analyses when possible. creator: Christian F. Kammerer creator: Michol Deutsch creator: Jacqueline K. Lungmus creator: Kenneth D. Angielczyk uri: https://doi.org/10.7717/peerj.9925 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Kammerer et al. title: Transcriptome analysis reveals potential immune function-related regulatory genes/pathways of female Lubo goat submandibular glands at different developmental stages link: https://peerj.com/articles/9947 last-modified: 2020-10-07 description: BackgroundThe submandibular glands, as major salivary glands, participate in rumen digestion in goats. Sialic acid, lysozyme, immunoglobulin A (IgA), lactoferrin and other biologically active substances secreted in the submandibular glands were reported in succession, which suggests that the submandibular gland may have immune functions in addition to participating in digestion. The aim of this study was to map the expression profile of differentially expressed genes (DEGs) at three different stages by transcriptome sequencing, screen immune-related genes and pathways by bioinformatics methods, and predict the immune function of submandibular glands at different developmental stages.MethodsNine submandibular gland tissue samples were collected from groups of 1-month-old kids, 12-month-old adolescent goats and 24-month-old adult goats (3 samples from each group), and high-throughput transcriptome sequencing was conducted on these samples. The DEGs among the three stages were screened and analysed. Key genes and signalling pathways were selected via protein-protein interaction (PPI) network analysis.ResultsThe results revealed 2,706, 2,525 and 52 DEGs between 1-month-old and 12-month-old goats, between 1-month-old and 24-month-old goats, and between 12-month-old and 24-month-old goats, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses indicated that most of the DEGs were enriched in immune- related GO terms and pathways. Based on functional enrichment analysis and network analysis, 10 genes (PTPRC, CD28, SELL, LCP2, MYC, LCK, ZAP70, ITGB2, SYK and CCR7), two signalling pathways (the T cell receptor signalling pathway and the NF-κβ signalling pathway) and eight GO terms (T cell receptor signalling pathway, neutrophil mediated immunity, B cell mediated immunity, regulation of alpha-beta T cell activation, positive regulation of T cell proliferation, regulation of leukocyte differentiation, positive regulation of antigen receptor-mediated signalling pathway, positive regulation of lymphocyte proliferation) that may play key roles in the immune functions of the goat submandibular glands at different developmental stages were identified. Moreover, we found that eight antibacterial peptide-encoding genes were downregulated in the tuberculosis and salivary secretion pathways, while all immunoglobulins were upregulated in 10 immune system pathways. These findings indicate that the submandibular glands may be important immunological organs during the growth process of goats and that the immune function of these glands gradually weakens with age up to 12 months but remains relatively stable after 12 months of age. Overall, this study will improve our understanding of transcriptional regulation related to goat submandibular gland immune function. creator: Aili Wang creator: Tianle Chao creator: Zhibin Ji creator: Rong Xuan creator: Shuang Liu creator: Maosen Guo creator: Guizhi Wang creator: Jianmin Wang uri: https://doi.org/10.7717/peerj.9947 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: SlMYB102 expression enhances low-temperature stress resistance in tomato plants link: https://peerj.com/articles/10059 last-modified: 2020-10-07 description: Herein, we identified the tomato SlMYB102 gene as a MYB family transcription factor of the R2R3-MYB subfamily. We additionally determined that the SlMYB102 promoter region contains photoresponsive, abiotic stress-responsive, and hormone-responsive regulatory elements, and we detected higher SlMYB102 expression in the reproductive organs of tomato than that in vegetative organs, with the expression being highest in ripe fruits and in roots. SlMYB102 expression was also shown to be cold-inducible. The protein encoded by SlMYB102 localized to the nucleus wherein it was found to mediate the transcriptional activation of target genes through its C-terminal domain. Overexpression of SlMYB102 in tomato plants conferred enhanced tolerance to cold stress. Under such cold stress conditions, we found that proline levels in the leaves of SlMYB102 overexpressing transgenic plants were higher than those in WT plants. In addition, S1MYB102 overexpression was associated with the enhanced expression of cold response genes including SlCBF1, SlCBF3, SlDREB1, SlDEB2, and SlICE1. We also found that the overexpression of SlMYB102 further enhanced the cold-induced upregulation of SlP5CS and SlAPX2. Taken together, these results suggest that SlMYB102 may be involved in the C-repeat binding transcription factor (CBF) and proline synthesis pathways, thereby improving tomato plant cold resistance. creator: Meiling Wang creator: Juan Hao creator: Xiuhua Chen creator: Xichun Zhang uri: https://doi.org/10.7717/peerj.10059 license: https://creativecommons.org/licenses/by-nc/4.0 rights: ©2020 Wang et al. title: Factors influencing unmet need for contraception amongst adolescent girls and women in Cambodia link: https://peerj.com/articles/10065 last-modified: 2020-10-07 description: BackgroundUnmet need is the gap between women’s need and their practice of using contraception. Unmet need for contraception in female adolescents and women in Cambodia is a public health concern which may lead to unintended pregnancies or abortions that can contribute to maternal morbidity and mortality.MethodsBronfenbrenner’s Social Ecological Model was used as a theoretical framework to analyze data from the 2014 Cambodian Demographic and Health Survey to ascertain demographic and social factors potentially associated with unmet need for contraception. Bivariate and weighted multiple logistic regression analyses with adjusted odds ratios (AOR) were conducted for 4,823 Cambodian, sexually active females aged 15–29 years.ResultsThe percentage of unmet need for contraception was 11.7%. At the individual level of the Social Ecological Model, there was an increased likelihood of unmet need in adolescent girls 15–19 years and women 20–24 years. Unmet need was decreased in currently employed women. At the microenvironment level, there was an increased likelihood of unmet need with the husband’s desire for more children and when the decision for a woman’s access to healthcare was made by someone else in the household. At the macroenvironment level, unmet need was decreased in women who could access a health facility near their residence to obtain medical care. There were no urban rural differences found in the Cambodian sample population.ConclusionUnmet need for contraception in Cambodian females adolescents and women is associated with younger age, unemployment and low personal autonomy for accessing healthcare but not with education or wealth status. There is a need to implement culturally appropriate reproductive and sexual health literacy programs to increase access to modern contraception and to raise women’s autonomy. creator: Farwa Rizvi creator: Joanne Williams creator: Steven Bowe creator: Elizabeth Hoban uri: https://doi.org/10.7717/peerj.10065 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Rizvi et al. title: Lactobacillus johnsonii BS15 improves intestinal environment against fluoride-induced memory impairment in mice—a study based on the gut–brain axis hypothesis link: https://peerj.com/articles/10125 last-modified: 2020-10-07 description: BackgroundExcessive fluoride can lead to chronic neurodegeneration characterized by neuron and myelin loss and memory dysfunction. The gut–brain axis hypothesis suggests that gut microbiota plays a crucial role in regulating brain function. Thus, using probiotics to adjust the gut microenvironment may be a potential therapy for mental diseases.MethodsMice in the prob group were administrated with Lactobacillus johnsonii BS15 for 28 days prior to and throughout a 70-day exposure to sodium fluoride. The drinking water of all groups (F and prob groups) except the control group were replaced by high-fluoride water (100 mg NaF/L) on day 28. Animals in each group were divided into two subsets: one underwent behavioral test, and the other was sacrificed for sampling. The mRNA expression level and protein content related to inflammatory reaction in the ileum and hippocampus were respectively detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA). The mRNA expression levels of proteins related to myelin structure, apoptosis, and memory in the hippocampus and tight junction proteins in the ileum were determined by RT-qPCR and/or immunohistochemistry. Gut permeability markers (D-lactate and diamine oxidase (DAO)) in the serum were also examined by ELISA.ResultsThe results showed that fluoride exposure induced a lower spontaneous exploration (P < 0.05) in T-maze test, which indicated an impairment of memory. Spontaneous exploration of BS15-treated mice was significantly higher (P < 0.05) than that in F group. Fluoride reduced (P < 0.05) levels of myelin structural protein (proteolipid protein) and neurogenesis-associated proteins (brain-derived neurotrophic factor and cAMP/Ca2+ responsive element-binding protein), induced disordered inflammatory cytokines (TNF-α, IFN-γ, and IL-6; P < 0.05), increased pro-apoptotic genes (caspase-3; P < 0.05), and decreased anti-apoptotic genes (Bcl-2; P < 0.05) in the hippocampus, of which the influences were reversed by BS15. BS15 treatment exerted significant preventive effects on reversing the gut inflammation induced by excessive fluoride intake by reducing (P < 0.05) the levels of pro-inflammatory cytokines (tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ)) and remarkably increasing (P < 0.05) the level of anti-inflammatory cytokines (IL-10). Moreover, the serum DAO activity and D-lactate concentration significantly increased by fluoride were also reduced (P < 0.05) by BS15. This result indicated the profitable effect of BS15 on gut permeability.ConclusionL. johnsonii BS15 intake could benefit the neuroinflammation and demyelination in the hippocampus by improving the gut environment and ameliorating fluorine-induced memory dysfunction. creator: Jinge Xin creator: Dong Zeng creator: Hesong Wang creator: Ning Sun creator: Abdul Khalique creator: Ying Zhao creator: Liqian Wu creator: Kangcheng Pan creator: Bo Jing creator: Xueqin Ni uri: https://doi.org/10.7717/peerj.10125 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Xin et al. title: CKS2 and RMI2 are two prognostic biomarkers of lung adenocarcinoma link: https://peerj.com/articles/10126 last-modified: 2020-10-07 description: BackgroundLung adenocarcinoma (ACA) is the most common subtype of non-small-cell lung cancer. About 70%–80% patients are diagnosed at an advanced stage; therefore, the survival rate is poor. It is urgent to discover accurate markers that can differentiate the late stages of lung ACA from the early stages. With the development of biochips, researchers are able to efficiently screen large amounts of biological analytes for multiple purposes.MethodsOur team downloaded GSE75037 and GSE32863 from the Gene Expression Omnibus (GEO) database. Next, we utilized GEO’s online tool, GEO2R, to analyze the differentially expressed genes (DEGs) between stage I and stage II–IV lung ACA. The using the Cytoscape software was used to analyze the DEGs and the protein-protein interaction (PPI) network was further constructed. The function of the DEGs were further analyzed by cBioPortal and Gene Expression Profiling Interactive Analysis (GEPIA) online tools. We validated these results in 72 pairs human samples.ResultsWe identified 109 co-DEGs, most of which were involved in either proliferation, S phase of mitotic cell cycle, regulation of exit from mitosis, DNA replication initiation, DNA replication, and chromosome segregation. Utilizing cBioPortal and University of California Santa Cruz databases, we further confirmed 35 hub genes. Two of these genes, encoding CDC28 protein kinase regulatory subunit 2 (CKS2) and RecQ-mediated genome instability 2 (RMI2), were upregulated in lung ACA compared with adjacent normal tissues. The Kaplan–Meier curves revealed upregulation of CKS2 and RMI2 are associated with worse survival. Using CMap analysis, we discovered 10 small molecular compounds that reversed the altered DEGs, the top five are phenoxybenzamine, adiphenine, resveratrol, and trifluoperazine. We also evaluated 72 pairs resected samples, results revealed that upregulation of CKS2 and RMI2 in lung ACA were associated with larger tumor size. Our results allow the deeper recognizing of the mechanisms of the progression of lung ACA, and may indicate potential therapeutic strategies for the therapy of lung ACA. creator: Dayong Xiao creator: Siyuan Dong creator: Shize Yang creator: Zhenghua Liu uri: https://doi.org/10.7717/peerj.10126 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Xiao et al. title: A digital mapping application for quantifying and displaying air temperatures at high spatiotemporal resolutions in near real-time across Australia link: https://peerj.com/articles/10106 last-modified: 2020-10-07 description: Surface air temperature (Ta) required for real-time environmental modelling applications should be spatially quantified to capture the nuances of local-scale climates. This study created near real-time air temperature maps at a high spatial resolution across Australia. This mapping is achieved using the thin plate spline interpolation in concert with a digital elevation model and ‘live’ recordings garnered from 534 telemetered Australian Bureau of Meteorology automatic weather station (AWS) sites. The interpolation was assessed using cross-validation analysis in a 1-year period using 30-min interval observation. This was then applied to a fully automated mapping system—based in the R programming language—to produce near real-time maps at sub-hourly intervals. The cross-validation analysis revealed broad similarities across the seasons with mean-absolute error ranging from 1.2 °C (autumn and summer) to 1.3 °C (winter and spring), and corresponding root-mean-square error in the range 1.6 °C to 1.7 °C. The R2 and concordance correlation coefficient (Pc ) values were also above 0.8 in each season indicating predictions were strongly correlated to the validation data. On an hourly basis, errors tended to be highest during the late afternoons in spring and summer from 3 pm to 6 pm, particularly for the coastal areas of Western Australia. The mapping system was trialled over a 21-day period from 1 June 2020 to 21 June 2020 with majority of maps completed within 28-min of AWS site observations being recorded. All outputs were displayed in a web mapping application to exemplify a real-time application of the outputs. This study found that the methods employed would be highly suited for similar applications requiring real-time processing and delivery of climate data at high spatiotemporal resolutions across a considerably large land mass. creator: Mathew Webb creator: Budiman Minasny uri: https://doi.org/10.7717/peerj.10106 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Webb and Minasny title: COVID-19: molecular and serological detection methods link: https://peerj.com/articles/10180 last-modified: 2020-10-07 description: Since COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was declared as a pandemic disease by the World Health Organization in early 2020, many countries, organizations and companies have tried to find the best way to diagnose the virus and contain its spreading. SARS-CoV-2 is a positive-sense single RNA (+ssRNA) coronavirus and mainly spreads through droplets, respiratory secretions, and direct contact. The early detection of the virus plays a central role in lowering COVID19 incidents and mortality rates. Thus, finding a simple, accurate, cheap and quick detection approach for SARS-CoV-2 at early stage of the viral infection is urgent and at high demand all around the world. The Food and Drug Administration and other health agencies have declared Emergency Use Authorization to develop diagnostic methods for COVID-19 and fulfill the demand. However, not all developed methods are appropriate and selecting a suitable method is challenging. Among all detection methods, rRT-PCR is the gold standard method. Unlike molecular methods, serological methods lack the ability of early detection with low accuracy. In this review, we summarized the current knowledge about COVID-19 detection methods aiming to highlight the advantages and disadvantages of molecular and serological methods. creator: Ahmed E. Dhamad creator: Muna A. Abdal Rhida uri: https://doi.org/10.7717/peerj.10180 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Dhamad and Abdal Rhida title: The highly rearranged mitochondrial genomes of three economically important scale insects and the mitochondrial phylogeny of Coccoidea (Hemiptera: Sternorrhyncha) link: https://peerj.com/articles/9932 last-modified: 2020-10-06 description: The mitochondrial genomes (mitogenomes) of scale insects are less known in comparison to other insects, which hinders the phylogenetic and evolutionary studies of Coccoidea and higher taxa. Herein, the complete mitogenomes of Unaspis yanonensis, Planococcus citri and Ceroplastes rubens were sequenced for Coccoidea. The 15,220-bp long mitogenome of U. yanonensis contained the typical set of 37 genes including 13 PCGs, 22 tRNA genes and two rRNA genes; the 15,549-bp long mitogenome of P. citri lacked the tRNA gene trnV; the 15,387-bp long mitogenome of C. rubens exhibited several shortened PCGs and lacked five tRNA genes. The mitochondrial gene arrangement of the three mitogenomes was different from other scale insects and Drosophila yakuba. Most PCGs used standard ATN (ATA, ATT, ATC and ATG) start codons and complete TAN (TAA or TAG) termination codons. The ND4L had the highest evolutionary rate but COX1 and CYTB were the lowest. Most tRNA genes had cloverleaf secondary structures, whereas the reduction of dihydrouridine (DHU) arms and TψC arms were detected. Tandem repeats, stem-loop (SL) structures and poly-[TA]n stretch were found in the control regions (CRs) of the three mitogenomes. The phylogenetic analyses using Bayesian inference (BI) and maximum likelihood methods (ML) showed identical results, both supporting the inner relationship of Coccoidea as Coccidae + (Pseudococcidae + Diaspididae). creator: Hong-Ling Liu creator: Qing-Dong Chen creator: Song Chen creator: De-Qiang Pu creator: Zhi-Teng Chen creator: Yue-Yue Liu creator: Xu Liu uri: https://doi.org/10.7717/peerj.9932 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liu et al. title: Master regulator genes and their impact on major diseases link: https://peerj.com/articles/9952 last-modified: 2020-10-06 description: Master regulator genes (MRGs) have become a hot topic in recent decades. They not only affect the development of tissue and organ systems but also play a role in other signal pathways by regulating additional MRGs. Because a MRG can regulate the concurrent expression of several genes, its mutation often leads to major diseases. Moreover, the occurrence of many tumors and cardiovascular and nervous system diseases are closely related to MRG changes. With the development in omics technology, an increasing amount of investigations will be directed toward MRGs because their regulation involves all aspects of an organism’s development. This review focuses on the definition and classification of MRGs as well as their influence on disease regulation. creator: Wanwan Cai creator: Wanbang Zhou creator: Zhe Han creator: Junrong Lei creator: Jian Zhuang creator: Ping Zhu creator: Xiushan Wu creator: Wuzhou Yuan uri: https://doi.org/10.7717/peerj.9952 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Cai et al. title: Increased photosynthesis and grain yields in maize grown with less irrigation water combined with density adjustment in semiarid regions link: https://peerj.com/articles/9959 last-modified: 2020-10-06 description: In order to design a water-saving and high-yield maize planting model suitable for semiarid areas, we conducted trials by combining supplementary irrigation with different planting densities. Three planting densities (L: 52,500, M: 75,000, and H: 97,500 plants ha–1) and four supplementary irrigation modes (NI: no irrigation; IV: 375 m3 ha–1 during the 11-leaf stage; IS: 375 m3 ha–1 in the silking stage; and IVS: 375 m3 ha–1 during both stages) were tested. The irrigation treatments significantly increased the leaf relative water content, but the high planting density significantly decreased the relative water content during the silking and filling stages. After supplementary irrigation during the 11-leaf stage, IV and IVS significantly increased the photosynthetic capacity, but decreased the leaf water use efficiency. IS and IVS significantly increased the photosynthetic capacity after supplementary irrigation in the silking stage over two years. During the filling stage, IV, IS, and IVS increased the two-year average net photosynthetic rate by 17.0%, 27.2%, and 30.3%, respectively. The intercellular CO2 concentration increased as the density increased, whereas the stomatal conductance, transpiration rate, net photosynthetic rate, and leaf water use efficiency decreased, and the high planting density significantly reduced the leaf photosynthetic capacity. The highest grain yield was obtained using the IVS treatment under the medium planting density, but it did not differ significantly from that with the IS treatment. Furthermore, the IVS treatment used two times more water than the IS treatment. Thus, the medium planting density combined with supplementary irrigation during the silking stage was identified as a suitable water-saving planting model to improve the photosynthetic capacity and grain yield, and to cope with drought and water shortages in semiarid regions. creator: Donghua Liu creator: Qianmin Jia creator: Juan Li creator: Peng Zhang creator: Xiaolong Ren creator: Zhikuan Jia uri: https://doi.org/10.7717/peerj.9959 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liu et al. title: Phylogeographic analysis and species distribution modelling of the wood frog Batrachyla leptopus (Batrachylidae) reveal interglacial diversification in south western Patagonia link: https://peerj.com/articles/9980 last-modified: 2020-10-06 description: BackgroundThe evolutionary history of southern South American organisms has been strongly influenced by Pleistocene climate oscillations. Amphibians are good models to evaluate hypotheses about the influence of these climate cycles on population structure and diversification of the biota, because they are sensitive to environmental changes and have restricted dispersal capabilities. We test hypotheses regarding putative forest refugia and expansion events associated with past climatic changes in the wood frog Batrachyla leptopus distributed along ∼1,000 km of length including glaciated and non-glaciated areas in southwestern Patagonia.MethodsUsing three mitochondrial regions (D-loop, cyt b, and coI) and two nuclear loci (pomc and crybA1), we conducted multilocus phylogeographic analyses and species distribution modelling to gain insights of the evolutionary history of this species. Intraspecific genealogy was explored with maximum likelihood, Bayesian, and phylogenetic network approaches. Diversification time was assessed using molecular clock models in a Bayesian framework, and demographic scenarios were evaluated using approximate Bayesian computation (ABC) and extended Bayesian skyline plot (EBSP). Species distribution models (SDM) were reconstructed using climatic and geographic data.ResultsPopulation structure and genealogical analyses support the existence of four lineages distributed north to south, with moderate to high phylogenetic support (Bootstrap > 70%; BPP > 0.92). The diversification time of B. leptopus’ populations began at ∼0.107 mya. The divergence between A and B lineages would have occurred by the late Pleistocene, approximately 0.068 mya, and divergence between C and D lineages was approximately 0.065 mya. The ABC simulations indicate that lineages coalesced at two different time periods, suggesting the presence of at least two glacial refugia and a postglacial colonization route that may have generated two southern lineages (p = 0.93, type I error: <0.094, type II error: 0.134). EBSP, mismatch distribution and neutrality indexes suggest sudden population expansion at ∼0.02 mya for all lineages. SDM infers fragmented distributions of B. leptopus associated with Pleistocene glaciations. Although the present populations of B. leptopus are found in zones affected by the last glacial maximum (∼0.023 mya), our analyses recover an older history of interglacial diversification (0.107–0.019 mya). In addition, we hypothesize two glacial refugia and three interglacial colonization routes, one of which gave rise to two expanding lineages in the south. creator: José J. Nuñez creator: Elkin Y. Suárez-Villota creator: Camila A. Quercia creator: Angel P. Olivares creator: Jack W. Sites Jr uri: https://doi.org/10.7717/peerj.9980 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Nuñez et al. title: Light quality affects the proliferation of in vitro cultured plantlets of Camellia oleifera Huajin link: https://peerj.com/articles/10016 last-modified: 2020-10-06 description: BackgroundCamellia oleifera is an important oil-yielding woody plant native to China. Tea oil extracted from the seeds is rich in health-beneficial compounds. Huajin is a high-yielding elite variety of C. oleifera, with large fruits and remarkable resilience, widely cultivated in southern China; however, its seedling quality tends to be uneven. At present, techniques such as grafting, and cuttings are primarily adopted to propagate C. oleifera. These approaches are susceptible to environmental constraints owing to the long growth period, resulting in the lack of C. oleifera seedlings. Methods to make the cultivation more economical are warranted; this can be facilitated by tissue culture technology to provide good-quality seedlings in a short time.MethodsIn vitro cultured plantlets of C. oleifera Huajin were exposed to red light (RL), blue light (BL), red:blue light at a 4:1 ratio (R4:B1), and red:blue light at a 1:4 ratio (R1:B4); white light (WL) was used as the control treatment. To investigate the influence of light spectral quality on the proliferation coefficient, photosynthetic pigments, soluble proteins, plant height, leaf shape, Rubisco enzyme activity, and stomata and leaf anatomical features.ResultsThe highest proliferation coefficient was observed under combined red and blue (4:1) light. In addition, this treatment resulted in the second highest chlorophyll content, the thickest palisade and spongy tissues, and consequently, the thickest leaves. The same treatment resulted in the second highest stomatal density, albeit concomitantly with the smallest average stomatal length and width.DiscussionThese results indicate that high-quality propagation of Huajin shoots can be achieved by culturing the plants in vitro under a combination of red and blue (4:1) lights. Previous studies have shown that red and blue lights improve rooting and transplanting rates of tissue culture seedlings. Hence, future research should focus on the effect of light quality on rooting and transplanting of tissue culture plantlets of Huajin and its specific molecular mechanisms. creator: Chaoyin He creator: Yanling Zeng creator: Yuzhong Fu creator: Jiahao Wu creator: Qin Liang uri: https://doi.org/10.7717/peerj.10016 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 He et al. title: The tale of springs and streams: how different aquatic ecosystems impacted the mtDNA population structure of two riffle beetles in the Western Carpathians link: https://peerj.com/articles/10039 last-modified: 2020-10-06 description: The Western Carpathians are a particularly interesting part of the Carpathian Arc. According to recent molecular data upon aquatic and terrestrial taxa, this mountain area is an important biodiversity hotspot of Europe. Moreover, the W Carpathians include rich systems of karst springs inhabited by specific fauna, where molecular diversity and phylogeographic patterns are yet to be fully explored. Our study aims to compare population genetic structure and molecular diversity of two related and commonly co-occurring riffle beetles, Elmis aenea (PWJ Müller, 1806) and Limnius perrisi (Dufour, 1843) in the springs and streams of the W Carpathians using the mitochondrial DNA barcoding fragment of the cytochrome c oxidase subunit I gene (COI). The relatively stable thermal and chemical conditions of springs throughout unfavourable climatic settings make these highly specific lotic systems potentially ideal for a long-term survival of some aquatic biota. Populations of both elmid species were relatively homogeneous genetically, with a single dominant haplotype. However, we revealed that E. aenea significantly dominated in the springs, while L. perrisi preferred streams. Relative isolation of the springs and their stable conditions were reflected in significantly higher molecular diversity of the E. aenea population in comparison to L. perrisi. The results of Bayesian Skyline Plot analysis also indicated the exceptional position of springs regarding maintaining the population size of E. aenea. On the other hand, it seems that streams in the W Carpathians provide more effective dispersal channels for L. perrisi, whose population expanded much earlier compared to E. aenea. Present study points out that different demographic histories of these two closely related elmid species are manifested by their different habitat preference and molecular diversity. creator: Jana Bozáňová creator: Zuzana Čiamporová Zat’ovičová creator: Fedor Čiampor Jr creator: Tomasz Mamos creator: Michał Grabowski uri: https://doi.org/10.7717/peerj.10039 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Bozáňová et al. title: Neighbourhood socioeconomic characteristics and blood pressure among Jamaican youth: a pooled analysis of data from observational studies link: https://peerj.com/articles/10058 last-modified: 2020-10-06 description: IntroductionNeighbourhood characteristics are associated with several diseases, but few studies have investigated the association between neighbourhood and health in Jamaica. We evaluated the relationship between neighbourhood socioeconomic status (SES) and blood pressure (BP) among youth, 15–24 years old, in Jamaica.MethodsA pooled analysis was conducted using data from three studies (two national surveys and a birth cohort), conducted between 2005–2008, with individual level BP, anthropometric and demographic data, and household SES. Data on neighbourhood SES were obtained from the Mona Geo-Informatics Institute. Neighbourhood was defined using community boundaries from the Social Development Commission in Jamaica. Community characteristics (poverty, unemployment, dependency ratio, population density, house size, and proportion with tertiary education) were combined into SES scores using principal component analysis (PCA). Multivariable analyses were computed using mixed effects multilevel models.ResultsAnalyses included 2,556 participants (1,446 females; 1,110 males; mean age 17.9 years) from 306 communities. PCA yielded two neighbourhood SES variables; the first, PCA-SES1, loaded highly positive for tertiary education and larger house size (higher value = higher SES); while the second, PCA-SES2, loaded highly positive for unemployment and population density (higher value = lower SES). Among males, PCA-SES1 was inversely associated with systolic BP (β-1.48 [95%CI −2.11, −0.84] mmHg, p < 0.001, for each standard deviation unit increase in PCA-SES1 score) in multivariable model accounting for age, household SES, study, BMI, fasting glucose, physical activity and diet. PCA-SES1 was not significantly associated with systolic BP among females (β −0.48 [−1.62, 0.66], p = 0.410) in a similar model. Associations for PCA-SES2 was assessed using linear splines to account for non-linear effects. The were no significant associations between systolic BP and PCA-SES2 among males. Among females, higher PCA-SES2 (i.e. lower SES) was associated with higher systolic BP at spline 2 [z-score -1 to 0] (β4.09 [1.49, 6.69], p = 0.002), but with lower systolic BP at spline 3 [z-core 0 to 1] (β-2.81 [−5.04, −0.59], p = 0.013). There were no significant associations between diastolic BP and PCA-SES1, but PCA-SES2 showed non-linear associations with diastolic BP particularly among males.ConclusionHigher neighbourhood SES was inversely associated with systolic BP among male Jamaican youth; there were non-linear associations between neighbourhood SES and systolic BP among females and for diastolic BP for both males and females. creator: Trevor S. Ferguson creator: Novie O.M. Younger-Coleman creator: Jasneth Mullings creator: Damian Francis creator: Lisa-Gaye Greene creator: Parris Lyew-Ayee creator: Rainford Wilks uri: https://doi.org/10.7717/peerj.10058 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Ferguson et al. title: Validation and clinical application of a targeted next-generation sequencing gene panel for solid and hematologic malignancies link: https://peerj.com/articles/10069 last-modified: 2020-10-06 description: BackgroundNext-generation sequencing (NGS) is a high-throughput technology that has become widely integrated in molecular diagnostics laboratories. Among the large diversity of NGS-based panels, the Trusight Tumor 26 (TsT26) enables the detection of low-frequency variants across 26 genes using the MiSeq platform.MethodsWe describe the inter-laboratory validation and subsequent clinical application of the panel in 399 patients presenting a range of tumor types, including gastrointestinal (GI, 29%), hematologic (18%), lung (13%), gynecological and breast (8% each), among others.ResultsThe panel is highly accurate with a test sensitivity of 92%, and demonstrated high specificity and positive predictive values (95% and 96%, respectively). Sequencing testing was successful in two-thirds of patients, while the remaining third failed due to unsuccessful quality-control filtering. Most detected variants were observed in the TP53 (28%), KRAS (16%), APC (10%) and PIK3CA (8%) genes. Overall, 372 variants were identified, primarily distributed as missense (81%), stop gain (9%) and frameshift (7%) altered sequences and mostly reported as pathogenic (78%) and variants of uncertain significance (19%). Only 14% of patients received targeted treatment based on the variant determined by the panel. The variants most frequently observed in GI and lung tumors were: KRAS c.35G > A (p.G12D), c.35G > T (p.G12V) and c.34G > T (p.G12C).ConclusionsPrior panel validation allowed its use in the laboratory daily practice by providing several relevant and potentially targetable variants across multiple tumors. However, this study is limited by high sample inadequacy rate, raising doubts as to continuity in the clinical setting. creator: Iván Prieto-Potin creator: Nerea Carvajal creator: Jenifer Plaza-Sánchez creator: Rebeca Manso creator: Carmen Laura Aúz-Alexandre creator: Cristina Chamizo creator: Sandra Zazo creator: Almudena López-Sánchez creator: Socorro María Rodríguez-Pinilla creator: Laura Camacho creator: Raquel Longarón creator: Beatriz Bellosillo creator: Rosa Somoza creator: Javier Hernández-Losa creator: Víctor Manuel Fernández-Soria creator: Ricardo Ramos-Ruiz creator: Ion Cristóbal creator: Jesús García-Foncillas creator: Federico Rojo uri: https://doi.org/10.7717/peerj.10069 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Prieto-Potin et al. title: A noninvasive flexible conformal sensor for accurate real-time monitoring of local cerebral edema based on electromagnetic induction link: https://peerj.com/articles/10079 last-modified: 2020-10-06 description: Cerebral edema (CE) is a non-specific pathological swelling of the brain secondary to any type of neurological injury. The real-time monitoring of focal CE mostly found in early stage is of great significance to reduce mortality and disability. Magnetic Induction Phase Shift (MIPS) is expected to achieve non-invasive continuous monitoring of CE. However, most existing MIPS sensors are made of hard materials which makes it difficult to accurately retrieve CE information. In this article, we designed a conformal two-coil structure and a single-coil structure, and studied their sensitivity map using finite element method (FEM). After that, the conformal MIPS sensor that is preferable for local CE monitoring was fabricated by flexible printed circuit (FPC). Next, physical experiments were conducted to investigate its performance on different levels of simulated CE solution volume, measurement distance, and bending. Subsequently, 14 rabbits were chosen to establish CE model and another three rabbits were selected as controls. The 24-hour MIPS real-time monitoring experiments was carried out to verify that the feasibility. Results showed a gentler attenuation trend of the conformal two-coil structure, compared with the single-coil structure. In addition, the novel flexible conformal MIPS sensor has a characteristic of being robust to bending according to the physical experiments. The results of animal experiments showed that the sensor can be used for CE monitoring. It can be concluded that this flexible conformal MIPS sensor is desirable for local focusing measurement of CE and subsequent multidimensional information extraction for predicting model. Also, it enables a much more comfortable environment for long-time bedside monitoring. creator: Jingbo Chen creator: Gen Li creator: Mingsheng Chen creator: Gui Jin creator: Shuanglin Zhao creator: Zelin Bai creator: Jun Yang creator: Huayou Liang creator: Jia Xu creator: Jian Sun creator: Mingxin Qin uri: https://doi.org/10.7717/peerj.10079 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: Attention behaviours but not pain-related behaviours are modified by the presence of a twin in lambs undergoing castration by rubber ring link: https://peerj.com/articles/10081 last-modified: 2020-10-06 description: The social context of social species such as sheep can modify their physiological and behavioural responses to stressors, through social buffering and social facilitation. Social buffering can lead to amelioration of stress, while social facilitation can lead to stress responses in an observer animal in the presence of a conspecific in distress. The current study investigated twin lambs undergoing ring castration, grouped either homogeneously with a castrated lamb (actor), or heterogeneously with a non-castrated lamb (observer) to examine the impact of social grouping on behavioural responses. Each lamb was scored for two classes of behaviour: pain-related behaviours and postures that are putatively related to the response to castration; and attentional behaviours directed at its twin. Thus, each lamb participated in the experiment as an actor exhibiting pain-related behaviours and postures, and as an observer of its twin. When behaviours of lambs were assessed as actors, there was a significant (P < 0.05) effect of castration but no significant effect of social grouping on expression of pain-related behaviours. When behaviours of lambs were assessed as observers, homogeneous grouping of castrated lambs increased attention towards the other twin in comparison to non-castrated lambs grouped homogeneously or lambs grouped heterogeneously (P < 0.01). Non-castrated lambs grouped homogeneously demonstrated significantly lower numbers of head direction changes (P < 0.001) and lower number of ear posture changes (P < 0.05) than heterogeneously grouped or castrated lambs. This study indicates that social attention between twin lambs is not clearly dependent on pain status of the actor lamb. It suggests that in order for the observer lamb to provide significant attention to the actor lamb displaying pain-related behaviour, the observer lamb also needs to be experiencing pain concurrently. Furthermore, there is some evidence that the presence of pain-related behaviours can lead to increased attention to the surrounding environment in non-castrated lambs. Understanding the effect of concurrent experience and varying social context assists us to improve our understanding of results of other experiments on pain-related behavioural responses. creator: Andrew Inhyuk Cho creator: Caroline Lee creator: Alison Small uri: https://doi.org/10.7717/peerj.10081 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Cho et al. title: Mathematical achievement: the role of spatial and motor skills in 6–8 year-old children link: https://peerj.com/articles/10095 last-modified: 2020-10-06 description: Several studies have tried to establish the factors that underlie mathematical ability across development. Among them, spatial and motor abilities might play a relevant role, but no studies jointly contemplate both types of abilities to account for mathematical performance. The present study was designed to observe the roles of spatial and motor skills in mathematical performance. A total of 305 children aged between 6 and 8 years took part in this study. A generalized linear regression model with mathematical performance as a dependent variable was performed. Results revealed that Block design (as a visuospatial reasoning measure) accounted for mathematical performance, especially among 6- and 7-year-olds but not in 8-year-olds. After controlling for the effect of the block design, mental rotation and manual dexterity predicted mathematical performance. These findings highlight the role of underlying cognitive (spatial) and motor abilities in supporting mathematical achievement in primary school children. creator: Laura M. Fernández-Méndez creator: María José Contreras creator: Irene Cristina Mammarella creator: Tommaso Feraco creator: Chiara Meneghetti uri: https://doi.org/10.7717/peerj.10095 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Fernández-Méndez et al. title: Fecal microbiota in client-owned obese dogs changes after weight loss with a high-fiber-high-protein diet link: https://peerj.com/articles/9706 last-modified: 2020-10-05 description: BackgroundThe fecal microbiota from obese individuals can induce obesity in animal models. In addition, studies in humans, animal models and dogs have revealed that the fecal microbiota of subjects with obesity is different from that of lean subjects and changes after weight loss. However, the impact of weight loss on the fecal microbiota in dogs with obesity has not been fully characterized.MethodsIn this study, we used 16S rRNA gene sequencing to investigate the differences in the fecal microbiota of 20 pet dogs with obesity that underwent a weight loss program. The endpoint of the weight loss program was individually tailored to the ideal body weight of each dog. In addition, we evaluated the qPCR based Dysbiosis Index before and after weight loss.ResultsAfter weight loss, the fecal microbiota structure of dogs with obesity changed significantly (weightedANOSIM; p = 0.016, R = 0.073), showing an increase in bacterial richness (p = 0.007), evenness (p = 0.007) and the number of bacterial species (p = 0.007). The fecal microbiota composition of obese dogs after weight loss was characterized by a decrease in Firmicutes (92.3% to 78.2%, q = 0.001), and increase in Bacteroidetes (1.4% to 10.1%, q = 0.002) and Fusobacteria (1.6% to 6.2%, q = 0.040). The qPCR results revealed an overall decrease in the Dysbiosis Index, driven mostly due to a significant decrease in E. coli (p = 0.030), and increase in Fusobacterium spp. (p = 0.017).ConclusionThe changes observed in the fecal microbiota of dogs with obesity after weight loss with a weight loss diet rich in fiber and protein were in agreement with previous studies in humans, that reported an increase of bacterial biodiversity and a decrease of the ratio Firmicutes/Bacteroidetes. creator: Sandra Bermudez Sanchez creator: Rachel Pilla creator: Benjamin Sarawichitr creator: Alessandro Gramenzi creator: Fulvio Marsilio creator: Joerg M. Steiner creator: Jonathan A. Lidbury creator: Georgiana R.T. Woods creator: Alexander J. German creator: Jan S. Suchodolski uri: https://doi.org/10.7717/peerj.9706 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Bermudez Sanchez et al. title: Elevation of MPF and MAPK gene expression, GSH content and mitochondrial distribution quality induced by melatonin promotes porcine oocyte maturation and development in vitro link: https://peerj.com/articles/9913 last-modified: 2020-10-05 description: The MPF and MAPK genes play crucial roles during oocyte maturation processes. However, the pattern of MPF and MAPK gene expression induced by melatonin (MT) and its correlation to oocyte maturation quality during the process of porcine oocyte maturation in vitro remains unexplored. To unravel it, in this study, we cultured the porcine oocytes in maturation medium supplemented with 0, 10−6, 10−9, and 10−12 mol/L melatonin. Later, we analyzed the MPF and MAPK gene expression levels by RT-PCR and determined the maturation index (survival and maturation rate of oocytes). The GSH content in the single oocyte, and cytoplasmic mitochondrial maturation distribution after porcine oocyte maturation in vitro was also evaluated. We also assessed the effects of these changes on parthenogenetic embryonic developmental potential. The oocytes cultured with 10−9mol/L melatonin concentration showed higher oocyte maturation rate, and MPF and MAPK genes expression levels along with better mitochondrial distribution than the 0, 10−6, and 10−12 mol/L melatonin concentrations (p < 0.05). No significant difference was observed in the survival rates when the oocytes were cultured with different melatonin concentrations. The expression of the MPF gene in the oocytes cultured with 10−6 mol/L melatonin was higher than with 10−12 and 0 mol/L melatonin, and the expression of the MAPK gene in 10−6 and 10−12 group was higher than the control (p < 0.05). As far as the embryonic developmental potential is concerned, the cleavage and blastocyst rate of oocytes cultured with 10−6 and 10−9 mol/L melatonin was significantly higher than the 10−12 mol/L melatonin and control. In conclusion, 10−9–10−6 mol/L melatonin significantly induced the MPF and MAPK gene expression; besides, it could also be correlated with GSH content of single oocyte, mitochondrial maturation distribution, and the first polar body expulsion. These changes were also found to be associated with parthenogenetic embryo developmental potential in vitro. creator: Zimo Zhao creator: Ling Yang creator: Dan Zhang creator: Zi Zheng creator: Ning Li creator: Qianjun Li creator: Maosheng Cui uri: https://doi.org/10.7717/peerj.9913 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhao et al. title: Comparison of pediatric scoring systems for mortality in septic patients and the impact of missing information on their predictive power: a retrospective analysis link: https://peerj.com/articles/9993 last-modified: 2020-10-05 description: BackgroundScores can assess the severity and course of disease and predict outcome in an objective manner. This information is needed for proper risk assessment and stratification. Furthermore, scoring systems support optimal patient care, resource management and are gaining in importance in terms of artificial intelligence.ObjectiveThis study evaluated and compared the prognostic ability of various common pediatric scoring systems (PRISM, PRISM III, PRISM IV, PIM, PIM2, PIM3, PELOD, PELOD 2) in order to determine which is the most applicable score for pediatric sepsis patients in terms of timing of disease survey and insensitivity to missing data.MethodsWe retrospectively examined data from 398 patients under 18 years of age, who were diagnosed with sepsis. Scores were assessed at ICU admission and re-evaluated on the day of peak C-reactive protein. The scores were compared for their ability to predict mortality in this specific patient population and for their impairment due to missing data.ResultsPIM (AUC 0.76 (0.68–0.76)), PIM2 (AUC 0.78 (0.72–0.78)) and PIM3 (AUC 0.76 (0.68–0.76)) scores together with PRSIM III (AUC 0.75 (0.68–0.75)) and PELOD 2 (AUC 0.75 (0.66–0.75)) are the most suitable scores for determining patient prognosis at ICU admission. Once sepsis is pronounced, PELOD 2 (AUC 0.84 (0.77–0.91)) and PRISM IV (AUC 0.8 (0.72–0.88)) become significantly better in their performance and count among the best prognostic scores for use at this time together with PRISM III (AUC 0.81 (0.73–0.89)). PELOD 2 is good for monitoring and, like the PIM scores, is also largely insensitive to missing values.ConclusionOverall, PIM scores show comparatively good performance, are stable as far as timing of the disease survey is concerned, and they are also relatively stable in terms of missing parameters. PELOD 2 is best suitable for monitoring clinical course. creator: Christian Niederwanger creator: Thomas Varga creator: Tobias Hell creator: Daniel Stuerzel creator: Jennifer Prem creator: Magdalena Gassner creator: Franziska Rickmann creator: Christina Schoner creator: Daniela Hainz creator: Gerard Cortina creator: Benjamin Hetzer creator: Benedikt Treml creator: Mirjam Bachler uri: https://doi.org/10.7717/peerj.9993 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Niederwanger et al. title: Immune cell landscape analysis reveals prognostic immune cells and its potential mechanism in squamous cell lung carcinoma link: https://peerj.com/articles/9996 last-modified: 2020-10-05 description: BackgroundSquamous cell lung carcinoma (LUSC) was closely associated with smoking which was known to have a distant immunosuppression effect. In this study, we aimed to explore the relationship between immune cells and clinical outcomes of LUSC patients with smoking history.MethodsThe immune cell infiltration and RNA expression profiles of LUSC patients were collected from The Cancer Genome Atlas (TCGA). Then, the correlation between immune cell infiltration and clinical characteristics was explored. According to the level of immune cell infiltration, LUSC patients with smoking history were divided into high or low group to screen the differentially expressed lncRNAs and mRNAs. The prediction of target genes was performed by miRanda. Finally, the prognostic value of a certain signature was confirmed in an independent dataset.ResultsHigher abundance of tumor-infiltrating T follicular helper (Tfh) cells together with a lower abundance of resting memory CD4 T cells had been found in LUSC current reformed smokers for ≤15 years and current smoking patients. Moreover, Tfh cell infiltration was not only associated with better overall survival (OS) but also varied from different degrees of TNM stage. Low expression of lncRNA PWRN1 and its potential regulating genes DMRTB1, PIRT, APOBEC1, and ZPBP2 were associated with better OS. Combining PWRN1 and four regulating genes as a signature, patients with higher-level expression of the signature had shorter survival time in not only the TCGA but also in the GEO dataset.ConclusionsIt was found that Tfh cells presented higher infiltration in LUSC current reformed smokers for ≤15 years and current smokers, while resting memory CD4 T cells had lower infiltration. The signature consisting of PWRN1 as well as its predicted targeted mRNAs was dysregulated in different levels of Tfh cell infiltration and might indicate patients’ OS. creator: Yongyong Wang creator: Jianji Guo uri: https://doi.org/10.7717/peerj.9996 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang and Guo title: Cetaceans along the southeastern Brazilian coast: occurrence, distribution and niche inference at local scale link: https://peerj.com/articles/10000 last-modified: 2020-10-05 description: It is deemed important to understand cetacean occurrence and distribution to comprehend their ecological roles. The geographical occurrence of species’ niche can be used to better describe their potential distribution. The niche can be defined using environmental variables. Those variables are considered static and not affected by biological activities. The present study goal was to assess the occurrence and distribution of cetaceans along the southeastern Brazilian coast, as well as to define the fundamental and realized niche of each species and to investigate niche overlap at local scale. The environmental requirements for each species were also investigated throughout statistical tests. Sighting data were obtained through oceanographic surveys conducted between 2012 and 2015. The environmental variables available on MARSPEC and the software NicheA were used for the ecological niche modeling. A total of twelve cetacean species were identified and the potential distribution areas of the six commonest ones were defined. Even though the species presented different environmental preferences, most of them had partial overlap among niches and potential distribution areas. The environmental heterogeneity of the study area might support the co-occurrence of different species with different environmental requirements. creator: Giovanna Corrêa e Figueiredo creator: Karina Bohrer do Amaral creator: Marcos César de Oliveira Santos uri: https://doi.org/10.7717/peerj.10000 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Figueiredo et al. title: Are giant clams (Tridacna maxima) distractible? A multi-modal study link: https://peerj.com/articles/10050 last-modified: 2020-10-05 description: To properly assess risk, an animal must focus its attention on relevant external stimuli; however, attention can be reallocated when distracting stimuli are present. This reallocation of attention may interfere with an individual’s ability to effectively assess risk and may impede its response. Multiple stimuli presented together can have additive effects as distractors, and these include stimuli in different modalities. Although changes in noise and water flow are detectable by some bivalves, this has not been studied in the context of risk assessment or distraction. We experimentally exposed giant clams (Tridacna maxima) to changes in water particle movement through underwater sound (motorboat noise) and increased water flow to determine whether these stimuli, individually or together, modified risk assessment or caused distraction. We found that clams responded to sound, flow, and their combination by increasing frequency of mantle retractions (a potential anti-predator response) when exposed to a stimulus. Sound alone did not change risk assessment in either the latency to close or to reemerge following closure. However, when exposed to both stimuli simultaneously, clams increased their latency to close. We suggest that clams perceive sound and flow in an additive way, and are thus distracted. Interestingly, and uniquely, clams discriminate these multimodal stimuli through a single sensory modality. For sessile clams, anthropogenic noise is detectable, yet unavoidable, suggesting that they be especially vulnerable to marine noise pollution. creator: Ryan Doyle creator: Jonathan Kim creator: Angelika Pe creator: Daniel T. Blumstein uri: https://doi.org/10.7717/peerj.10050 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Doyle et al. title: Predictors of Gull-billed tern (Gelochelidon nilotica) nest survival in artificial coastal saltpans, Bohai Bay, China link: https://peerj.com/articles/10054 last-modified: 2020-10-05 description: BackgroundCoastal saltpans are a common supratidal human-modified wetland habitat found within many coastal landscape mosaics. Commercial salt production and aquaculture practices often result in the creation of exposed coastal substrates that could provide suitable breeding habitat for waterbird populations; however, few studies have quantified waterbird breeding success in these artificial wetlands.MethodsHere we examine the nesting behavior of the Gull-billed tern (Gelochelidon nilotica) breeding in the Nanpu coastal saltpans of Bohai Bay, Yellow Sea, China over three consecutive nesting seasons (2017–2019) by using nest survival model in Program MARK.ResultsThe results revealed that nest survival of Gull-billed terns in coastal saltpans (0.697) was higher than previously published estimates from other regions, with an estimated daily survival rate (DSR) of 0.982 ± 0.001 (±95% CI). High nest survival was mainly attributed to low levels of human disturbances and low predation rates, while exposure to strong winds, flooding and silting were the main factors causing nest failure. Model-averaged estimates revealed that eggs laid in nests located on ‘habitat islands’ with feather or clam shell substrates were most likely to hatch. Initiation date, nest age, clutch size and quadratic effects of nearest-neighbor distance, nearest distance to road and nearest distance to water were all significant predictors of nest success, but the nest survival declined overall from 2017 to 2019 due to the degradation and loss of breeding habitat anthropogenically caused by rising water levels.DiscussionCoastal saltpans represent an alternative breeding habitat for the Gull-billed tern populations in Bohai Bay, but conservation management should prioritize flood prevention to improve the extent and quality of breeding habitat, concurrent with efforts to create further ‘habitat islands’ with suitable nesting substrate. creator: Fuxing Wu creator: Weipan Lei creator: Huw Lloyd creator: Zhengwang Zhang uri: https://doi.org/10.7717/peerj.10054 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wu et al. title: Description of a new member of the family Erysipelotrichaceae: Dakotella fusiforme gen. nov., sp. nov., isolated from healthy human feces link: https://peerj.com/articles/10071 last-modified: 2020-10-05 description: A Gram-positive, non-motile, rod-shaped facultative anaerobic bacterial strain SG502T was isolated from healthy human fecal samples in Brookings, SD, USA. The comparison of the 16S rRNA gene placed the strain within the family Erysipelotrichaceae. Within this family, Clostridium innocuum ATCC 14501T, Longicatena caecimuris strain PG-426-CC-2, Eubacterium dolichum DSM 3991T and E. tortuosum DSM 3987T(=ATCC 25548T) were its closest taxa with 95.28%, 94.17%, 93.25%, and 92.75% 16S rRNA sequence identities respectively. The strain SG502T placed itself close to C. innocuum in the 16S rRNA phylogeny. The members of genus Clostridium within family Erysipelotrichaceae was proposed to be reassigned to genus Erysipelatoclostridium to resolve the misclassification of genus Clostridium. Therefore, C. innocuum was also classified into this genus temporarily with the need to reclassify it in the future because of its difference in genomic properties. Similarly, genome sequencing of the strain and comparison with its 16S phylogenetic members and proposed members of the genus Erysipelatoclostridium, SG502T warranted a separate genus even though its 16S rRNA similarity was >95% when comapred to C. innocuum. The strain was 71.8% similar at ANI, 19.8% [17.4–22.2%] at dDDH and 69.65% similar at AAI to its closest neighbor C. innocuum. The genome size was nearly 2,683,792 bp with 32.88 mol% G+C content, which is about half the size of C. innocuum genome and the G+C content revealed 10 mol% difference. Phenotypically, the optimal growth temperature and pH for the strain SG502T were 37 °C and 7.0 respectively. Acetate was the major short-chain fatty acid product of the strain when grown in BHI-M medium. The major cellular fatty acids produced were C18:1ω9c, C18:0and C16:0. Thus, based on the polyphasic analysis, for the type strain SG502T (=DSM 107282T= CCOS 1889T), the name Dakotella fusiforme gen. nov., sp. nov., is proposed. creator: Sudeep Ghimire creator: Supapit Wongkuna creator: Joy Scaria uri: https://doi.org/10.7717/peerj.10071 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Ghimire et al. title: Analysis of pedestrian activity before and during COVID-19 lockdown, using webcam time-lapse from Cracow and machine learning link: https://peerj.com/articles/10132 last-modified: 2020-10-05 description: At the turn of February and March 2020, COVID-19 pandemic reached Europe. Many countries, including Poland imposed lockdown as a method of securing social distance between potentially infected. Stay-at-home orders and movement control within public space not only affected the touristm industry, but also the everyday life of the inhabitants. The hourly time-lapse from four HD webcams in Cracow (Poland) are used in this study to estimate how pedestrian activity changed during COVID-19 lockdown. The collected data covers the period from 9 June 2016 to 19 April 2020 and comes from various urban zones. One zone is tourist, one is residential and two are mixed. In the first stage of the analysis, a state-of-the-art machine learning algorithm (YOLOv3) is used to detect people. Additionally, a non-standard application of the YOLO method is proposed, oriented to the images from HD webcams. This approach (YOLOtiled) is less prone to pedestrian detection errors with the only drawback being the longer computation time. Splitting the HD image into smaller tiles increases the number of detected pedestrians by over 50%. In the second stage, the analysis of pedestrian activity before and during the COVID-19 lockdown is conducted for hourly, daily and weekly averages. Depending on the type of urban zone, the number of pedestrians decreased from 33% in residential zones to 85% in tourist zones located in the Old Town. The presented method allows for more efficient detection and counting of pedestrians from HD time-lapse webcam images compared to SSD, YOLOv3 and Faster R-CNN. The result of the research is a published database with the detected number of pedestrians from the four-year observation period for four locations in Cracow. creator: Robert Szczepanek uri: https://doi.org/10.7717/peerj.10132 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Szczepanek title: Effects of early life stage exposure of largemouth bass to atrazine or a model estrogen (17α-ethinylestradiol) link: https://peerj.com/articles/9614 last-modified: 2020-10-02 description: Endocrine disrupting contaminants are of continuing concern for potentially contributing to reproductive dysfunction in largemouth and smallmouth bass in the Chesapeake Bay watershed (CBW) and elsewhere. Exposures to atrazine (ATR) have been hypothesized to have estrogenic effects on vertebrate endocrine systems. The incidence of intersex in male smallmouth bass from some regions of CBW has been correlated with ATR concentrations in water. Fish early life stages may be particularly vulnerable to ATR exposure in agricultural areas, as a spring influx of pesticides coincides with spawning and early development. Our objectives were to investigate the effects of early life stage exposure to ATR or the model estrogen 17α-ethinylestradiol (EE2) on sexual differentiation and gene expression in gonad tissue. We exposed newly hatched largemouth bass (LMB, Micropterus salmoides) from 7 to 80 days post-spawn to nominal concentrations of 1, 10, or 100 µg ATR/L or 1 or 10 ng EE2/L and monitored histological development and transcriptomic changes in gonad tissue. We observed a nearly 100% female sex ratio in LMB exposed to EE2 at 10 ng/L, presumably due to sex reversal of males. Many gonad genes were differentially expressed between sexes. Multidimensional scaling revealed clustering by gene expression of the 1 ng EE2/L and 100 µg ATR/L-treated male fish. Some pathways responsive to EE2 exposure were not sex-specific. We observed differential expression in male gonad in LMB exposed to EE2 at 1 ng/L of several genes involved in reproductive development and function, including star, cyp11a2, ddx4 (previously vasa), wnt5b, cyp1a and samhd1. Expression of star, cyp11a2 and cyp1a in males was also responsive to ATR exposure. Overall, our results confirm that early development is a sensitive window for estrogenic endocrine disruption in LMB and are consistent with the hypothesis that ATR exposure induces some estrogenic responses in the developing gonad. However, ATR-specific and EE2-specific responses were also observed. creator: Jessica K. Leet creator: Catherine A. Richter creator: Robert S. Cornman creator: Jason P. Berninger creator: Ramji K. Bhandari creator: Diane K. Nicks creator: James L. Zajicek creator: Vicki S. Blazer creator: Donald E. Tillitt uri: https://doi.org/10.7717/peerj.9614 license: https://creativecommons.org/publicdomain/zero/1.0/ rights: title: Acarbose-metformin is more effective in glycemic variability control than repaglinide-metformin in T2DM patients inadequately controlled with metformin: a retrospective cohort study link: https://peerj.com/articles/9905 last-modified: 2020-10-02 description: BackgroundAcarbose and repaglinide are widely used either by themselves or in combination with other medications. However, their efficacy in diabetes control has not been compared when used in combination with metformin.MethodsThe present study aimed to compare their effects on glycemic variability (GV) control when taken with metformin for type 2 diabetes mellitus (T2DM) inadequately controlled with metformin alone. In this retrospective cohort study, T2DM patients who were treated with either acarbose-metformin or repaglinide-metformin combination were recruited. Either acarbose 100 mg or repaglinide 2 mg triple daily was taken for the subsequent 12 weeks in combination with metformin. Demographic data, biochemical data and 7-point glycemic self-monitoring conducted with capillary blood (SMBG) data were reviewed after one week and 12 weeks. The primary outcome including glucose control and changes in GV as well as other factors affecting GV and the incidence of hypoglycemia were also analyzed.ResultsOf the 305 T2DM patients enrolled, data from 273 subjects, 136 in the acarbose-metformin group (M+A) and 137 in the repaglinide-metformin group (M+R) were analyzed. Both regimens improved glycemic control at 12 weeks post commencement of new medications. GV, expressed as the mean amplitude of plasma glycemic excursions (MAGE, 5.0 ± 2.6 vs. 2.8 ± 1.6 mmol/L, p < 0.001 in M+A; 5.1 ± 2.5 vs. 2.9 ± 1.3 mmol/L, p < 0.001 in M+R), standard deviation of blood glucose (SDBG, 3.6 ± 1.3 vs. 2.0 ± 0.9 mmol/L, p < 0.001 in M+A; 3.7 ± 1.3 vs. 2.4 ± 1.3 p < 0.001 in M+R), coefficient of variation of blood glucose (CVBG, (0.30 ± 0.09 vs. 0.21 ± 0.1, p < 0.001 in M+A; 0.31 ± 0.09 vs. 0.24 ± 0.12, p < 0.001 in M+R), postprandial amplitude of glycemic excursions (PPGE, 5.2 ± 2.6 vs. 2.8 ± 1.6 mmol/L, p < 0.001 in M+A; 5.3 ± 2.5 vs. 2.9 ± 1.3 mmol/L, p < 0.001 in M+R) or largest amplitude of glycemic excursions (LAGE, 9.8 ± 3.6 vs. 5.4 ± 2.4 mmol/L, p < 0.001 in M+A; 10.1 ± 3.4 vs. 6.3 ± 3.2 mmol/L, p < 0.001 in M+R) decreased significantly after the addition of acarbose or repaglinide (p < 0.05 respectively). Compared with repaglinide-metformin, acarbose-metformin was more effective in GV control at 12 weeks post commencement of new medications (p < 0.05). This study indicates that both acarbose-metformin and repaglinide-metformin combinations could effectively reduce GV and the acarbose-metformin combination seems to be more effective than the repaglinide-metformin combination. However, this conclusion should be confirmed by future large-scaled and more comprehensive studies due to the limitations of the present study. creator: Guoli Du creator: Wanrun Xie creator: Yinxia Su creator: Yao Ma creator: Xiaoming Gao creator: Sheng Jiang creator: Huazheng Liang uri: https://doi.org/10.7717/peerj.9905 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Du et al. title: A novel dual MEK/PDK1 inhibitor 9za retards the cell cycle at G0/G1 phase and induces mitochondrial apoptosis in non-small cell lung cancer cells link: https://peerj.com/articles/9981 last-modified: 2020-10-02 description: BackgroundA novel dual MEK/PDK1 inhibitor named 9za has been synthesized by our research team. Preliminary study showed that 9za possessed potent cytotoxicity and proapoptosis in non-small cell lung cancer (NSCLC) cells. Nevertheless, the precise underlying mechanism is vague.MethodsIn this work, we adopted the MTT assay, the Cell Cycle Detection Kit, and the JC-1 staining assay to detect the cell viability, the cell cycle distribution and the mitochondrial membrane potential (MMP), respectively. Cell apoptosis was measured by the morphology observation under a light microscope, Annexin V-FITC/propidium iodide (PI) apoptosis detection and the colorimetric TUNEL assay. Western blot was used to monitor the cell cycle-, apoptosis-related proteins and relevant proteins involved in the signaling pathways.ResultsThe MTT assay demonstrated that 9za sharply decreased the viability of NSCLC cells. Cell cycle analysis revealed that low concentrations of 9za arrested the cell cycle at the G0/G1 phase , which was further confirmed by the decreased levels of Cyclin D1, cyclin-dependent kinase 4 (CDK4) and cyclin-dependent kinase 6 (CDK6). Additionally, morphological observations, Annexin V-FITC/propidium iodide (PI) apoptosis analysis and TUNEL assays indicated that high concentrations of 9za induced cell apoptosis. Furthermore, the JC-1 staining assay revealed that the mitochondrial membrane potential was downregulated following 9za exposure. Western blot also showed that 9za markedly decreased the expression levels of total Bcl-2, Cytochrome C in the mitochondria and BCL2 associated X (BAX) in the cytoplasm. However, the levels of BAX in the mitochondria, Cytochrome C in the cytoplasm, active caspase-9, active caspase-3 and cleaved–PARP showed the opposite changes. Moreover, the dose-dependent decreased phosphorylation levels of PDK1, protein kinase B (Akt), MEK and extracellular signal regulated kinase 1/2 (ERK1/2) after 9za treatment verified that 9za was indeed a dual MEK/PDK1 inhibitor, as we expected. Compared with a single MEK inhibitor PD0325901 or a single PDK1 inhibitor BX517, the dual MEK/PDK1 inhibitor 9za could strengthen the cytotoxic and proapoptotic effect, indicating that the double blocking of the MEK and PDK1 signaling pathways plays stronger cell growth inhibition and apoptosis induction roles than the single blocking of the MEK or PDK1 signaling pathway in NSCLC cells. Our work elucidated the molecular mechanisms for 9za as a novel drug candidate against NSCLC. creator: Rangru Liu creator: Zutao Yu creator: Zhuo Chen creator: Danqi Liu creator: Fengying Huang creator: Qianbin Li creator: Gaoyun Hu creator: Xinan Yi creator: Xi Li creator: Honghao Zhou creator: Zhaoqian Liu uri: https://doi.org/10.7717/peerj.9981 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liu et al. title: The effect of long-term volleyball training on the level of somatic parameters of female volleyball players in various age categories link: https://peerj.com/articles/9992 last-modified: 2020-10-02 description: Volleyball is an exceedingly popular physical activity in the adolescent population, especially with females. The study objective was to assess the effect of volleyball training and natural ontogenetic development on the somatic parameters of adolescent girls. The study was implemented in a group of 130 female volleyball players (aged 12.3 ± 0.5 – 18.1 ± 0.6 years) along with 283 females from the general population (aged 12.3 ± 0.5 – 18.2 ± 0.5 years). The measured parameters included: body height (cm), body mass (kg), body fat (kg, %), visceral fat (cm2), body water (l), fat free mass (kg) and skeletal muscle mass (kg, %). Starting at the age of 13, the volleyball players had significantly lower body fat ratio and visceral fat values than those in the general population (p < 0.001 in body fat % and p < 0.01 in visceral fat). In volleyball players, the mean body fat (%) values were 17.7 ± 6.6 in 12-year-old players, 16.7 ± 4.9 in 13-year-old players, 18.5 ± 3.9 in 16-year-old players, and 19.3 ± 3.1 in 18-year-old players. In the general population, the mean body fat (%) values were 19.6 ± 6.3 in 12-year-old girls, 21.7 ± 6.4 in 13-year-old girls, 23.4 ± 6.1 in 16-year-old girls, and 25.8 ± 7.0 in 18-year-old girls. The visceral fat (cm2) mean values were 36.4 ± 19.3 in 12-year-old players, 39.2 ± 16.3 in 13-year-old players, 45.7 ± 14.7 in 16-year-old players, and 47.2 ± 12.4 in 18-year-old players. In the general population, the mean visceral fat (cm2) values were 41.4 ± 21.1 in 12-year-old girls, 48.4 ± 21.5 in 13-year-old girls, 58.0 ± 24.7 in 16-year-old girls, and 69.1 ± 43.7 in 18-year-old girls. In volleyball players, lower body fat ratio corresponded with a higher skeletal muscle mass ratio. The differences found in skeletal muscle mass ratio were also significant starting at the age of 13 (p < 0.001). The mean skeletal muscle mass (%) values were 44.1 ± 3.4 in 12-year-old volleyball players, 45.4 ± 2.5 in 13-year-old players, 45.0 ± 2.2 in 16-year-old players, and 44.7 ± 1.8 in 18-year-old players. In the general population, the mean skeletal muscle mass (%) values were 42.8 ± 3.2 in 12-year-old girls, 42. ± 4.1 in 13-year-old girls, 41.9 ± 3.3 in 16-year-old girls, and 40.6 ± 3.7 in 18-year-old girls. Differences in body composition between the individual age groups were similar between the volleyball players and girls in the general population. The results indicate that regular volleyball training influences the body composition of young females however the development of body composition parameters is subject to their ontogenetic development. creator: Petr Kutáč creator: David Zahradnik creator: Miroslav Krajcigr creator: Václav Bunc uri: https://doi.org/10.7717/peerj.9992 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kutáč et al. title: Differentially expressed genes related to oxidoreductase activity and glutathione metabolism underlying the adaptation of Phragmites australis from the salt marsh in the Yellow River Delta, China link: https://peerj.com/articles/10024 last-modified: 2020-10-02 description: The common reed (Phragmites australis) is a dominant species in the coastal wetlands of the Chinese Yellow River Delta, where it tolerates a wide range of salinity. Recent environmental changes have led to the increase of soil salinity in this region, which has degraded much of the local vegetation. Clones of common reeds from the tidal marsh may have adapted to local high salinity habitat through selection on genes and metabolic pathways conferring salt tolerance. This study aims to reveal molecular mechanisms underlying salt tolerance in the tidal reed by comparing them to the salt-sensitive freshwater reed under salt stress. We employed comparative transcriptomics to reveal the differentially expressed genes (DEGs) between these two types of common reeds under different salinity conditions. The results showed that only three co-expressed genes were up-regulated and one co-expressed gene was down-regulated between the two reed types. On the other hand, 1,371 DEGs were exclusively up-regulated and 285 DEGs were exclusively down-regulated in the tidal reed compared to the control, while 115 DEGs were exclusively up-regulated and 118 DEGs were exclusively down-regulated in the freshwater reed compared to the control. From the pattern of enrichment of transcripts involved in salinity response, the tidal reed was more active and efficient in scavenging reactive oxygen species (ROS) than the freshwater reed, with the tidal reed showing significantly higher gene expression in oxidoreductase activity. Furthermore, when the reeds were exposed to salt stress, transcripts encoding glutathione metabolism were up-regulated in the tidal reed but not in the freshwater reed. DEGs related to encoding glutathione reductase (GR), glucose-6-phosphate 1-dehydrogenase (G6PDH), 6-phosphogluconate dehydrogenase (6PD), glutathione S-transferase (GST) and L-ascorbate peroxidase (LAP) were revealed as especially highly differentially regulated and therefore represented candidate genes that could be cloned into plants to improve salt tolerance. Overall, more genes were up-regulated in the tidal reed than in the freshwater reed from the Yellow River Delta when under salt stress. The tidal reed efficiently resisted salt stress by up-regulating genes encoding for oxidoreductase activity and glutathione metabolism. We suggest that this type of common reed could be extremely useful in the ecological restoration of degraded, high salinity coastal wetlands in priority. creator: Liwen Zhang creator: Lin Chen creator: Feng Lu creator: Ziting Liu creator: Siqun Lan creator: Guangxuan Han uri: https://doi.org/10.7717/peerj.10024 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: Comparing local ancestry inference models in populations of two- and three-way admixture link: https://peerj.com/articles/10090 last-modified: 2020-10-02 description: Local ancestry estimation infers the regional ancestral origin of chromosomal segments in admixed populations using reference populations and a variety of statistical models. Integrating local ancestry into complex trait genetics has the potential to increase detection of genetic associations and improve genetic prediction models in understudied admixed populations, including African Americans and Hispanics. Five methods for local ancestry estimation that have been used in human complex trait genetics are LAMP-LD (2012), RFMix (2013), ELAI (2014), Loter (2018), and MOSAIC (2019). As users rather than developers, we sought to perform direct comparisons of accuracy, runtime, memory usage, and usability of these software tools to determine which is best for incorporation into association study pipelines. We find that in the majority of cases RFMix has the highest median accuracy with the ranking of the remaining software dependent on the ancestral architecture of the population tested. Additionally, we estimate the O(n) of both memory and runtime for each software and find that for both time and memory most software increase linearly with respect to sample size. The only exception is RFMix, which increases quadratically with respect to runtime and linearly with respect to memory. Effective local ancestry estimation tools are necessary to increase diversity and prevent population disparities in human genetics studies. RFMix performs the best across methods, however, depending on application, other methods perform just as well with the benefit of shorter runtimes. Scripts used to format data, run software, and estimate accuracy can be found at https://github.com/WheelerLab/LAI_benchmarking. creator: Ryan Schubert creator: Angela Andaleon creator: Heather E. Wheeler uri: https://doi.org/10.7717/peerj.10090 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Schubert et al. title: An immunoinformatics approach for the design of a multi-epitope subunit vaccine for urogenital schistosomiasis link: https://peerj.com/articles/8795 last-modified: 2020-10-02 description: Discovery of T and B memory cells capable of eliciting long-term immunity against schistosomiasisis is important for people in endemic areas. Changes in schistosomes environment due to developmental cycle, induces up-regulation of Heat Shock Proteins (HSPs) which assist the parasite in coping with the hostile conditions associated with its life cycle. This study therefore focused on exploring the role of HSPs in urogenital schistosomiasis to develop new multi-epitope subunit vaccine against the disease using immunoinformatic approaches. The designed subunit vaccine was subjected to in silico antigenicity, immunogenicity, allergenicity and physicochemical parameters analysis. A 3D structure of the vaccine construct was predicted, followed by disulphide engineering for stability, codon adaptation and in silico cloning for proper expression and molecular protein–protein docking of vaccine construct in the vector against toll-like receptor 4 receptor, respectively. Consequently, a 493 amino acid multi-epitope vaccine construct of antigenicity probability of 0.91 was designed. This was predicted to be stable, non-allergenic in nature and safe for human use. creator: Olugbenga S. Onile creator: Adeyinka I. Fadahunsi creator: Ameerah A. Adekunle creator: Bolaji F. Oyeyemi creator: Chiaka I. Anumudu uri: https://doi.org/10.7717/peerj.8795 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Onile et al. title: A basilosaurid archaeocete (Cetacea, Pelagiceti) from the Late Eocene of Oregon, USA link: https://peerj.com/articles/9809 last-modified: 2020-10-02 description: BackgroundBasilosaurid archaeocetes are known from the Late Eocene of virtually all coastlines bearing coeval marine rocks except the North Pacific Basin, until now. Here we report on three consecutive posterior thoracic vertebrae of a large, basilosaurid archaeocete from a Late Eocene horizon in the Keasey Formation in Oregon.MethodsThese vertebrae were morphologically and morphometrically compared to other vertebrae of similar age from around the world.ResultsThe specimens were determined to be different from all currently named species of fossil cetacean, but most similar to those found in the Gulf Coast region of North America. These vertebrae represent the first confirmed specimen of a Late Eocene basilosaurid from the North Pacific. These and other basilosaurids known only from vertebrae are reviewed here in the context of Late Eocene paleoceanography and cetacean evolution. creator: Mark D. Uhen creator: David Taylor uri: https://doi.org/10.7717/peerj.9809 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Uhen and Taylor title: Two extremely rare new species of fossorial salamanders of the genus Oedipina (Plethodontidae) from northwestern Ecuador link: https://peerj.com/articles/9934 last-modified: 2020-10-02 description: We describe two new species of salamanders of the genus Oedipina, subgenus Oedopinola, from two localities on the northwestern foothills of Ecuador, at elevations between 921 and 1,067 m. These are the southernmost members of the genus. We examined different museum collections and we found just three specimens of Oedipina from Ecuador, obtained throughout the history of herpetological collections in the country. We identify two of the three specimens as new species, but refrain from assigning a specific identity to the third, pending further study. Oedipina villamizariorum sp. n. is a medium-sized member of the genus, with a narrow, relatively pointed head and blunt snout; dorsolaterally oriented eyes, moderate in size; and digits that are moderately long and having pointed tips. Oedipina ecuatoriana sp. n., somewhat larger, has a narrow head and broadly rounded snout; this new species differs from all known Oedipina by the distinctive presence of paired prefrontal bones and a reduced phalangeal formula: 0-0-1-0; 0-1-2-1-1. We provide detailed descriptions of the osteology of both new species. Finally, we present a phylogenetic hypothesis for the genus, including one of the two new species, based on partial sequences of mitochondrial DNA. creator: Carolina Reyes-Puig creator: David B. Wake creator: Ramachandran Kotharambath creator: Jeffrey W. Streicher creator: Claudia Koch creator: Diego F. Cisneros-Heredia creator: Mario H. Yánez-Muñoz creator: Santiago Ron uri: https://doi.org/10.7717/peerj.9934 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Reyes-Puig et al. title: The relationship between protein domains and homopeptides in the Plasmodium falciparum proteome link: https://peerj.com/articles/9940 last-modified: 2020-10-02 description: The proteome of the malaria parasite Plasmodium falciparum is notable for the pervasive occurrence of homopeptides or low-complexity regions (i.e., regions that are made from a small subset of amino-acid residue types). The most prevalent of these are made from residues encoded by adenine/thymidine (AT)-rich codons, in particular asparagine. We examined homopeptide occurrences within protein domains in P. falciparum. Homopeptide enrichments occur for hydrophobic (e.g., valine), or small residues (alanine or glycine) in short spans (<5 residues), but these enrichments disappear for longer lengths. We observe that short asparagine homopeptides (<10 residues long) have a dramatic relative depletion inside protein domains, indicating some selective constraint to keep them from forming. We surmise that this is possibly linked to co-translational protein folding, although there are specific protein domains that are enriched in longer asparagine homopeptides (≥10 residues) indicating a functional linkage for specific poly-asparagine tracts. Top gene ontology functional category enrichments for homopeptides associated with diverse protein domains include “vesicle-mediated transport”, and “DNA-directed 5′-3′ RNA polymerase activity”, with various categories linked to “binding” evidencing significant homopeptide depletions. Also, in general homopeptides are substantially enriched in the parts of protein domains that are near/in IDRs. The implications of these findings are discussed. creator: Yue Wang creator: Hsin Jou Yang creator: Paul M. Harrison uri: https://doi.org/10.7717/peerj.9940 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: Fish predation hinders the success of coral restoration efforts using fragmented massive corals link: https://peerj.com/articles/9978 last-modified: 2020-10-02 description: As coral reefs continue to decline globally, coral restoration practitioners have explored various approaches to return coral cover and diversity to decimated reefs. While branching coral species have long been the focus of restoration efforts, the recent development of the microfragmentation coral propagation technique has made it possible to incorporate massive coral species into restoration efforts. Microfragmentation (i.e., the process of cutting large donor colonies into small fragments that grow fast) has yielded promising early results. Still, best practices for outplanting fragmented corals of massive morphologies are continuing to be developed and modified to maximize survivorship. Here, we compared outplant success among four species of massive corals (Orbicella faveolata, Montastraea cavernosa, Pseudodiploria clivosa, and P. strigosa) in Southeast Florida, US. Within the first week following coral deployment, predation impacts by fish on the small (<5 cm2) outplanted colonies resulted in both the complete removal of colonies and significant tissue damage, as evidenced by bite marks. In our study, 8–27% of fragments from four species were removed by fish within one week, with removal rates slowing down over time. Of the corals that remained after one week, over 9% showed signs of fish predation. Our findings showed that predation by corallivorous fish taxa like butterflyfishes (Chaetodontidae), parrotfishes (Scaridae), and damselfishes (Pomacentridae) is a major threat to coral outplants, and that susceptibility varied significantly among coral species and outplanting method. Moreover, we identify factors that reduce predation impacts such as: (1) using cement instead of glue to attach corals, (2) elevating fragments off the substrate, and (3) limiting the amount of skeleton exposed at the time of outplanting. These strategies are essential to maximizing the efficiency of outplanting techniques and enhancing the impact of reef restoration. creator: Gammon Koval creator: Nicolas Rivas creator: Martine D’Alessandro creator: Dalton Hesley creator: Rolando Santos creator: Diego Lirman uri: https://doi.org/10.7717/peerj.9978 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Koval et al. title: A new species of alien terrestrial planarian in Spain: Caenoplana decolorata link: https://peerj.com/articles/10013 last-modified: 2020-10-02 description: Terrestrial planarians found in a plant nursery in Spain in 2012 are described as a new species, Caenoplana decolorata. Dorsally they are mahogany brown with a cream median line. Ventrally they are pastel turquoise fading to brown laterally. Molecular data indicate that they are a member of the genus Caenoplana, but that they differ from other Caenoplana species found in Europe. One mature specimen has been partially sectioned, and the musculature and copulatory apparatus is described, confirming the generic placement but distinguishing the species from other members of the genus. It is probable that the species originates from Australia. creator: Eduardo Mateos creator: Hugh D. Jones creator: Marta Riutort creator: Marta Álvarez-Presas uri: https://doi.org/10.7717/peerj.10013 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Mateos et al. title: Early childhood caries and its associations with sugar consumption, overweight and exclusive breastfeeding in low, middle and high-income countries: an ecological study link: https://peerj.com/articles/9413 last-modified: 2020-10-01 description: AimThis ecological study examined the associations between the prevalence of early childhood caries (ECC), overweight, country’s per capita sugar consumption and duration of exclusive breastfeeding.MethodsPer capita consumption of sugar in kilograms, percentage of children exclusively breastfed until 6 months of age, percentage of 0–5-year-old children with overweight status, and percentage of 3–5-year-old children with ECC were compared among low-income countries (LICs), middle-income countries (MICs) and high-income countries (HICs). The association between the prevalence of ECC and the study variables, and the effect modification by income region were assessed using multivariable linear regression models. Regression coefficients, confidence intervals, partial eta squared and P-values for effect modification were calculated.ResultsThe per capita sugar consumption in LICs was significantly lower than in MICs (P = 0.001) and HICs (P < 0.001). The percentage of infants who exclusively breastfed up to 6 months was significantly lower in HICs than in LICs (P < 0.001) and MICs (P = 0.003). The prevalence of overweight was significantly lower in LICs than in MICs (P < 0.001) and HICs (P = 0.021). The prevalence of ECC was significantly lower in HICs than in MICs (P < 0.001). Income was a significant modifier of the associations between the prevalence of ECC, per capita sugar consumption (P = 0.005), and exclusive breastfeeding up to 6 months (P = 0.03). The associations between the prevalence of ECC and per capita sugar consumption at the global level and for MICs were stronger (partial eta squared = 0.05 and 0.13 respectively) than for LICs and HICs (partial eta squared <0.0001 and 0.003 respectively). Only in MICs was there a significant association between the prevalence of ECC and per capita sugar consumption (P = 0.002), and between the prevalence of ECC and the percentage of children exclusively breastfed up to 6 months (P = 0.02).ConclusionThough the quantity of sugar consumption and exclusive breastfeeding may be a significant risk indicator for ECC in MICs, sugar consumption may be more of a risk indicator for ECC in HICs than in LICs, and vice versa for exclusive breastfeeding. Although ECC and overweight are both sugar-related diseases, we found no significant relationship between them. creator: Morenike O. Folayan creator: Maha El Tantawi creator: Francisco Ramos-Gomez creator: Wael Sabbah uri: https://doi.org/10.7717/peerj.9413 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Folayan et al. title: Dragonflies and damselflies (Odonata) from Puerto Rico: a checklist with notes on distribution and habitat link: https://peerj.com/articles/9711 last-modified: 2020-10-01 description: BackgroundConservation of tropical freshwater fauna requires a solid understanding of species biodiversity patterns. We provide an up to date annotated list of Odonata of Puerto Rico, which is based on current reports. The list is complemented with notes on the geographic and altitudinal distribution of this order on the island. We also compare current composition relative to early reports conducted when Puerto Rico was mostly an agricultural region.MethodsWe surveyed adult Odonata all over Puerto Rico with the aid of undergraduate students. Students were trained on capturing, preserving, and data basing specimens. All material was centralized, identified by the lead author, and deposited in the Zoology Museum at the University of Puerto Rico (MZUPR), Río Piedras campus. Surveys were complemented with focal collections by the authors and a literature review of published records for Puerto Rico and the Caribbean. We requested records from specialists to obtain the most complete list of species for the island.ResultsAn annotated list of Odonata from Puerto Rico is presented, reporting 49 species distributed in two suborders and four families. We provide information on species distribution among municipalities and elevations around Puerto Rico. A historic list of species was developed for the 1930s-1940s, when agriculture covered most of Puerto Rico, based on literature and museum specimens. Both current and historic records are similar and suggest that the Odonata fauna is dominated by generalist species and has changed little since the agricultural period. Our list provides a point of reference to understand biodiversity patterns in Puerto Rico and the Caribbean and for assessing the effects of land use change on aquatic insect diversity. creator: Alonso Ramírez creator: Norman Maldonado-Benítez creator: Ashley Mariani-Ríos creator: Javier Figueroa-Santiago uri: https://doi.org/10.7717/peerj.9711 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Ramírez et al. title: Differential effects of cisplatin on cybrid cells with varying mitochondrial DNA haplogroups link: https://peerj.com/articles/9908 last-modified: 2020-10-01 description: BackgroundDrug therapy yields different results depending on its recipient population. Cisplatin, a commonly used chemotherapeutic agent, causes different levels of resistance and side effects for different patients, but the mechanism(s) are presently unknown. It has been assumed that this variation is a consequence of differences in nuclear (n) DNA, epigenetics, or some external factor(s). There is accumulating evidence that an individual’s mitochondrial (mt) DNA may play a role in their response to medications. Variations within mtDNA can be observed, and an individual’s mtDNA can be categorized into haplogroups that are defined by accumulations of single nucleotide polymorphisms (SNPs) representing different ethnic populations.MethodsThe present study was conducted on transmitochondrial cytoplasmic hybrids (cybrids) that possess different maternal-origin haplogroup mtDNA from African (L), Hispanic [A+B], or Asian (D) backgrounds. Cybrids were created by fusing Rho0 ARPE-19 cells (lacking mtDNA) with platelets, which contain numerous mitochondria but no nuclei. These cybrid cells were cultured to passage five, treated with cisplatin, incubated for 48 h, then analyzed for cell metabolic activity (tetrazolium dye (MTT) assay), mitochondrial membrane potential (JC-1 assay), cytotoxicity (lactate dehydrogenase (LDH) assay), and gene expression levels for ALK, BRCA1, EGFR, and ERBB2/HER2.ResultsResults indicated that untreated cybrids with varying mtDNA haplogroups had similar relative metabolic activity before cisplatin treatment. When treated with cisplatin, (1) the decline in metabolic activity was greatest in L (27.4%, p < 0.012) < D (24.86%, p = 0.0001) and [A+B] cybrids (24.67%, p = 0.0285) compared to untreated cybrids; (2) mitochondrial membrane potential remained unchanged in all cybrids (3) LDH production varied between cybrids (L >[A+B], p = 0.0270). (4) The expression levels decreased for ALK in L (p < 0.0001) and [A+B] (p = 0.0001) cybrids but not in D cybrids (p = 0.285); and decreased for EGFR in [A+B] cybrids (p = 0.0246) compared to untreated cybrids.ConclusionOur findings suggest that an individual’s mtDNA background may be associated with variations in their response to cisplatin treatment, thereby affecting the efficiency and the severity of side effects from the treatment. creator: Sina Abedi creator: Gregory Yung creator: Shari R. Atilano creator: Kunal Thaker creator: Steven Chang creator: Marilyn Chwa creator: Kevin Schneider creator: Nitin Udar creator: Daniela Bota creator: M. Cristina Kenney uri: https://doi.org/10.7717/peerj.9908 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Abedi et al. title: Connectivity of bacterial assemblages along the Loa River in the Atacama Desert, Chile link: https://peerj.com/articles/9927 last-modified: 2020-10-01 description: The Loa River is the only perennial artery that crosses the Atacama Desert in northern Chile. It plays an important role in the ecological and economic development of the most water-stressed region, revealing the impact of the mining industry, which exacerbate regional water shortages for many organisms and ecological processes. Despite this, the river system has remained understudied. To our knowledge, this study provides the first effort to attempt to compare the microbial communities at spatial scale along the Loa River, as well as investigate the physicochemical factors that could modulate this important biological component that still remains largely unexplored. The analysis of the spatial bacterial distribution and their interconnections in the water column and sediment samples from eight sites located in three sections along the river catchment (upper, middle and lower) was conducted using 16S rRNA gene-based Illumina MiSeq sequencing. Among a total of 543 ASVs identified at the family level, over 40.5% were cosmopolitan in the river and distributed within a preference pattern by the sediment substrate with 162 unique ASVs, while only 87 were specific to the column water. Bacterial diversity gradually decreased from the headwaters, where the upper section had the largest number of unique families. Distinct groupings of bacterial communities often associated with anthropogenic disturbance, including Burkholderiaceae and Flavobacteriaceae families were predominant in the less-impacted upstream section. Members of the Arcobacteraceae and Marinomonadaceae were prominent in the agriculturally and mining-impacted middle sector while Rhodobacteraceae and Coxiellaceae were most abundant families in downstream sites. Such shifts in the community structure were also related to the influence of salinity, chlorophyll, dissolved oxygen and redox potential. Network analyses corroborated the strong connectivity and modular structure of bacterial communities across this desert river, shedding light on taxonomic relatedness of co-occurring species and highlighting the need for planning the integral conservation of this basin. creator: Ana Zárate creator: Cristina Dorador creator: Ruben Araya creator: Mariela Guajardo creator: July Z. Florez creator: Gonzalo Icaza creator: Diego Cornejo creator: Jorge Valdés uri: https://doi.org/10.7717/peerj.9927 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zárate et al. title: Persistent mosquito fogging can be detrimental to non-target invertebrates in an urban tropical forest link: https://peerj.com/articles/10033 last-modified: 2020-10-01 description: BackgroundHuman population growth has led to biodiversity declines in tropical cities. While habitat loss and fragmentation have been the main drivers of urban biodiversity loss, man-made interventions to reduce health risks have also emerged as an unintentional threat. For instance, insecticide fogging to control mosquito populations has become the most common method of preventing the expansion of mosquito-borne diseases such as Dengue. However, the effectiveness of fogging in killing mosquitoes has been called into question. One concern is the unintended effect of insecticide fogging on non-target invertebrates that are crucial for the maintenance of urban ecosystems. Here, we investigate the impacts of fogging on: (1) target invertebrate taxon (Diptera, including mosquitoes); (2) non-target invertebrate taxa; and (3) the foraging behavior of an invertebrate pollinator taxon (Lepidoptera) within an urban tropical forest.MethodsWe carried out fogging with Pyrethroid insecticide (Detral 2.5 EC) at 10 different sites in a forest situated in the state of Selangor, Peninsular Malaysia. Across the sites, we counted the numbers of knocked-down invertebrates and identified them based on morphology to different taxa. We constructed Bayesian hierarchical Poisson regression models to investigate the effects of fogging on: (1) a target invertebrate taxon (Diptera) 3-h post-fogging; (2) selected non-target invertebrate taxa 3-h post-fogging; and (3) an invertebrate pollinator taxon (Lepidoptera) 24-h post-fogging.ResultsA total of 1,874 invertebrates from 19 invertebrate orders were knocked down by the fogging treatment across the 10 sites. Furthermore, 72.7% of the invertebrates counted 3-h post-fogging was considered dead. Our regression models showed that given the data and prior information, the probability that fogging had a negative effect on invertebrate taxa 3-h post-fogging was 100%, with reductions to 11% of the pre-fogging count of live individuals for the target invertebrate taxon (Diptera), and between 5% and 58% of the pre-fogging count of live individuals for non-target invertebrate taxa. For the invertebrate pollinator, the probability that fogging had a negative effect 24-h post-fogging was also 100%, with reductions to 53% of the pre-fogging count of live individuals.DiscussionOur Bayesian models unequivocally demonstrate that fogging has detrimental effects on one pollinator order and non-target invertebrate orders, especially taxa that have comparatively lower levels of chitinisation. While fogging is effective in killing the target order (Diptera), no mosquitos were found dead in our experiment. In order to maintain urban biodiversity, we recommend that health authorities and the private sector move away from persistent insecticide fogging and to explore alternative measures to control adult mosquito populations. creator: Nicole S.M. Lee creator: Gopalasamy R. Clements creator: Adeline S.Y. Ting creator: Zhi H. Wong creator: Sze H. Yek uri: https://doi.org/10.7717/peerj.10033 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Lee et al. title: Low-cost air pollution monitoring system—an opportunity for reducing the health risk associated with physical activity in polluted air link: https://peerj.com/articles/10041 last-modified: 2020-10-01 description: The issue of air pollution by particulate matter (PM) concerns many places in the world. At the same time, many residents undertake physical activity (recreation, rehabilitation, sport) in the open air. Generally, the amount of dust concentration depends on both the place (center or periphery of the city) and the time of day. In the present study we describe the outcome of monitoring of the state of air pollution by particle matter (PM10) in the Kraków agglomeration area in order to show that it can provide information concerning air quality in the area where people practice varied kinds of sports in the open air. The measurements of PM10 have been made by a few stations with identical construction working as one network. The details of the air pollution monitoring system and its data quality verification have been described. The network stations made multipoint observations across the Kraków Metropolitan Area during the year 2017 in eight locations. The locations selected represent a diverse spectrum of terrain conditions in which the Kraków agglomeration community undertakes physical activity. For most months of 2017, the minimum monthly average 4-hour PM10 concentrations were recorded between 10–14 h, regardless of location, whereas the maximum was between 18–22. We also noticed a huge differences in the average monthly value of PM10 in some locations within the Kraków agglomeration—ranging between 4.9–339.0 µg m−3. This indicates that some regions of the city are more suitable for performance of physical activity in the open air than others. In conclusion, we postulate that a low-cost air pollution monitoring system is capable of providing valuable information concerning air quality in a given region, which seems to be of importance also to people who practice varied sports activities in the open air. creator: Zenon Nieckarz creator: Jerzy A. Zoladz uri: https://doi.org/10.7717/peerj.10041 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Nieckarz and Zoladz title: Brain hothubs and dark functional networks: correlation analysis between amplitude and connectivity for Broca’s aphasia link: https://peerj.com/articles/10057 last-modified: 2020-10-01 description: Source localization and functional brain network modeling are methods of identifying critical regions during cognitive tasks. The first activity estimates the relative differences of the signal amplitudes in regions of interest (ROI) and the second activity measures the statistical dependence among signal fluctuations. We hypothesized that the source amplitude–functional connectivity relationship decouples or reverses in persons having brain impairments. Five Broca’s aphasics with five matched cognitively healthy controls underwent overt picture-naming magnetoencephalography scans. The gamma-band (30–45 Hz) phase-locking values were calculated as connections among the ROIs. We calculated the partial correlation coefficients between the amplitudes and network measures and detected four node types, including hothubs with high amplitude and high connectivity, coldhubs with high connectivity but lower amplitude, non-hub hotspots, and non-hub coldspots. The results indicate that the high-amplitude regions are not necessarily highly connected hubs. Furthermore, the Broca aphasics utilized different hothub sets for the naming task. Both groups had dark functional networks composed of coldhubs. Thus, source amplitude–functional connectivity relationships could help reveal functional reorganizations in patients. The amplitude–connectivity combination provides a new perspective for pathological studies of the brain’s dark functional networks. creator: Feng Lin creator: Shao-Qiang Cheng creator: Dong-Qing Qi creator: Yu-Er Jiang creator: Qian-Qian Lyu creator: Li-Juan Zhong creator: Zhong-Li Jiang uri: https://doi.org/10.7717/peerj.10057 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Lin et al. title: Iris sanguinea is conspecific with I. sibirica (Iridaceae) according to morphology and plastid DNA sequence data link: https://peerj.com/articles/10088 last-modified: 2020-10-01 description: A taxonomic revision of Iris subser. Sibiricae is provided based on morphological and molecular analyses and the study of protologues and original material. Two to three species have been recognized in this subseries by botanists. To address the question of species delimitations and relationships within this group, we analyzed four non-coding regions of plastid DNA (trnS–trnG, trnL–trnF, rps4–trnSGGA, and psbA–trnH) for samples from 26 localities across the distribution ranges of two currently recognized species, I. sanguinea and I. sibirica. Variance analysis, based on nine characters, revealed no separation between taxa. Moreover, no morphological character could be used to define clear boundaries between taxa. Our results strongly support that I. subser. Sibiricae is monotypic and comprises only I. sibirica, instead of two or three species. Iris sibirica is morphologically variable and one of the most widespread Eurasian species of Iridaceae. Previously accepted taxa, I. sanguinea and I. typhifolia, are synonymised with I. sibirica and also two names, I. orientalis and I. sibirica var. haematophylla, which are typified here, are placed in the synonymy of I. sibirica. Information on the distribution of I. sibirica and the main features used to distinguish between I. sibirica and I. subser. Chrysographes species are provided. creator: Eugeny Boltenkov creator: Elena Artyukova creator: Marina Kozyrenko creator: Andrey Erst creator: Anna Trias-Blasi uri: https://doi.org/10.7717/peerj.10088 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Boltenkov et al. title: Positive association between COVID-19 deaths and influenza vaccination rates in elderly people worldwide link: https://peerj.com/articles/10112 last-modified: 2020-10-01 description: BackgroundThe coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is an ongoing global health crisis, directly and indirectly impacting all spheres of human life. Some pharmacological measures have been proposed to prevent COVID-19 or reduce its severity, such as vaccinations. Previous reports indicate that influenza vaccination appears to be negatively correlated with COVID-19-associated mortality, perhaps as a result of heterologous immunity or changes in innate immunity. The understanding of such trends in correlations could prevent deaths from COVID-19 in the future. The aim of this study was therefore to analyze the association between COVID-19 related deaths and influenza vaccination rate (IVR) in elderly people worldwide.MethodsTo determine the association between COVID-19 deaths and influenza vaccination, available data sets from countries with more than 0.5 million inhabitants were analyzed (in total 39 countries). To accurately estimate the influence of IVR on COVID-19 deaths and mitigate effects of confounding variables, a sophisticated ranking of the importance of different variables was performed, including as predictor variables IVR and some potentially important geographical and socioeconomic variables as well as variables related to non-pharmaceutical intervention. The associations were measured by non-parametric Spearman rank correlation coefficients and random forest functions.ResultsThe results showed a positive association between COVID-19 deaths and IVR of people ≥65 years-old. There is a significant increase in COVID-19 deaths from eastern to western regions in the world. Further exploration is needed to explain these findings, and additional work on this line of research may lead to prevention of deaths associated with COVID-19. creator: Christian Wehenkel uri: https://doi.org/10.7717/peerj.10112 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wehenkel