title: PeerJ description: Articles published in PeerJ link: https://peerj.com/articles/index.rss3?journal=peerj&month=2021-02 creator: info@peerj.com PeerJ errorsTo: info@peerj.com PeerJ language: en title: DisCoVering potential candidates of RNAi-based therapy for COVID-19 using computational methods link: https://peerj.com/articles/10505 last-modified: 2021-02-26 description: The ongoing pandemic of a novel coronavirus (SARS-CoV-2) leads to international concern; thus, emergency interventions need to be taken. Due to the time-consuming experimental methods for proposing useful treatments, computational approaches facilitate investigating thousands of alternatives simultaneously and narrow down the cases for experimental validation. Herein, we conducted four independent analyses for RNA interference (RNAi)-based therapy with computational and bioinformatic methods. The aim is to target the evolutionarily conserved regions in the SARS-CoV-2 genome in order to down-regulate or silence its RNA. miRNAs are denoted to play an important role in the resistance of some species to viral infections. A comprehensive analysis of the miRNAs available in the body of humans, as well as the miRNAs in bats and many other species, were done to find efficient candidates with low side effects in the human body. Moreover, the evolutionarily conserved regions in the SARS-CoV-2 genome were considered for designing novel significant siRNA that are target-specific. A small set of miRNAs and five siRNAs were suggested as the possible efficient candidates with a high affinity to the SARS-CoV-2 genome and low side effects. The suggested candidates are promising therapeutics for the experimental evaluations and may speed up the procedure of treatment design. Materials and implementations are available at: https://github.com/nrohani/SARS-CoV-2. creator: Narjes Rohani creator: Fatemeh Ahmadi Moughari creator: Changiz Eslahchi uri: https://doi.org/10.7717/peerj.10505 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Rohani et al. title: Composition and distribution of fish environmental DNA in an Adirondack watershed link: https://peerj.com/articles/10539 last-modified: 2021-02-26 description: BackgroundEnvironmental DNA (eDNA) surveys are appealing options for monitoring aquatic biodiversity. While factors affecting eDNA persistence, capture and amplification have been heavily studied, watershed-scale surveys of fish communities and our confidence in such need further exploration.MethodsWe characterized fish eDNA compositions using rapid, low-volume filtering with replicate and control samples scaled for a single Illumina MiSeq flow cell, using the mitochondrial 12S ribosomal RNA locus for taxonomic profiling. Our goals were to determine: (1) spatiotemporal variation in eDNA abundance, (2) the filtrate needed to achieve strong sequencing libraries, (3) the taxonomic resolution of 12S ribosomal sequences in the study environment, (4) the portion of the expected fish community detectable by 12S sequencing, (5) biases in species recovery, (6) correlations between eDNA compositions and catch per unit effort (CPUE) and (7) the extent that eDNA profiles reflect major watershed features. Our bioinformatic approach included (1) estimation of sequencing error from unambiguous mappings and simulation of taxonomic assignment error under various mapping criteria; (2) binning of species based on inferred assignment error rather than by taxonomic rank; and (3) visualization of mismatch distributions to facilitate discovery of distinct haplotypes attributed to the same reference. Our approach was implemented within the St. Regis River, NY, USA, which supports tribal and recreational fisheries and has been a target of restoration activities. We used a large record of St. Regis-specific observations to validate our assignments.ResultsWe found that 300 mL drawn through 25-mm cellulose nitrate filters yielded greater than 5 ng/µL DNA at most sites in summer, which was an approximate threshold for generating strong sequencing libraries in our hands. Using inferred sequence error rates, we binned 12S references for 110 species on a state checklist into 85 single-species bins and seven multispecies bins. Of 48 bins observed by capture survey in the St. Regis, we detected eDNA consistent with 40, with an additional four detections flagged as potential contaminants. Sixteen unobserved species detected by eDNA ranged from plausible to implausible based on distributional data, whereas six observed species had no 12S reference sequence. Summed log-ratio compositions of eDNA-detected taxa correlated with log(CPUE) (Pearson’s R = 0.655, P < 0.001). Shifts in eDNA composition of several taxa and a genotypic shift in channel catfish (Ictalurus punctatus) coincided with the Hogansburg Dam, NY, USA. In summary, a simple filtering apparatus operated by field crews without prior expertise gave useful summaries of eDNA composition with minimal evidence of field contamination. 12S sequencing achieved useful taxonomic resolution despite the short marker length, and data exploration with standard bioinformatic tools clarified taxonomic uncertainty and sources of error. creator: Robert S. Cornman creator: James E. McKenna, Jr. creator: Jennifer A. Fike uri: https://doi.org/10.7717/peerj.10539 license: https://creativecommons.org/publicdomain/zero/1.0/ rights: title: Soil properties and root traits jointly shape fine-scale spatial patterns of bacterial community and metabolic functions within a Korean pine forest link: https://peerj.com/articles/10902 last-modified: 2021-02-26 description: Spatial heterogeneity of soil bacterial community depends on scales. The fine-scale spatial heterogeneity of bacterial community composition and functions remains unknown. We analyzed the main driving factors of fine-scale spatial patterns of soil bacterial community composition and carbon metabolic functions across a 30 m × 40 m plot within a Korean pine forest by combining Illumina 16S rRNA sequencing with Biolog Ecoplates based on 53 soil samples. Clear spatial patterns in bacterial community composition and metabolic functions were observed in the forest soil. The bacterial community composition and metabolic functions both showed distance-decay of similarity within a distance of meters. Structural equation model analysis revealed that environmental variables and geographic distance together explained 37.9% and 63.1% of community and metabolic functions, respectively. Among all environmental factors, soil organic carbon (SOC) and root biomass emerged as the most important drivers of the bacterial community structure. In contrast, soil pH explained the largest variance in metabolic functions. Root biomass explained the second-largest variance in soil bacterial community composition, but root traits made no difference in metabolic functions variance. These results allow us to better understand the mechanisms controlling belowground diversity and plant-microbe interactions in forest ecosystems. creator: Jialing Teng creator: Jing Tian creator: Guirui Yu creator: Yakov Kuzyakov uri: https://doi.org/10.7717/peerj.10902 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Teng et al. title: High prevalence of circulating DS-1-like human rotavirus A and genotype diversity in children with acute gastroenteritis in Thailand from 2016 to 2019 link: https://peerj.com/articles/10954 last-modified: 2021-02-26 description: BackgroundHuman rotavirus A (RVA) infection is the primary cause of acute gastroenteritis (AGE) in infants and young children worldwide, especially in children under 5 years of age and is a major public health problem causing severe diarrhea in children in Thailand. This study aimed to investigate the prevalence, genotype diversity, and molecular characterization of rotavirus infection circulating in children under 15 years of age diagnosed with AGE in Thailand from January 2016 to December 2019.MethodsA total of 2,001 stool samples were collected from children with gastroenteritis (neonates to children <15 years of age) and tested for RVA by real-time polymerase chain reaction (RT-PCR). Amplified products were sequenced and submitted to an online genotyping tool for analysis.ResultsOverall, 301 (15.0%) stool samples were positive for RVA. RVA occurred most frequently among children aged 0-24 months. The seasonal incidence of rotavirus infection occurred typically in Thailand during the winter months (December-March). The G3P[8] genotype was identified as the most prevalent genotype (33.2%, 100/301), followed by G8P[8] (10.6%, 32/301), G9P[8] (6.3%, 19/301), G2P[4] (6.0%, 18/301), and G1P[6] (5.3%, 16/301). Uncommon G and P combinations such as G9P[4], G2P[8], G3P[4] and G3P[9] were also detected at low frequencies. In terms of genetic backbone, the unusual DS-1-like G3P[8] was the most frequently detected (28.2%, 85/301), and the phylogenetic analysis demonstrated high nucleotide identity with unusual DS-1-like G3P[8] detected in Thailand and several countries.ConclusionsA genetic association between RVA isolates from Thailand and other countries ought to be investigated given the local and global dissemination of rotavirus as it is crucial for controlling viral gastroenteritis, and implications for the national vaccination programs. creator: Siripat Pasittungkul creator: Fajar Budi Lestari creator: Jiratchaya Puenpa creator: Watchaporn Chuchaona creator: Nawarat Posuwan creator: Jira Chansaenroj creator: John Mauleekoonphairoj creator: Natthinee Sudhinaraset creator: Nasamon Wanlapakorn creator: Yong Poovorawan uri: https://doi.org/10.7717/peerj.10954 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Pasittungkul et al. title: Comparison of machine learning approaches for enhancing Alzheimer’s disease classification link: https://peerj.com/articles/10549 last-modified: 2021-02-25 description: Alzheimer’s disease (AD) is a progressive neurodegenerative disorder, accounting for nearly 60% of all dementia cases. The occurrence of the disease has been increasing rapidly in recent years. Presently about 46.8 million individuals suffer from AD worldwide. The current absence of effective treatment to reverse or stop AD progression highlights the importance of disease prevention and early diagnosis. Brain structural Magnetic Resonance Imaging (MRI) has been widely used for AD detection as it can display morphometric differences and cerebral structural changes. In this study, we built three machine learning-based MRI data classifiers to predict AD and infer the brain regions that contribute to disease development and progression. We then systematically compared the three distinct classifiers, which were constructed based on Support Vector Machine (SVM), 3D Very Deep Convolutional Network (VGGNet) and 3D Deep Residual Network (ResNet), respectively. To improve the performance of the deep learning classifiers, we applied a transfer learning strategy. The weights of a pre-trained model were transferred and adopted as the initial weights of our models. Transferring the learned features significantly reduced training time and increased network efficiency. The classification accuracy for AD subjects from elderly control subjects was 90%, 95%, and 95% for the SVM, VGGNet and ResNet classifiers, respectively. Gradient-weighted Class Activation Mapping (Grad-CAM) was employed to show discriminative regions that contributed most to the AD classification by utilizing the learned spatial information of the 3D-VGGNet and 3D-ResNet models. The resulted maps consistently highlighted several disease-associated brain regions, particularly the cerebellum which is a relatively neglected brain region in the present AD study. Overall, our comparisons suggested that the ResNet model provided the best classification performance as well as more accurate localization of disease-associated regions in the brain compared to the other two approaches. creator: Qi Li creator: Mary Qu Yang uri: https://doi.org/10.7717/peerj.10549 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Li and Yang title: Characterization and functional analysis of the Hydroxycinnamoyl-CoA: shikimate hydroxycinnamoyl transferase (HCT) gene family in poplar link: https://peerj.com/articles/10741 last-modified: 2021-02-25 description: Hydroxycinnamoyl-CoA: shikimate hydroxycinnamoyl transferase (HCT) divides the mass flux to H, G and S units in monolignol biosynthesis and affects lignin content. Ten HCT homologs were identified in the Populus trichocarpa (Torr. & Gray) genome. Both genome duplication and tandem duplication resulted in the expansion of HCT orthologs in Populus. Comprehensive analysis including motif analysis, phylogenetic analysis, expression profiles and co-expression analysis revealed the divergence and putative function of these candidate PoptrHCTs. PoptrHCT1 and 2 were identified as likely involved in lignin biosynthesis. PoptrHCT9 and 10- are likely to be involved in plant development and the response to cold stress. Similar functional divergence was also identified in Populus tomentosa Carr. Enzymatic assay of PtoHCT1 showed that PtoHCT1 was able to synthesize caffeoyl shikimate using caffeoyl-CoA and shikimic acid as substrates. creator: Nan Chao creator: Qi Qi creator: Shuang Li creator: Brent Ruan creator: Xiangning Jiang creator: Ying Gai uri: https://doi.org/10.7717/peerj.10741 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Chao et al. title: Genetic structure and historical and contemporary gene flow of Astyanaxmexicanus in the Gulf of Mexico slope: a microsatellite-based analysis link: https://peerj.com/articles/10784 last-modified: 2021-02-25 description: Background Astyanax mexicanus from the river basins of the Gulf of Mexico slope are small freshwater fish that usually live in large groups in different freshwater environments. The group is considered successful due to its high capacity for dispersal and adaptation to different habitats, and the species present high morphological variability throughout their distribution in Mexico. This has produced the most extreme morphotype of the group; the hypogeous or troglobite, which has no eyes or coloration, and is probably the cause of taxonomic uncertainty in the recognition of species across the entire range. Most studies of A. mexicanus have mainly focused on cave individuals, as well as their adjacent surface locations, providing an incomplete evolutionary history, particularly in terms of factors related to dispersal and the potential corridors used, barriers to gene flow, and distribution of genetic variability. The aim of the present study is to determine the population structure and the degree and direction of genetic flow in this complex taxonomic group, incorporating geographic locations not previously included in analyses using microsatellite loci. Our aim is to contribute to the knowledge of the intricate evolutionary history of A. mexicanus throughout most of its range.MethodsThe present study included a set of several cave and surface locations of A. mexicanus, which have been widely sampled along the Gulf of Mexico slope, in a genetic population analysis using 10 microsatellite loci.ResultsTen genetic populations or lineages were identified. In these populations, gene flow was recorded at two time periods. Historical gene flow, both inter and intra-basin, was observed among surface populations, from surface to cave populations, and among cave populations, whereas recording of contemporary gene flow was limited to intra-basin exchanges and observed among surface populations, surface to cave populations, and cave populations. creator: Rodolfo Pérez-Rodríguez creator: Sarai Esquivel-Bobadilla creator: Adonaji Madeleine Orozco-Ruíz creator: José Luis Olivas-Hernández creator: Francisco Javier García-De León uri: https://doi.org/10.7717/peerj.10784 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Pérez-Rodríguez et al. title: Phenotype characteristics of gastric epithelial mucus in patients with different gastric diseases: from superficial gastritis to gastric cancer link: https://peerj.com/articles/10822 last-modified: 2021-02-25 description: BackgroundGastric gland mucin is important for maintaining the basic function of the gastric mucosa, protecting it from foreign substances and reducing the occurrence of gastric diseases. Exploring the phenotype of gastric gland mucus changes during the progression of gastric disease is of great clinical significance.MethodsA total of 483 patients with different gastric diseases were collected in this study, including 82 superficial gastritis (SG), 81 atrophic gastritis (AG), 168 dysplasia (GD), and 152 gastric cancer (GC). Mucin staining was performed using HID-ABpH2.5-PAS method and was further grouped according to the mucin coloration.ResultsThe phenotypic characteristics of mucin during disease progression were divided into neutral, acidic, and mucus-free types. Furthermore, acidic mucus can be divided into type I, type II, and type III. The SG group was dominated by neutral mucus (100%), and the AG was dominated by acid mucus (81.48%), which gradually increased with the severity of atrophy (P < 0.05). The GD and GC groups were dominated by mucus-free (43.45%, 78.29%), and as the degree of GD worsened, neutral and acidic mucus gradually decreased and mucus-free increased (P < 0.001). From the SG, AG, GD, and GC progression, neutral and acidic mucus gradually decreased, and mucus- free gradually increased. Acidic mucin revealed that type III (red-brown black) mucin was predominant in AG, GD, and GC, and increased with the degree of AG, GD, as well as the biological behavior of GC. In the lesion adjacent to high-grade GD or GC, type III acid mucin is predominant.ConclusionThere were three mucin phenotypes in the process of gastric diseases. With the disease progression, the trend of phenotypic change was that neutral and acidic mucus gradually decreased and mucus-free increased. The appearance of type III mucin suggested a relatively serious phase of gastric diseases and may be a more suitable candidate for follow-up monitoring of patients with GC risk. creator: Nannan Dong creator: Rui Guo creator: Yuehua Gong creator: Yuan Yuan uri: https://doi.org/10.7717/peerj.10822 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Dong et al. title: Comparison of microleakage between different restorative materials to restore marginal gap at crown margin link: https://peerj.com/articles/10823 last-modified: 2021-02-25 description: BackgroundAn occurrence of secondary caries around the indirect restoration margin is reported to remain a leading cause of failures.ObjectiveThis study aimed to test the interfacial microleakage of conventional glass-ionomer (CGI), resin-modified glass ionomer (RMGI) and Nano-hybrid composite (CR) restorations at a full veneer margin crown.MethodsNinety human extracted molar teeth were divided into three groups (n = 30). Each group was subdivided into three subgroups (n = 10) according to the extent of the structural defects; The structural defect in G1 had a depth of 1.5 mm, width and length at 2 mm and 1 mm intrusion within the crown cervical margin. The corresponding structural defect dimension values for G2 were 2, 5, 4 and 2 mm with defects extending onto the root structure. Meanwhile, G3: structural deficiency of 2 mm depth, 3 mm width and 3 mm length and with 1.5 mm extension into the prepared teeth. These structural defects in each subgroup were restored with CGI, RMGI and CR. Artificial carious lesion formation was induced at the cervical finish line with a demineralizing solution. The artificial carious lesions were restored as per the group distribution. Subsequently, teeth samples were prepared and cemented with Nickel-chromium full coverage restorations utilizing glass-ionomer luting cement. Teeth samples were thermocycled, isolated with nail varnish, and immersed in 0.1% methylene blue for 24 h. The teeth samples were sectioned longitudinally, dye penetration was evaluated with a stereomicroscope. The data were analyzed with Kruskal–Wallis and Mann–Whitney U tests.ResultsCGI-G1 recorded the highest micro-leakage score at 1.450; while CR-G3 recorded the least score (0.350). At a cementum-restoration interface, CR-G1 (0.850) documented the lowest micro-leakage; RMGI-G3 had a greater value at 1.700.ConclusionsThe hybrid CR could be effectively used to restore the restoration of a marginal gap around crown margins. creator: Satheesh B. Haralur creator: Ghaseb Ahmed AL Ghaseb creator: Norah Ali Alqahtani creator: Bader Alqahtani uri: https://doi.org/10.7717/peerj.10823 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Haralur et al. title: Sheath-tailed bats (Chiroptera: Emballonuridae) from the early Pleistocene Rackham’s Roost Site, Riversleigh World Heritage Area, and the distribution of northern Australian emballonurid species link: https://peerj.com/articles/10857 last-modified: 2021-02-25 description: Sheath-tailed bats (Family Emballonuridae) from the early Pleistocene Rackham’s Roost Site cave deposit in the Riversleigh World Heritage Area, north-western Queensland are the oldest recorded occurrence for the family in Australia. The fossil remains consist of maxillary and dentary fragments, as well as isolated teeth, but until now their precise identity has not been assessed. Our study indicates that at least three taxa are represented, and these are distinguished from other Australian emballonurids based on morphometric analysis of craniodental features. Most of the Rackham’s Roost Site emballonurid remains are referrable to the modern species Taphozous georgianus Thomas, 1915, but the extant species T. troughtoni Tate, 1952 also appears to be present, as well as a very large, as-yet undetermined species of Saccolaimus Temminck, 1838. We identify craniodental features that clearly distinguish T. georgianus from the externally very similar T. troughtoni. Results suggest that the distributions of T. georgianus and T. troughtoni may have overlapped in north-western Queensland since at least the early Pleistocene. creator: Tyler R. King creator: Troy J. Myers creator: Kyle N. Armstrong creator: Michael Archer creator: Suzanne J. Hand uri: https://doi.org/10.7717/peerj.10857 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 King et al. title: The Doublesex/Mab-3 domain transcription factor DMD-10 regulates ASH-dependent behavioral responses link: https://peerj.com/articles/10892 last-modified: 2021-02-25 description: The Doublesex/Mab-3 Domain transcription factor DMD-10 is expressed in several cell types in C. elegans, including in the nervous system. We sought to investigate whether DMD-10 is required for normal neuronal function using behavioral assays. We found that mutation of dmd-10 did not broadly affect behavior. dmd-10 mutants were normal in several behavioral assays including a body bends assay for locomotion, egg laying, chemotaxis and response to gentle touch to the body. dmd-10 mutants did have defects in nose-touch responsiveness, which requires the glutamate receptor GLR-1. However, using quantitative fluorescence microscopy to measure levels of a GLR-1::GFP fusion protein in the ventral nerve cord, we found no evidence supporting a difference in the number of GLR-1 synapses or in the amount of GLR-1 present in dmd-10 mutants. dmd-10 mutants did have decreased responsiveness to high osmolarity, which, along with nose-touch, is sensed by the polymodal sensory neuron ASH. Furthermore, mutation of dmd-10 impaired behavioral response to optogenetic activation of ASH, suggesting that dmd-10 promotes neuronal signaling in ASH downstream of sensory receptor activation. Together our results suggest that DMD-10 is important in regulating the frequency of multiple ASH-dependent behavioral responses. creator: Julia Durbeck creator: Celine Breton creator: Michael Suter creator: Eric S. Luth creator: Annette M. McGehee uri: https://doi.org/10.7717/peerj.10892 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Durbeck et al. title: Resistance training with different repetition duration to failure: effect on hypertrophy, strength and muscle activation link: https://peerj.com/articles/10909 last-modified: 2021-02-25 description: BackgroundThis study investigated the effects of two 14-week resistance training protocols with different repetition duration (RD) performed to muscle failure (MF) on gains in strength and muscle hypertrophy as well as on normalized electromyographic (EMG) amplitude and force-angle relationships.MethodsThe left and right legs of ten untrained males were assigned to either one of the two protocols (2-s or 6-s RD) incorporating unilateral knee extension exercise. Both protocols were performed with 3–4 sets, 50–60% of the one-repetition maximum (1RM), and 3 min rest. Rectus femoris and vastus lateralis cross-sectional areas (CSA), maximal voluntary isometric contraction (MVIC) at 30o and 90o of knee flexion and 1RM performance were assessed before and after the training period. In addition, normalized EMG amplitude-angle and force-angle relationships were assessed in the 6th and 39th experimental sessions.ResultsThe 6-s RD protocol induced larger gains in MVIC at 30o of knee angle measurement than the 2-s RD protocol. Increases in MVIC at 90o of knee angle, 1RM, rectus femoris and vastus lateralis CSA were not significant between the 2-s and 6-s RD protocols. Moreover, different normalized EMG amplitude-angle and force-angle values were detected between protocols over most of the angles analyzed.ConclusionPerforming longer RD could be a more appropriate strategy to provide greater gains in isometric maximal muscle strength at shortened knee positions. However, similar maximum dynamic strength and muscle hypertrophy gains would be provided by protocols with different RD. creator: Lucas Túlio Lacerda creator: Rodrigo Otávio Marra-Lopes creator: Marcel Bahia Lanza creator: Rodrigo César Ribeiro Diniz creator: Fernando Vitor Lima creator: Hugo Cesar Martins-Costa creator: Gustavo Ferreira Pedrosa creator: André Gustavo Pereira Andrade creator: Armin Kibele creator: Mauro Heleno Chagas uri: https://doi.org/10.7717/peerj.10909 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Lacerda et al. title: A community perspective on the concept of marine holobionts: current status, challenges, and future directions link: https://peerj.com/articles/10911 last-modified: 2021-02-25 description: Host-microbe interactions play crucial roles in marine ecosystems. However, we still have very little understanding of the mechanisms that govern these relationships, the evolutionary processes that shape them, and their ecological consequences. The holobiont concept is a renewed paradigm in biology that can help to describe and understand these complex systems. It posits that a host and its associated microbiota with which it interacts, form a holobiont, and have to be studied together as a coherent biological and functional unit to understand its biology, ecology, and evolution. Here we discuss critical concepts and opportunities in marine holobiont research and identify key challenges in the field. We highlight the potential economic, sociological, and environmental impacts of the holobiont concept in marine biological, evolutionary, and environmental sciences. Given the connectivity and the unexplored biodiversity specific to marine ecosystems, a deeper understanding of such complex systems requires further technological and conceptual advances, e.g., the development of controlled experimental model systems for holobionts from all major lineages and the modeling of (info)chemical-mediated interactions between organisms. Here we propose that one significant challenge is to bridge cross-disciplinary research on tractable model systems in order to address key ecological and evolutionary questions. This first step is crucial to decipher the main drivers of the dynamics and evolution of holobionts and to account for the holobiont concept in applied areas, such as the conservation, management, and exploitation of marine ecosystems and resources, where practical solutions to predict and mitigate the impact of human activities are more important than ever. creator: Simon M. Dittami creator: Enrique Arboleda creator: Jean-Christophe Auguet creator: Arite Bigalke creator: Enora Briand creator: Paco Cárdenas creator: Ulisse Cardini creator: Johan Decelle creator: Aschwin H. Engelen creator: Damien Eveillard creator: Claire M.M. Gachon creator: Sarah M. Griffiths creator: Tilmann Harder creator: Ehsan Kayal creator: Elena Kazamia creator: François H. Lallier creator: Mónica Medina creator: Ezequiel M. Marzinelli creator: Teresa Maria Morganti creator: Laura Núñez Pons creator: Soizic Prado creator: José Pintado creator: Mahasweta Saha creator: Marc-André Selosse creator: Derek Skillings creator: Willem Stock creator: Shinichi Sunagawa creator: Eve Toulza creator: Alexey Vorobev creator: Catherine Leblanc creator: Fabrice Not uri: https://doi.org/10.7717/peerj.10911 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Dittami et al. title: Towards a multi-basin SWAT model for the migration of nutrients and pesticides to Puck Bay (Southern Baltic Sea) link: https://peerj.com/articles/10938 last-modified: 2021-02-25 description: BackgroundThis paper analyzes the impact of changes in fertilization on crop yields and the runoff of nutrients from a small agricultural catchment (176 km2) to a shallow bay, using the SWAT model. Puck Bay is part of the Gulf of Gdansk and belongs to the Baltic Sea. The whole area of Puck Bay (364 km2) is protected (Natura 2000) yet despite this it suffers from eutrophication problems due to the relatively minimal depth and difficult water exchange.MethodsThe paper presents a comparison of the calculated yields and the runoff of nutrients and pesticides in the SWAT model, for a small agricultural coastal catchment. Calculations were made for 13 crop scenarios with weather data from 2011 to 2019. For each crop, an agriculture calendar was made. Two variants of fertilization were considered (autofertilization mode and according to the calendar). The nutrient runoff was calculated depending on the adopted scenario. In addition, the fate of selected pesticides was simulated.ResultsDepending on the crop, the annual load of NO3into the stream ranged from 0.74 to 3.65 kg ha−1. The annual load of organic phosphorous into the stream was between 0.686 and 3.64 kg ha−1. This is lower than in the majority of EU or Baltic countries. The surface runoff of dissolved Glyphosate was equal to 286 mg ha−1. The annual loads of nutrients from the catchment area are equivalent in both fertilization modes. Regardless of the selected fertilization mode, in addition to the dosage, the form of nutrients is important for the model. creator: Paweł Wielgat creator: Dominika Kalinowska creator: Adam Szymkiewicz creator: Piotr Zima creator: Beata Jaworska-Szulc creator: Ewa Wojciechowska creator: Nicole Nawrot creator: Karolina Matej-Lukowicz creator: Lidia Anita Dzierzbicka-Glowacka uri: https://doi.org/10.7717/peerj.10938 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wielgat et al. title: Comparative anatomy and salt management of Sonneratia caseolaris (L.) Engl. (Lythraceae) grown in saltwater and freshwater link: https://peerj.com/articles/10962 last-modified: 2021-02-25 description: Sonneratia caseolaris is a pioneer species in mangrove. It can naturally grow in both saltwater and freshwater. The study was aimed at investigating and comparing the anatomical character of the S. caseolaris plants growing in different conditions and how they coped with salinity. The anatomical characteristics of roots, stems, petioles and leaf blade were investigated. The plant samples were prepared into permanent slides using a paraffin method, while the wood samples were made into permanent slides using a sliding microtome technique. Tissue clearing of leaf blade and scanning electron microscopic analysis of wood were performed. In addition, sodium chloride content in various organs and tissues was examined. It was found that cable root, stem and leaf blade showed some different anatomical characteristics between the two conditions. Periderm is a prominent tissue in saltwater roots. Tanniferous cells were observed in pneumatophores, petioles, stems and leaf blades of saltwater plants, but not found in pneumatophores and lamina of freshwater plants. Mesophyll thickness was lower in the saltwater condition. The vessel density was significantly higher in the saltwater condition than in the freshwater condition, whereas the vessel diameters in the freshwater condition were significantly higher than those in the saltwater condition. From the results, it can be concluded that root periderm plays an important role in salt exclusion, and the occurrence of tanniferous cells is associated with salt elimination. creator: Sukrit Tatongjai creator: Ekaphan Kraichak creator: Prasart Kermanee uri: https://doi.org/10.7717/peerj.10962 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Tatongjai et al. title: Sensitivity analysis of the infection transmissibility in the UK during the COVID-19 pandemic link: https://peerj.com/articles/10992 last-modified: 2021-02-25 description: The coronavirus (COVID-19) outbreak started in December 2019 and rapidly spread around the world affecting millions of people. With the growth of infection rate, many countries adopted different policies to control the spread of the disease. The UK implemented strict rules instructing individuals to stay at home except in some special circumstances starting from 23 March 2020. Accordingly, this study focuses on sensitivity analysis of transmissibility of the infection as the effects of removing restrictions, for example by returning different occupational groups to their normal working environment and its effect on the reproduction number in the UK. For this reason, available social contact matrices are adopted for the population of UK to account for the average number of contacts. Different scenarios are then considered to analyse the variability of total contacts on the reproduction number in the UK as a whole and each of its four nations. Our data-driven retrospective analysis shows that if more than 38.5% of UK working-age population return to their normal working environment, the reproduction number in the UK is expected to be higher than 1. However, analysis of each nation, separately, shows that local reproduction number in each nation may be different and requires more adequate analysis. Accordingly, we believe that using statistical methods and historical data can provide good estimation of local transmissibility and reproduction number in any region. As a consequence of this analysis, efforts to reduce the restrictions should be implemented locally via different control policies. It is important that these policies consider the social contacts, population density, and the occupational groups that are specific to each region. creator: Pardis Biglarbeigi creator: Kok Yew Ng creator: Dewar Finlay creator: Raymond Bond creator: Min Jing creator: James McLaughlin uri: https://doi.org/10.7717/peerj.10992 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Biglarbeigi et al. title: A new kentriodontid (Cetacea: Odontoceti) from the early to middle Miocene of the western North Pacific and a revision of kentriodontid phylogeny link: https://peerj.com/articles/10945 last-modified: 2021-02-24 description: A new species of an extinct dolphin belonging to the kentriodontids, i.e., Kentriodon sugawarai sp. nov., is described from the upper lower to lowest middle Miocene Kadonosawa Formation in Ninohe City, Iwate Prefecture, northern Japan. The holotype of Kentriodon sugawarai sp. nov., consists of a partial skull with ear bones, mandibular fragments, and some postcranial bones. This new species shares five unique characters with other species of Kentriodon. In addition, the new species differs from other species of the genus in displaying a narrow width of the squamosal lateral to the exoccipital in posterior view, the dorsolateral edge of the opening of the ventral infraorbital foramen that is formed by the maxilla and the lacrimal or the jugal, and at least three anterior dorsal infraorbital foramina. Our phylogenetic analysis based on 393 characters for 103 Odontoceti taxa yielded a consensus tree showing all previously identified kentriodontids as a monophyletic group that comprises the sister group of the crown Dephinoidea, which in turn include Delphinidae, Phocoenidae and Monodontidae. Our analysis also indicates that the distinct innovation of the acoustic apparatus (i.e., 13 out of 29 derived characters are from tympanoperiotic) would have occurred in the ancestral lineage of the Delphinoidea (sensu lato) including the monophyletic Kentriodontidae during their initial evolution and diversification. creator: Zixuan Guo creator: Naoki Kohno uri: https://doi.org/10.7717/peerj.10945 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Guo and Kohno title: Addiction peer recovery coach training pilot: assessment of confidence levels link: https://peerj.com/articles/10783 last-modified: 2021-02-24 description: BackgroundPeer recovery coaches (PRCs) have become a critical tool in addiction treatment in many areas of the world. Despite this fact, no identified research has examined the process or impact of PRC training. Furthermore, no scales were identified to measure trainee confidence in various PRC techniques. The goal of this article is to analyze the process and immediate impact of PRC training of twelve American Indians (AIs) in a culturally-specific program. We focus most specifically on trainee confidence levels.MethodsNo written consent was obtained and completion of the assessment was considered consent. Trainees completed self-assessments before and after the training. The self-assessment examined nine areas ranging from understanding the role of PRCs to knowledge of effective PRC techniques. Paired t-tests were used to assess for changes in individual trainee responses between the pre- and post-assessments.ResultsPre-training responses ranged from moderate to high. Questions with the lowest average confidence levels address PRC activities or specific techniques to facilitate recovery. All nine questions showed statistically significant mean improvements in the post-training self-assessments. Questions regarding specific PRC activities and techniques showed the greatest improvement. Questions relating to helping people more generally showed the smallest improvement. Average post-training responses fell within a very narrow range indicating relatively consistent confidence levels across skills. Analysis indicates participants were possibly over-confident in certain areas (i.e., maintaining boundaries). This small pilot represents an initial attempt to measure confidence levels of PRC trainees. The findings may inform future training by identifying certain areas where emphasis might be most helpful for trainees. In addition, it is hoped that this work will encourage more systematic analysis of the impact of PRC training on individuals. creator: Nicholas Guenzel creator: Hongying Dai uri: https://doi.org/10.7717/peerj.10783 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Guenzel and Dai title: Evidence of phenotypic plasticity along an altitudinal gradient in the dung beetle Onthophagus proteus link: https://peerj.com/articles/10798 last-modified: 2021-02-24 description: BackgroundHigh altitude insects are an ecologically specialized group and possess a suite of adaptions which allow persistence in the inhospitable conditions often associated with mountain tops. Changes in body coloration and reductions or increases in body size are thought to be examples of such adaptions. Melanic individuals, or individuals containing high levels of eumelanin, possess several traits which increase resistance to ultraviolet radiation and desiccation, while aiding thermoregulation. Trait variation is often observed in dung beetles and is associated with dimorphism and sexual selection. In this study, we identified trait changes which occur across an altitudinal gradient by measuring morphological color and body size traits in a montane insect.MethodsUsing standard digital photography and Image J, we examined individuals of Afromontane dung beetle Onthophagus proteus. Individuals were classified according to sex and color morph to identify intrasexual variance. Nine morphometric traits were measured per beetle to identify patterns of morphology across discrete 500 m altitude segments.ResultsThe results of this study provide one of the first descriptions of trait changes associated with elevation in an African dung beetle. We suggest that color polymorphism in Onthophagus proteus might be at least partly driven by environmental factors as there is significantly increased melanism with increasing elevation and significant differences in color hues between altitude bands. We also suggest changes in horn length are density dependent, as we observed an increase in cephalic horn length at high elevations where O. proteus is the most abundant species. creator: Roisin A. Stanbrook creator: W. Edwin Harris creator: Charles P. Wheater creator: Martin Jones uri: https://doi.org/10.7717/peerj.10798 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Stanbrook et al. title: Distribution, fractionation and sources of rare earth elements in suspended particulate matter in a tropical agricultural catchment, northeast Thailand link: https://peerj.com/articles/10853 last-modified: 2021-02-24 description: Forty-eight suspended particulate matter (SPM) samples were collected from the Mun River, northeast Thailand and its junction with the Mekong River, to investigate the relationship between the distribution of rare earth elements (REE) in SPM and the soils in the watershed. The total REE contents (∑REE) in SPM in the Mun River ranged from 78.5 to 377.8 mg/kg with the average of 189.3 mg/kg, which was lower than ∑REE of 222.3 mg/kg at the Mekong River (one sample at junction). The Post Archean Australia Shale (PAAS)-normalized ratios of light REE (LREE), middle REE (MREE) and heavy REE (HREE) were averaged to 1.0, 1.3 and 1.0, which showed a clear enrichment in MREE. In short, along the Mun River, the REE contents in SPM were decreasing, and the PAAS-normalized patterns of REE showed gradually flat. The REE content in SPM and soils are highest in the upper catchment, indicating that soil/bedrock is the most important source of REE in SPM. Additionally, the positive Eu anomaly was enhanced by the higher Ca content in SPM (R = 0.45), which may be caused by more feldspars or carbonates with Ca and Eu substituting Ca. The results present the REE behaviors of SPM in the Mun River and relationship between REE in SPM and soil/bedrock, the findings may support the other studies in catchment weathering. creator: Kunhua Yang creator: Guilin Han creator: Jie Zeng creator: Wenxiang Zhou uri: https://doi.org/10.7717/peerj.10853 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Yang et al. title: Characteristics of planktonic and sediment bacterial communities in a heavily polluted urban river link: https://peerj.com/articles/10866 last-modified: 2021-02-24 description: Urban rivers represent a unique ecosystem in which pollution occurs regularly, altering the biogeochemical characteristics of waterbodies and sediments. However, little is presently known about the spatiotemporal patterns of planktonic and sediment bacterial community diversities and compositions in urban rivers. Herein, Illumina MiSeq high-throughput sequencing was performed to reveal the spatiotemporal dynamics of bacterial populations in Liangtan River, a heavily polluted urban river in Chongqing City (China). The results showed the richness and diversity of sediment bacteria were significantly higher than those of planktonic bacteria, whereas a strong overlap (46.7%) in OTUs was identified between water and sediment samples. Bacterial community composition remarkably differed in waters and sediments. Planktonic bacterial communities were dominated by Proteobacteria, Bacteroidetes, Cyanobacteria and Actinobacteria, while sediment bacterial communities mainly included Proteobacteria, Actinobacteria, Chloroflexi and Bacteroidetes. Additionally, several taxonomic groups of potential bacterial pathogens showed an increasing trend in water and sediment samples from residential and industrial areas (RI). Variation partition analysis (VPA) indicated that temperature and nutrient were identified as the main drivers determining the planktonic and sediment bacterial assemblages. These results highlight that bacterial communities in the polluted urban river exhibit spatiotemporal variation due to the combined influence of environmental factors associated with sewage discharge and hydropower dams. creator: Heqing Huang creator: Jianhui Liu creator: Fanghui Zhang creator: Kangwen Zhu creator: Chunhua Yang creator: Qiujie Xiang creator: Bo Lei uri: https://doi.org/10.7717/peerj.10866 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Huang et al. title: Extremely low nucleotide diversity among thirty-six new chloroplast genome sequences from Aldama (Heliantheae, Asteraceae) and comparative chloroplast genomics analyses with closely related genera link: https://peerj.com/articles/10886 last-modified: 2021-02-24 description: Aldama (Heliantheae, Asteraceae) is a diverse genus in the sunflower family. To date, nearly 200 Asteraceae chloroplast genomes have been sequenced, but the plastomes of Aldama remain undescribed. Plastomes in Asteraceae usually show little sequence divergence, consequently, our hypothesis is that species of Aldama will be overall conserved. In this study, we newly sequenced 36 plastomes of Aldama and of five species belonging to other Heliantheae genera selected as outgroups (i.e., Dimerostemma asperatum, Helianthus tuberosus, Iostephane heterophylla, Pappobolus lanatus var. lanatus, and Tithonia diversifolia). We analyzed the structure and gene content of the assembled plastomes and performed comparative analyses within Aldama and with other closely related genera. As expected, Aldama plastomes are very conserved, with the overall gene content and orientation being similar in all studied species. The length of the plastome is also consistent and the junction between regions usually contain the same genes and have similar lengths. A large ∼20 kb and a small ∼3 kb inversion were detected in the Large Single Copy (LSC) regions of all assembled plastomes, similarly to other Asteraceae species. The nucleotide diversity is very low, with only 1,509 variable sites in 127,466 bp (i.e., 1.18% of the sites in the alignment of 36 Aldama plastomes, with one of the IRs removed, is variable). Only one gene, rbcL, shows signatures of positive selection. The plastomes of the selected outgroups feature a similar gene content and structure compared to Aldama and also present the two inversions in the LSC region. Deletions of different lengths were observed in the gene ycf2. Multiple SSRs were identified for the sequenced Aldama and outgroups. The phylogenetic analysis shows that Aldama is not monophyletic due to the position of the Mexican species A. dentata. All Brazilian species form a strongly supported clade. Our results bring new understandings into the evolution and diversity of plastomes at the species level. creator: Benoit Loeuille creator: Verônica Thode creator: Carolina Siniscalchi creator: Sonia Andrade creator: Magdalena Rossi creator: José Rubens Pirani uri: https://doi.org/10.7717/peerj.10886 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Loeuille et al. title: Pushing up or pushing out—an initial investigation into horizontal- versus vertical-force training on swimming start performance: a pilot study link: https://peerj.com/articles/10937 last-modified: 2021-02-24 description: BackgroundThe block phase in the swimming start requires a quick reaction to the starting signal and a large take-off velocity that is primarily horizontal in direction. Due to the principle of specificity of training, there is a potential benefit of performing a greater proportion of horizontal force production exercises in a swimmers’ dry-land resistance training sessions. Therefore, the purpose of this pilot study was to provide an insight into the effects of a horizontal- (HF) vs vertical-force (VF) training intervention on swim start performance.MethodsEleven competitive swimmers (six males (age 20.9 ± 1.8 years, body mass 77.3 ± 9.7 kg, height 1.78 ± 0.05 m) and five females (age 21.4 ± 2.0 years, body mass 67.5 ± 7.4 kg, height 1.69 ± 0.05 m)) completed 2 weekly sessions of either a horizontal- or vertical-force focused resistance training programme for 8 weeks. Squat jump force-time characteristics and swim start kinetic and kinematic parameters were collected pre- and post-intervention.ResultsAcross the study duration, the swimmers completed an average of nine swimming sessions per week with an average weekly swim volume of 45.5 ± 17.7 km (HF group) and 53 ± 20.0 km (VF group), but little practice of the swim start per week (n = 9). Within-group analyses indicated a significant increase in predicted one repetition maximum (1RM) hip thrust strength in the HF group, as well as significant increases in grab resultant peak force but reductions in resultant peak force of the block phase for the VF group. No significant between-group differences in predicted 1RM hip thrust and back squat strength, squat jump force-time and swim start performance measures were observed after 8 weeks of training. Significant correlations in the change scores of five block kinetic variables to time to 5 m were observed, whereby increased block kinetic outputs were associated with a reduced time to 5 m. This may be indicative of individual responses to the different training programmes.DiscussionThe results of this current study have been unable to determine whether a horizontal- or vertical-force training programme enhances swim start performance after an 8-week training intervention. Some reasons for the lack of within and between group effects may reflect the large volume of concurrent training and the relative lack of any deliberate practice of the swim start. Larger samples and longer training duration may be required to determine whether significant differences occur between these training approaches. Such research should also look to investigate how a reduction in the concurrent training loads and/or an increase in the deliberate practice of the swim start may influence the potential changes in swim start performance. creator: Shiqi Thng creator: Simon Pearson creator: Justin W.L. Keogh uri: https://doi.org/10.7717/peerj.10937 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Thng et al. title: The effect of an integrated reading and anxiety intervention for poor readers with anxiety link: https://peerj.com/articles/10987 last-modified: 2021-02-24 description: A recent systematic review has reported that poor reading is reliably associated with anxiety. However, we currently lack evidence-based intervention for children who have both poor reading and anxiety (PRAX). In this study, we tested a new PRAX intervention in 8- to 12-year-old children using a double-baseline intervention case series design. Analyses of both group and individual data revealed that 12 weeks of PRAX intervention significantly improved children’s reading and spelling accuracy, and significantly reduced both anxiety disorders and symptoms. These results support PRAX intervention as a treatment for comorbid reading and anxiety problems in children and pave the way to a randomised controlled trial. creator: Deanna Francis creator: Jennifer L. Hudson creator: Saskia Kohnen creator: Lynn Mobach creator: Genevieve M. McArthur uri: https://doi.org/10.7717/peerj.10987 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Francis et al. title: Spatial temporal distribution of COVID-19 risk during the early phase of the pandemic in Malawi link: https://peerj.com/articles/11003 last-modified: 2021-02-24 description: BackgroundCOVID-19 has been one of the greatest challenges the world has faced since the second world war. This study aimed at investigating the distribution of COVID-19 in both space and time in Malawi.MethodsThe study used publicly available data of COVID-19 cases for the period from 2 April 2020 to 28 October 2020. Semiparametric spatial temporal models were fitted to the number of monthly confirmed cases as an outcome data, with time and district as independent variables, where district was the spatial unit, while accounting for sociodemographic factors.ResultsThe study found significant effects of location and time, with the two interacting. The spatial distribution of COVID-19 risk showed major cities being at greater risk than rural areas. Over time, the COVID-19 risk was increasing then decreasing in most districts with the rural districts being consistently at lower risk. High proportion of elderly people was positively associated with COVID-19 risk (β = 1.272, 95% CI [0.171, 2.370]) than low proportion of elderly people. There was negative association between poverty incidence and COVID-19 risk (β = −0.100, 95% CI [−0.136, −0.065]).ConclusionFuture or present strategies to limit the spread of COVID-19 should target major cities and the focus should be on time periods that had shown high risk. Furthermore, the focus should be on elderly and rich people. creator: Alfred Ngwira creator: Felix Kumwenda creator: Eddons C.S. Munthali creator: Duncan Nkolokosa uri: https://doi.org/10.7717/peerj.11003 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Ngwira et al. title: Low compositions of human toll-like receptor 7/8-stimulating RNA motifs in the MERS-CoV, SARS-CoV and SARS-CoV-2 genomes imply a substantial ability to evade human innate immunity link: https://peerj.com/articles/11008 last-modified: 2021-02-24 description: BackgroundThe innate immune system especially Toll-like receptor (TLR) 7/8 and the interferon pathway, constitutes an important first line of defense against single-stranded RNA viruses. However, large-scale, systematic comparisons of the TLR 7/8-stimulating potential of genomic RNAs of single-stranded RNA viruses are rare. In this study, a computational method to evaluate the human TLR 7/8-stimulating ability of single-stranded RNA virus genomes based on their human TLR 7/8-stimulating trimer compositions was used to analyze 1,002 human coronavirus genomes.ResultsThe human TLR 7/8-stimulating potential of coronavirus genomic (positive strand) RNAs followed the order of NL63-CoV > HKU1-CoV >229E-CoV ≅ OC63-CoV > SARS-CoV-2 > MERS-CoV > SARS-CoV. These results suggest that among these coronaviruses, MERS-CoV, SARS-CoV and SARS-CoV-2 may have a higher ability to evade the human TLR 7/8-mediated innate immune response. Analysis with a logistic regression equation derived from human coronavirus data revealed that most of the 1,762 coronavirus genomic (positive strand) RNAs isolated from bats, camels, cats, civets, dogs and birds exhibited weak human TLR 7/8-stimulating potential equivalent to that of the MERS-CoV, SARS-CoV and SARS-CoV-2 genomic RNAs.ConclusionsPrediction of the human TLR 7/8-stimulating potential of viral genomic RNAs may be useful for surveillance of emerging coronaviruses from nonhuman mammalian hosts. creator: Chu-Wen Yang creator: Mei-Fang Chen uri: https://doi.org/10.7717/peerj.11008 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Yang and Chen title: Comparison between cultured and wild Pacific white shrimp (Penaeus vannamei) vitellogenesis: next-generation sequencing and relative expression of genes directly and indirectly related to reproduction link: https://peerj.com/articles/10694 last-modified: 2021-02-23 description: Shrimp fisheries are among the most important fisheries worldwide, and shrimp culture has increased considerably in recent years. Most current studies on reproduction-related genes have been conducted on cultured shrimp. However, gene expression is intimately linked to physiological and environmental conditions, and therefore an organism’s growth environment has a great influence on reproduction. Thus, gene expression profiling, should be applied in fisheries studies. Here, we identified the expression patterns of 76 reproduction-related genes in P. vannamei via the analysis of pooled transcriptomes from a time-series experiment encompassing a full circadian cycle. The expression patterns of genes associated both directly (Vtg, ODP, and ProR) and indirectly (FAMet, CruA1, and CruC1) with reproduction were evaluated, as these genes could be used as molecular markers of previtellogenic and vitellogenic maturation stages. The evaluated genes were prominently upregulated during vitellogenic stages, with specific expression patterns depending on the organism’s environment, diet, and season. Vtg, ProR, ODP, and FaMet could serve as molecular markers for both wild and cultured organisms. creator: Araceli Lorena Montes-Dominguez creator: Jesus Arian Avena-Soto creator: Jorge Luis Lizarraga-Rodriguez creator: Rodrigo de Jesus Perez-Gala creator: Stephanie Jimenez-Gutierrez creator: Jesus Alberto Sotelo-Falomir creator: Fernando Marino Pinzon-Miranda creator: Francisco Martinez-Perez creator: Horacio Alberto Muñoz-Rubi creator: Dario Chavez-Herrera creator: Laura Rebeca Jimenez-Gutierrez uri: https://doi.org/10.7717/peerj.10694 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Montes-Dominguez et al. title: Fat mass prediction equations and reference ranges for Saudi Arabian Children aged 8–12 years using machine technique method link: https://peerj.com/articles/10734 last-modified: 2021-02-23 description: BackgroundThe number of children with obesity has increased in Saudi Arabia, which is a significant public health concern. Early diagnosis of childhood obesity and screening of the prevalence is needed using a simple in situ method. This study aims to generate statistical equations to predict body fat percentage (BF%) for Saudi children by employing machine learning technology and to establish gender and age-specific body fat reference range.MethodsData was combined from two cross-sectional studies conducted in Saudi Arabia for 1,292 boys and girls aged 8–12 years. Body fat was measured in both studies using bio-electrical impedance analysis devices. Height and weight were measured and body mass index was calculated and classified according to CDC 2,000 charts. A total of 603 girls and 374 boys were randomly selected for the learning phase, and 153 girls and 93 boys were employed in the validation set. Analyses of different machine learning methods showed that an accurate, sensitive model could be created. Two regression models were trained and fitted with the construction samples and validated. Gradient boosting algorithm was employed to achieve a better estimation and produce the equations, then the root means squared error (RMSE) equation was performed to decrease the error. Body fat reference ranges were derived for children aged 8–12 years.ResultsFor the gradient boosting models, the predicted fat percentage values were more aligned with the true value than those in regression models. Gradient boosting achieved better performance than the regression equation as it combined multiple simple models into a single composite model to take advantage of that weak classifier. The developed predictive model archived RMSE of 3.12 for girls and 2.48 boys. BF% and Fat mass index charts were presented in which cut-offs for 5th, 75th and 95th centiles are used to define ‘under-fat’, ‘normal’, ‘overfat’ and ‘subject with obesity’.ConclusionMachine learning models could represent a significant advancement for investigators studying adiposity-related issues in children. These models and newly developed centile charts could be useful tools for the estimation and classification of BF%. creator: Rabab B. Alkutbe creator: Abdulrahman Alruban creator: Hmidan Alturki creator: Anas Sattar creator: Hazzaa Al-Hazzaa creator: Gail Rees uri: https://doi.org/10.7717/peerj.10734 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Alkutbe et al. title: Contact toxicity of three insecticides for use in tier I pesticide risk assessments with Megachile rotundata (Hymenoptera: Megachilidae) link: https://peerj.com/articles/10744 last-modified: 2021-02-23 description: The current pesticide risk assessment paradigm may not adequately protect solitary bees as it focuses primarily on the honey bee (Apis mellifera). The alfalfa leafcutting bee (Megachile rotundata) is a potential surrogate species for use in pesticide risk assessment for solitary bees in North America. However, the toxicity of potential toxic reference standards to M. rotundata will need to be determined before pesticide risk assessment tests (tier I trials) can be implemented. Therefore, we assessed the acute topical toxicity and generated LD50 values for three insecticides: dimethoate (62.08 ng a.i./bee), permethrin (50.01 ng a.i./bee), and imidacloprid (12.82 ng a.i/bee). The variation in the mass of individual bees had a significant but small effect on these toxicity estimates. Overall, the toxicity of these insecticides to M. rotundata were within the 10-fold safety factor currently used with A. mellifera toxicity estimates from tier I trials to estimate risk to other bee species. Therefore, tier I pesticide risk assessments with solitary bees may not be necessary, and efforts could be directed to developing more realistic, higher-tier pesticide risk assessment trials for solitary bees. creator: Graham R. Ansell creator: Andrew J. Frewin creator: Angela E. Gradish creator: Cynthia D. Scott-Dupree uri: https://doi.org/10.7717/peerj.10744 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Ansell et al. title: Psychometric properties of the Multi-group Ethnic Identity Measure (MEIM) in a sample of Iranian young adults link: https://peerj.com/articles/10752 last-modified: 2021-02-23 description: BackgroundThis study examines the factor structure, reliability and test-retest validity of the 12-item Iranian version of the Multigroup Ethnic Identity Measure (MEIM). Additionally, the MEIM’s concurrent validity was tested by investigating the association between ethnic identity and subjective well-being.MethodThe scale was translated into Persian language and was administered to 426 students (193 female) at a major public university in Tehran along with the Positive Affect Negative Affect Schedule, and the Satisfaction with Life Scale.ResultsThe confirmatory factor analysis supported the two-factor first-order commitment, and exploration (consisting of 12 items), and the second-order unidimensional factor structure of general ethnic identity. Moreover, we found evidence for good internal consistency, test re-test reliability, and concurrent validity.ConclusionThe MEIM Persian version was found to be a valid and reliable measure to examine ethnic identity in this Iranian student population, for both males and females. These results support the utility of the Persian version of the MEIM for its use in Middle-Eastern contexts. creator: Mojtaba Habibi creator: Maede Sadat Etesami creator: Mohammad Ali Taghizadeh creator: Faezeh Sadat Akrami creator: Danilo Garcia uri: https://doi.org/10.7717/peerj.10752 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Habibi et al. title: Evaluation of key miRNAs during early pregnancy in Kazakh horse using RNA sequencing link: https://peerj.com/articles/10796 last-modified: 2021-02-23 description: BackgroundmiRNA has an important role in cell differentiation, biological development, and physiology. Milk production is an important quantitative trait in livestock and miRNA plays a role in the amount of milk produced.MethodsThe role of regulatory miRNAs involved in equine milk production is not fully understood. We constructed two miRNA libraries for Kazakh horse milk production from higher-producing (H group) and lower-producing (L group) individuals, and used RNA-Seq technology to identify the differentially expressed miRNAs between the two milk phenotypes of Kazakh horses.ResultsA total of 341 known and 333 novel miRNAs were detected from the H and L groups, respectively. Eighty-three differentially expressed miRNAs were identified between the H and L group s, of which 32 were known miRNAs (27 were up-regulated, five were down-regulated) and 51 were novel miRNAs (nine were up-regulated, 42 were down-regulated). A total of 2,415 genes were identified. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses showed that these genes were annotated to mammary gland development, mammary gland morphogenesis, tissue development and PI3K-Akt signaling pathways, insulin signaling pathway and TGF-beta signaling pathway, among others. Five miRNAs (miR-199a-3p, miR143, miR145, miR221, miR486-5p) were identified as affecting horse milk production and these five miRNAs were validated using qRT-PCR.ConclusionsWe described a methodology for the transcriptome-wide profiling of miRNAs in milk, which may help the design of new intervention strategies to improve the milk yield of Kazakh horses. creator: LingLing Liu creator: Chao Fang creator: YinZe Sun creator: WuJun Liu uri: https://doi.org/10.7717/peerj.10796 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Liu et al. title: Behavioral dominance interactions between Nicrophorus orbicollis and N. tomentosus burying beetles (Coleoptera: Silphidae) link: https://peerj.com/articles/10797 last-modified: 2021-02-23 description: Asymmetric interference competition, where one species is behaviorally dominant over another, appears widespread in nature with the potential to structure ecological communities through trade-offs between competitive dominance and environmental tolerance. The details of how species interact and the factors that contribute to behavioral dominance, however, are poorly known for most species, yet such details are important for understanding when and why trade-offs occur. Here, we examine behavioral interactions between two species of burying beetles (Coleoptera: Silphidae) that compete for limited breeding resources (i.e., small vertebrate carcasses) in nature, to identify behaviors involved in interference competition and to test if large body size, species identity, or time of arrival best predict behavioral dominance among species. To test these ideas, we placed same-sex individuals of Nicrophorus orbicollis (early to mid-summer breeder) and N. tomentosus (late summer to fall breeder) into an enclosure together with a 25–30 g mouse carcass (Mus musculus). We then video-recorded all behaviors, including neutral and aggressive interactions, for 13 h per trial (N = 14 trials). For each interaction, we assigned a winner based on which beetle retained its position instead of fleeing or retained possession of the carcass; the overall behavioral dominant was determined as the individual that won the most interactions over the length of the trial. We found that large body size was the best predictor of behavioral dominance. In most interactions, N. orbicollis was larger and dominant over N. tomentosus; however, when N. tomentosus was larger they outcompeted smaller N. orbicollis, illustrating the importance of body size in aggressive contests. The order of arrival to the carcass (priority effects) did not predict behavioral dominance. The larger size and abundance of N. orbicollis in nature suggest a competitive asymmetry between the species, supporting the idea that N. orbicollis constrains the ability of N. tomentosus to breed earlier in the summer. creator: Scott D. Schrempf creator: Kevin W. Burke creator: Jillian D. Wettlaufer creator: Paul R. Martin uri: https://doi.org/10.7717/peerj.10797 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Schrempf et al. title: The long non-coding RNA MEG3 plays critical roles in the pathogenesis of cholesterol gallstone link: https://peerj.com/articles/10803 last-modified: 2021-02-23 description: BackgroundCholesterol gallstone (CG) is the most common gallstone disease, which is induced by biliary cholesterol supersaturation. The purpose of this study is to investigate the pathogenesis of CG.MethodsSixteen mice were equally and randomly divided into model group and normal control group. The model group was fed with lithogenic diets to induce CG, and then gallbladder bile lipid analysis was performed. After RNA-seq library was constructed, differentially expressed mRNAs (DE-mRNAs) and differentially expressed lncRNAs (DE-lncRNAs) between model group and normal control group were analyzed by DESeq2 package. Using the cluster Profiler package, enrichment analysis for the DE-mRNAs was carried out. Based on Cytoscape software, the protein-protein interaction (PPI) network and competing endogenous RNA (ceRNA) network were built. Using quantitative real-time reverse transcription-PCR (qRT-PCR) analysis, the key RNAs were validated.ResultsThe mouse model of CG was suc cessfully established, and then 181 DE-mRNAs and 33 DE-lncRNAs between model and normal groups were obtained. Moreover, KDM4A was selected as a hub node in the PPI network, and lncRNA MEG3 was considered as a key lncRNA in the regulatory network. Additionally, the miR-107-5p/miR-149-3p/miR-346-3-MEG3 regulatory pairs and MEG3-PABPC4/CEP131/NUMB1 co-expression pairs existed in the regulatory network. The qRT-PCR analysis showed that KDM4A expression was increased, and the expressions of MEG3, PABPC4, CEP131, and NUMB1 were downregulated.ConclusionThese RNAs might be related to the pathogenesis of CG. creator: Changlin Qian creator: Weiqing Qiu creator: Jie Zhang creator: Zhiyong Shen creator: Hua Liu creator: Yongjie Zhang uri: https://doi.org/10.7717/peerj.10803 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Qian et al. title: Characterization of the GRAS gene family reveals their contribution to the high adaptability of wheat link: https://peerj.com/articles/10811 last-modified: 2021-02-23 description: GRAS transcription factors play important roles in many processes of plant development as well as abiotic and biotic stress responses. However, little is known about this gene family in bread wheat (Triticum aestivum), one of the most important crops worldwide. The completion of a quality draft genome allows genome-wide detection and evolutionary analysis of the GRAS gene family in wheat. In this study, 188 TaGRAS genes were detected and divided into 12 subfamilies based on phylogenetic analyses: DELLA, DLT, HAM, LISCL, SCL3, SCL4/7, SCR, SHR, PAT1, Os19, Os4 and LAS. Tandem and segmental duplications are the main contributors to the expansion of TaGRAS, which may contribute to the adaptation of wheat to various environmental conditions. A high rate of homoeolog retention during hexaploidization was detected, suggesting the nonredundancy and biological importance of TaGRAS homoeologs. Systematic analyses of TaGRAS indicated the conserved expression pattern and function of the same subfamily during evolution. In addition, we detected five genes belonging to the LISCL subfamily induced by both biotic and abiotic stresses and they may be potential targets for further research through gene editing. Using degradome and ChIP-seq data, we identified the targets of miR171 and histone modifications and further analyzed the contribution of epigenetic modification to the subfunctionalization of TaGRAS. This study laid a foundation for further functional elucidation of TaGRAS genes. creator: Yanfeng Liu creator: Wei Wang uri: https://doi.org/10.7717/peerj.10811 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Liu and Wang title: A modularity analysis helps improving the structure of the International Code of Zoological Nomenclature link: https://peerj.com/articles/10815 last-modified: 2021-02-23 description: BackgroundIn a recent work I transformed a complex and integrated text like the International Code of Zoological Nomenclature into a network of interconnected parts of text. This new approach allowed understanding that a continuous body of text cannot accurately reflect the true structure of the Code, and provided a scientific methodology to identify a priori parts that could be affected by future revisions. In this next step, I investigate further the structure of the Code, seeking to use the network in order to identify the various conceptual communities grouping the various articles and other text items of the Code.MethodsUsing the first version of the network of the Code, I perform a comprehensive modularity analysis in two rounds: the first round aims to identify the fewest and largest communities or modules for the entire network, whereas the second round identifies the sub-modules within each larger module. The potential conflicts between the current structure of the Code and the module composition are evaluated with a parcellation analysis.ResultsThe optimal modularity search identified 10 different modules in the entire network of varying size (ranging from 75 to 200 nodes). Each module can be further divided into smaller modules, that all-together allow describing the 65 conceptual groups of text items in the Code. Parcellation analysis revealed that two-thirds of the current chapters of the Code are in excellent or good accordance with the recovered conceptual modules, whereas the current composition of six chapters is in serious conflict with the conceptual structure of the Code.DiscussionJudging only the composition and not the order of appearance of the Articles in the Chapters of the Code, I show that in many cases the current structure of the Code is found to correspond quite well to the concepts presented therein. The most important conflict is found on the provisions related to the various groups of names governed by the Code: family-, genus-, and species-group names. Currently, these provisions are spread out in different Articles in different Chapters, along the entire length of the Code. The modularity analysis suggests that re-organizing the Code in chapters that will deal with all aspects related to a given group (e.g., chapters including information on name formation, availability, typification, and validity for a given group), could potentially improve reader experience and, consequently, the applicability of the Code. creator: Evangelos Vlachos uri: https://doi.org/10.7717/peerj.10815 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Vlachos title: Knee replacement outcome predicted by physiotherapists: a prospective cohort study link: https://peerj.com/articles/10838 last-modified: 2021-02-23 description: BackgroundKnee arthroplasty (KA) is commonly used for osteoarthritis of the knee joint and it is a highly successful procedure. Still, KA leaves 20% of patients dissatisfied with their outcome. The purpose of this study was to determine if a prognosis made by physiotherapists at the orthopaedic wards during the first post-operative days could predict the 6- and 12-months outcome of KA.MethodsPhysiotherapists at two orthopaedic wards in Denmark were asked to predict the 6- and 12-months outcome of the KA patients they have treated post-operatively on a 0–10 scale (10 representing the best prognosis). At 6 and 12 months post-operatively the patients answered the Oxford Knee Score (OKS), EuroQol 5D-3L and Patient Acceptable Symptom State (PASS). Multivariable logistic regression analyses were performed to assess the prediction of PASS and treatment success. We assessed predictive performance by examining measures of calibration and discrimination.ResultsA total of 361 patients were included. The models for PASS and Treatment Success showed poor to acceptable discriminative values (OR between 1.47 and 1.92 and areas under the curves of 0.62–0.73), however the calibration plots indicated significant uncertainties in the prediction.ConclusionPhysiotherapists prognoses of recovery after KA are associated with 6- and 12-months patient reported outcomes and satisfaction but have weak predictive value. This study suggests that physiotherapists’ prognoses may be useful as an additional source of information when identifying patients in need of additional post-operative care. creator: Marius Henriksen creator: Hiwa Mukriyani creator: Carsten Juhl uri: https://doi.org/10.7717/peerj.10838 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Henriksen et al. title: Hemocyte siRNA uptake is increased by 5′ cholesterol-TEG addition in Biomphalaria glabrata, snail vector of schistosome link: https://peerj.com/articles/10895 last-modified: 2021-02-23 description: Biomphalaria glabrata is one of the snail intermediate hosts of Schistosoma mansoni, the causative agent of intestinal schistosomiasis disease. Numerous molecular studies using comparative approaches between susceptible and resistant snails to S. mansoni infection have helped identify numerous snail key candidates supporting such susceptible/resistant status. The functional approach using RNA interference (RNAi) remains crucial to validate the function of such candidates. CRISPR-Cas systems are still under development in many laboratories, and RNA interference remains the best tool to study B. glabrata snail genetics. Herein, we describe the use of modified small interfering RNA (siRNA) molecules to enhance cell delivery, especially into hemocytes, the snail immune cells. Modification of siRNA with 5′ Cholesteryl TriEthylene Glycol (Chol-TEG) promotes cellular uptake by hemocytes, nearly eightfold over that of unmodified siRNA. FACS analysis reveals that more than 50% of hemocytes have internalized Chol-TEG siRNA conjugated to Cy3 fluorophores, 2 hours only after in vivo injection into snails. Chol-TEG siRNA targeting BgTEP1 (ThioEster-containing Protein), a parasite binding protein, reduced BgTEP1 transcript expression by 70–80% compared to control. The level of BgTEP1 protein secreted in the hemolymph was also decreased. However, despite the BgTEP1 knock-down at both RNA and protein levels, snail compatibility with its sympatric parasite is not affected suggesting functional redundancy among the BgTEP genes family in snail-schistosoma interaction. creator: Anaïs Portet creator: Richard Galinier creator: Damien Lassalle creator: Alexandre Faille creator: Benjamin Gourbal creator: David Duval uri: https://doi.org/10.7717/peerj.10895 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Portet et al. title: MCQ-Balance: a method to monitor patients with balance disorders and improve clinical interpretation of posturography link: https://peerj.com/articles/10916 last-modified: 2021-02-23 description: BackgroundAn estimated 20% to 30% of the global population has suffered a vertiginous episode. Among them, 20% do not receive a clear diagnosis. Improved methods, indicators and metrics are necessary to assess the sensory systems related to balance, especially when patients are undergoing treatment for vertiginous episodes. Patients with balance disorders should be monitored for changes at the individual level to gather objective information. In this study, we evaluate the use of the MCQ-Balance (Measure, Classify and Qualify) assessment for examining a patient’s balance progression using tests to measure static balance control and dynamic postural balance with a stabilometric platform.Materials and MethodsThe MCQ-Balance assessment comprises three stages: (i) measuring the progression of each variable between two separate and consecutive days (called sessions) using the Magnitude-Based Decision analysis; (ii) classifying the progression of the patient’s balance with a score; and (iii) qualifying the progression of the patient’s balance from the resulting scores using a set of rules. This method was applied to 42 patients with balance disorders of peripheral or central origin characterised by vertigo as the cardinal symptom. Balance progression was measured using the MCQ-Balance assessment over the course of three months, and these results were compared with the assessment of a clinical expert.ResultsThe MCQ-Balance assessment showed an accuracy of 83.4% and a Cohen’s Kappa coefficient of 0.752 compared to the assessment of a clinical expert.ConclusionThe MCQ-Balance assessment facilitates the monitoring of patient balance and provides objective information that has the potential to improve medical decision making and the adjustment of individual treatment. creator: Juan De la Torre creator: Javier Marin creator: Marco Polo creator: Eva M. Gómez-Trullén creator: Jose J. Marin uri: https://doi.org/10.7717/peerj.10916 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 De la Torre et al. title: Identification of the hub gene BUB1B in hepatocellular carcinoma via bioinformatic analysis and in vitro experiments link: https://peerj.com/articles/10943 last-modified: 2021-02-23 description: BackgroundHepatocellular carcinoma (HCC) is one of the most commonly diagnosed cancers and the fourth leading cause of cancer-related deaths in the world. Although the treatment of HCC has made great progress in recent years, the therapeutic effects on HCC are still unsatisfactory due to difficulty in early diagnosis, chemoresistance and high recurrence rate post-surgery.MethodsIn this study, we identified differentially expressed genes (DEGs) based on four Gene Expression Omnibus (GEO) datasets (GSE45267, GSE98383, GSE101685 and GSE112790) between HCC and normal hepatic tissues. A protein–protein interaction (PPI) network was established to identify the central nodes associated with HCC. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of the central nodes were conducted to find the hub genes. The expression levels of the hub genes were validated based on the ONCOMINE and Gene Expression Profiling Interactive Analysis (GEPIA) databases. Additionally, the genetic alterations of the hub genes were evaluated by cBioPortal. The role of the hub genes on the overall survival (OS) and relapse survival (RFS) of HCC patients was evaluated by Kaplan-Meier plotter. At last, the mechanistic role of the hub genes was illustrated by in vitro experiments.ResultsWe found the following seven hub genes: BUB1B, CCNB1, CCNB2, CDC20, CDK1, MAD2L1 and RRM2 using integrated bioinformatics analysis. All of the hub genes were significantly upregulated in HCC tissues. And the seven hub genes were associated with the OS and RFS of HCC patients. Finally, in vitro experiments indicated that BUB1B played roles in HCC cell proliferation, migration, invasion, apoptosis and cell cycle by partially affecting mitochondrial functions.ConclusionsIn summary, we identified seven hub genes that were associated with the expression and prognosis of HCC. The mechanistic oncogenic role of BUB1B in HCC was first illustrated. BUB1B might play an important role in HCC and could be potential therapeutic targets for HCC. creator: Jie Fu creator: Xiao Zhang creator: Likun Yan creator: Yaoli Shao creator: Xinxu Liu creator: Yuan Chu creator: Ge Xu creator: Xundi Xu uri: https://doi.org/10.7717/peerj.10943 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Fu et al. title: Functional analysis of CgWRKY57 from Cymbidium goeringii in ABA response link: https://peerj.com/articles/10982 last-modified: 2021-02-23 description: BackgroundThe orchid is one of the top ten Chinese flowers and has high ornamental value and elegant color. However, orchids are vulnerable to abiotic stresses during their growth and development, and the molecular mechanism of the abiotic stress response in orchids is unclear. WRKY proteins belong to a transcription factor family that plays important roles in biotic stress, abiotic stress, growth and development in plants, but little is known about the WRKY family in Cymbidium goeringii.MethodsThe specific fragment of the CgWRKY57 gene of C. goeringii was analyzed by bioinformatics. The expression of the CgWRKY57 gene of C. goeringii under 4 °C, 42 °C water and ABA stress as well as different tissues was detected by real-time fluorescence quantitative PCR. CgWRKY57 gene was overexpressed in wild type Arabidopsis thaliana by inflorescence infection method, and the function of transgenic lines under ABA stress was analyzed.ResultsCgWRKY57 was cloned from C. goeringii and found to encode 303 amino acids. The CgWRKY57 protein is an acidic, nonsecreted hydrophilic protein without a signal peptide or transmembrane domain. The CgWRKY57 protein is located to the nucleus and may function intracellularly according to its predicted subcellular localization. A domain analysis and homology comparison showed that the CgWRKY57 protein has a “WRKYGQK” domain and belongs to Group III of the WRKY family, and a phylogenetic analysis demonstrated that CgWRKY57 is closely related to OsWRKY47. CgWRKY57 was expressed in the roots, stems, leaves and floral organs of C. goeringii, and its expression level was highest in the roots according to real-time qPCR analysis. There were significant differences in CgWRKY57 expression under 4 °C, 42 °C ABA and water stress treatments, and its expression changed greatly under ABA stress. The expression of CgWRKY57 in transgenic plants was significantly higher than that in wild type plants under ABA stress, and the root length and germination rate were reduced in transgenic plants compared to wild type plants.ConclusionsThese results indicate that CgWRKY57 overexpression is responsive to ABA stress, and they provide a foundation for future analyses of the biological functions of the WRKY family in C. goeringii. creator: Huanhuan Liu creator: Lianping Wang creator: Xijun Jing creator: Yue Chen creator: Fengrong Hu uri: https://doi.org/10.7717/peerj.10982 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Liu et al. title: Refining the marine reptile turnover at the Early–Middle Jurassic transition link: https://peerj.com/articles/10647 last-modified: 2021-02-22 description: Even though a handful of long-lived reptilian clades dominated Mesozoic marine ecosystems, several biotic turnovers drastically changed the taxonomic composition of these communities. A seemingly slow paced, within-geological period turnover took place across the Early–Middle Jurassic transition. This turnover saw the demise of early neoichthyosaurians, rhomaleosaurid plesiosaurians and early plesiosauroids in favour of ophthalmosaurid ichthyosaurians and cryptoclidid and pliosaurid plesiosaurians, clades that will dominate the Late Jurassic and, for two of them, the entire Early Cretaceous as well. The fossil record of this turnover is however extremely poor and this change of dominance appears to be spread across the entire middle Toarcian–Bathonian interval. We describe a series of ichthyosaurian and plesiosaurian specimens from successive geological formations in Luxembourg and Belgium that detail the evolution of marine reptile assemblages across the Early–Middle Jurassic transition within a single area, the Belgo–Luxembourgian sub-basin. These fossils reveal the continuing dominance of large rhomaleosaurid plesiosaurians, microcleidid plesiosaurians and Temnodontosaurus-like ichthyosaurians up to the latest Toarcian, indicating that the structuration of the upper tier of Western Europe marine ecosystems remained essentially constant up to the very end of the Early Jurassic. These fossils also suddenly record ophthalmosaurid ichthyosaurians and cryptoclidid plesiosaurians by the early Bajocian. These results from a geographically-restricted area provide a clearer picture of the shape of the marine reptile turnover occurring at the early–Middle Jurassic transition. This event appears restricted to the sole Aalenian stage, reducing the uncertainty of its duration, at least for ichthyosaurians and plesiosaurians, to 4 instead of 14 million years. creator: Valentin Fischer creator: Robert Weis creator: Ben Thuy uri: https://doi.org/10.7717/peerj.10647 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Fischer et al. title: Sensing and processing whisker deflections in rodents link: https://peerj.com/articles/10730 last-modified: 2021-02-22 description: The classical view of sensory information mainly flowing into barrel cortex at layer IV, moving up for complex feature processing and lateral interactions in layers II and III, then down to layers V and VI for output and corticothalamic feedback is becoming increasingly undermined by new evidence. We review the neurophysiology of sensing and processing whisker deflections, emphasizing the general processing and organisational principles present along the entire sensory pathway—from the site of physical deflection at the whiskers to the encoding of deflections in the barrel cortex. Many of these principles support the classical view. However, we also highlight the growing number of exceptions to these general principles, which complexify the system and which investigators should be mindful of when interpreting their results. We identify gaps in the literature for experimentalists and theorists to investigate, not just to better understand whisker sensation but also to better understand sensory and cortical processing. creator: Thomas F. Burns creator: Ramesh Rajan uri: https://doi.org/10.7717/peerj.10730 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Burns and Rajan title: Association between problematic internet use and behavioral/emotional problems among Chinese adolescents: the mediating role of sleep disorders link: https://peerj.com/articles/10839 last-modified: 2021-02-22 description: BackgroundStudies that focus on the relationships of problematic Internet use (PIU), sleep disorders, and behavioral/emotional problems were limited. This study aimed to explore (1) the relationship between PIU and behavioral/emotional problems among Chinese adolescents and (2) whether sleep disorders mediate the relationship between PIU and behavioral/emotional problems.MethodsA total of 1,976 adolescents were recruited by cluster sampling from ten secondary schools in Guangzhou between January and April 2019, and 1,956 of them provided valid information (response rate: 98.9% ). Among them, 50.8% were males and the mean age was 13.6±1.5 years, ranging from 11 to 18 years. Data on behavioral/emotional problems, sleep disorders, and PIU were collected using a self-reported questionnaire. Linear regression models and mediation analyses were performed.ResultsOf the participants, 14.5% (284/1,956) reported moderate to severe PIU, and their average score for total difficulties was significantly higher than the score for average users (14.9±5.5 Vs 9.8±4.7). After adjusting for controlled variables, PIU was further proven to be positively related to elevated levels of behavioral/emotional problems (unstandardized β = 0.16, p < 0.05). In addition, sleep disorders partially mediated the forgoing associations.ConclusionsAdolescents with problematic Internet habits were at higher risk of developing behavioral and emotional problems than their normal-use peers, and sleep disorders partially mediated the effect. Close attention and effective guidance for adolescents with PIU and behavioral/emotional problems were recommended for parents and schools. creator: Wanxin Wang creator: Xueying Du creator: Yangfeng Guo creator: Wenyan Li creator: Sheng Zhang creator: Lan Guo creator: Ciyong Lu uri: https://doi.org/10.7717/peerj.10839 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: The click is not the trick: the efficacy of clickers and other reinforcement methods in training naïve dogs to perform new tasks link: https://peerj.com/articles/10881 last-modified: 2021-02-22 description: BackgroundA handheld metal noisemaker known as a “clicker” is widely used to train new behaviors in dogs; however, evidence for their superior efficacy compared to providing solely primary reinforcement or other secondary reinforcers in the acquisition of novel behavior in dogs is largely anecdotal.MethodsThree experiments were conducted to determine under what circumstances a clicker secondary reinforcer may result in acquisition of a novel behavior more rapidly or to a higher level compared to other readily available reinforcement methods. In Experiment 1, three groups of 30 dogs each were shaped to emit a novel sit and stay behavior of increasing duration with either the delivery of food alone, a verbal stimulus paired with food, or a clicker with food. The group that received only a primary reinforcer reached a significantly higher criterion of training success than the group trained with a verbal secondary reinforcer. Performance of the group experiencing a clicker as a secondary reinforcer was intermediate between the other two groups, but not significantly different from either. In Experiment 2, three groups of 25 dogs each were shaped to emit a nose targeting behavior and then perform that behavior at increasing distances from the experimenter using the same three methods of positive reinforcement as in Experiment 1. No statistically significant differences between the groups were found. In Experiment 3, three groups of 30 dogs each were shaped to emit a nose-targeting behavior upon an array of wooden blocks with task difficulty increasing throughout testing using the same three methods of positive reinforcement as previously tested. No statistically significant differences between the groups were found.ResultsOverall, the findings suggest that both primary reinforcement alone as well as a verbal or clicker secondary reinforcer can be used successfully in training a dog to perform a novel behavior, but that no positive reinforcement method demonstrated significantly greater efficacy than any other. creator: Rachel J. Gilchrist creator: Lisa M. Gunter creator: Samantha F. Anderson creator: Clive D.L. Wynne uri: https://doi.org/10.7717/peerj.10881 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Gilchrist et al. title: Comparative transcriptome analysis of roots, stems, and leaves of Pueraria lobata (Willd.) Ohwi: identification of genes involved in isoflavonoid biosynthesis link: https://peerj.com/articles/10885 last-modified: 2021-02-22 description: BackgroundPueraria lobata (Willd.) Ohwi is a valuable herb used in traditional Chinese medicine. Isoflavonoids are the major bioactive compounds in P. lobata, namely puerarin, daidzin, glycitin, genistin, daidzein, and glycitein, which have pharmacological properties of anti-cardiovascular, anti-hypertension, anti-inflammatory, and anti-arrhythmic.MethodsTo characterize the corresponding genes of the compounds in the isoflavonoid pathway, RNA sequencing (RNA-Seq) analyses of roots, stems, and leaves of P. lobata were carried out on the BGISEQ-500 sequencing platform.ResultsWe identified 140,905 unigenes in total, of which 109,687 were annotated in public databases, after assembling the transcripts from all three tissues. Multiple genes encoding key enzymes, such as IF7GT and transcription factors, associated with isoflavonoid biosynthesis were identified and then further analyzed. Quantitative real-time PCR (qRT-PCR) results of some genes encoding key enzymes were consistent with our RNA-Seq analysis. Differentially expressed genes (DEGs) were determined by analyzing the expression profiles of roots compared with other tissues (leaves and stems). This analysis revealed numerous DEGs that were either uniquely expressed or up-regulated in the roots. Finally, quantitative analyses of isoflavonoid metabolites occurring in the three P. lobata tissue types were done via high-performance liquid-chromatography and tandem mass spectrometry methodology (HPLC-MS/MS). Our comprehensive transcriptome investigation substantially expands the genomic resources of P. lobata and provides valuable knowledge on both gene expression regulation and promising candidate genes that are involved in plant isoflavonoid pathways. creator: Chenkai Wang creator: Nenggui Xu creator: Shuai Cui uri: https://doi.org/10.7717/peerj.10885 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: Higher bee abundance, but not pest abundance, in landscapes with more agriculture on a late-flowering legume crop in tropical smallholder farms link: https://peerj.com/articles/10732 last-modified: 2021-02-19 description: BackgroundLandscape composition is known to affect both beneficial insect and pest communities on crop fields. Landscape composition therefore can impact ecosystem (dis)services provided by insects to crops. Though landscape effects on ecosystem service providers have been studied in large-scale agriculture in temperate regions, there is a lack of representation of tropical smallholder agriculture within this field of study, especially in sub-Sahara Africa. Legume crops can provide important food security and soil improvement benefits to vulnerable agriculturalists. However, legumes are dependent on pollinating insects, particularly bees (Hymenoptera: Apiformes) for production and are vulnerable to pests. We selected 10 pigeon pea (Fabaceae: Cajunus cajan (L.)) fields in Malawi with varying proportions of semi-natural habitat and agricultural area within a 1 km radius to study: (1) how the proportion of semi-natural habitat and agricultural area affects the abundance and richness of bees and abundance of florivorous blister beetles (Coleoptera: Melloidae), (2) if the proportion of flowers damaged and fruit set difference between open and bagged flowers are correlated with the proportion of semi-natural habitat or agricultural area and (3) if pigeon pea fruit set difference between open and bagged flowers in these landscapes was constrained by pest damage or improved by bee visitation.MethodsWe performed three, ten-minute, 15 m, transects per field to assess blister beetle abundance and bee abundance and richness. Bees were captured and identified to (morpho)species. We assessed the proportion of flowers damaged by beetles during the flowering period. We performed a pollinator and pest exclusion experiment on 15 plants per field to assess whether fruit set was pollinator limited or constrained by pests.ResultsIn our study, bee abundance was higher in areas with proportionally more agricultural area surrounding the fields. This effect was mostly driven by an increase in honeybees. Bee richness and beetle abundances were not affected by landscape characteristics, nor was flower damage or fruit set difference between bagged and open flowers. We did not observe a positive effect of bee density or richness, nor a negative effect of florivory, on fruit set difference.DiscussionIn our study area, pigeon pea flowers relatively late—well into the dry season. This could explain why we observe higher densities of bees in areas dominated by agriculture rather than in areas with more semi-natural habitat where resources for bees during this time of the year are scarce. Therefore, late flowering legumes may be an important food resource for bees during a period of scarcity in the seasonal tropics. The differences in patterns between our study and those conducted in temperate regions highlight the need for landscape-scale studies in areas outside the temperate region. creator: Cassandra Vogel creator: Timothy L. Chunga creator: Xiaoxuan Sun creator: Katja Poveda creator: Ingolf Steffan-Dewenter uri: https://doi.org/10.7717/peerj.10732 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Vogel et al. title: Retinoic Acid Induced Protein 14 (Rai14) is dispensable for mouse spermatogenesis link: https://peerj.com/articles/10847 last-modified: 2021-02-19 description: BackgroundRetinoic Acid Induced Protein 14 (Rai14) is an evolutionarily conserved gene that is highly expressed in the testis. Previous experiments have reported that small interfering RNA (siRNA)-mediated gene knockdown (KD) of Rai14 in rat testis disrupted spermatid polarity and transport. Of note, a gene knockout (KO) model is considered the “gold standard” for in vivo assessment of crucial gene functions. Herein, we used CRISPR/Cas9-based gene editing to investigate the in vivo role of Rai14 in mouse testis.MethodsSperm concentration and motility were assayed using a computer-assisted sperm analysis (CASA) system. Histological and immunofluorescence (IF) staining and transmission electron microscopy (TEM) were used to visualize the effects of Rai14 KO in the testes and epididymides. Terminal deoxynucleotidyl transferase-dUTP nick-end labeling (TUNEL) was used to determine apoptotic cells. Gene transcript levels were calculated by real-time quantitative PCR.ResultsRai14 KO in mice depicted normal fertility and complete spermatogenesis, which is in sharp contrast with the results reported previously in a Rai14 KD rat model. Sperm parameters and cellular apoptosis did not appear to differ between wild-type (WT) and KO group. Mechanistically, in contrast to the well-known role of Rai14 in modulating the dynamics of F-actin at the ectoplasmic specialization (ES) junction in the testis, morphological changes of ES junction exhibited no differences between Rai14 KO and WT testes. Moreover, the F-actin surrounded at the ES junction was also comparable between the two groups.ConclusionIn summary, our study demonstrates that Rai14 is dispensable for mouse spermatogenesis and fertility. Although the results of this study were negative, the phenotypic information obtained herein provide an enhanced understanding of the role of Rai14 in the testis, and researchers may refer to these results to avoid conducting redundant experiments. creator: Yangyang Wu creator: Ting Wang creator: Zigao Zhao creator: Siyu Liu creator: Cong Shen creator: Hong Li creator: Mingxi Liu creator: Bo Zheng creator: Jun Yu creator: Xiaoyan Huang uri: https://doi.org/10.7717/peerj.10847 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wu et al. title: Regucalcin expression profiles in veal calf testis: validation of histological and molecular tests to detect sex steroids illicit administration link: https://peerj.com/articles/10894 last-modified: 2021-02-19 description: BackgroundSex steroids administration in meat producing animals is forbidden within the EU to preserve consumers’ safety, but continuous monitoring to identify resurgence of their misuse is needed. Among biomarkers related to sex steroids abuse in veal calves the regucalcin (RGN) mRNA perturbations in testis have been described in RNAlater samples. To setup novel diagnostic method, to update current tests available in National Residue Control Plans (NRCPs) and in legal dispute when illicit practices on farm animals are suspected, the reliability of RGN profiling was assessed by histological and molecular techniques.MethodsFormalin fixed paraffin embedded (FFPE) testis samples, chosen being the most effective preservation strategy adopted by histological NRCPs and allowing easier retrospective analysis if required by legal disputes, were analyzed from veal calves treated with nandrolone, 17β-estradiol and a cocktail of the two hormones. RGN levels were determined by quantitative Real Time PCR and Immunohistochemistry assays. Test performances were assessed and compared by multiple ROC curves.ResultsBoth tests resulted sensitive and specific, allowing to enrich, in future field investigation, novel integrated diagnostic protocols needed to unveil sex steroid abuse.DiscussionDeveloped RT-qPCR and IHC methods confirmed RGN as a useful and robust biomarker to detect illegal administration of sex steroid hormones in veal calves. The developed methods, successfully applied to ten years old FFPE blocks, could allow both retrospective analysis, when supplementary investigations are requested by authorities, and future implementation of current NRCPs. creator: Alessandro Benedetto creator: Elena Biasibetti creator: Chiara Beltramo creator: Valentina Audino creator: Simone Peletto creator: Elena Maria Bozzetta creator: Marzia Pezzolato uri: https://doi.org/10.7717/peerj.10894 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Benedetto et al. title: Metasychis varicollaris sp. nov. and report of Metasychis gotoi (Maldanidae, Annelida) from the China Seas link: https://peerj.com/articles/10608 last-modified: 2021-02-18 description: Polychaete species are widely distributed throughout Indo-Pacific and European waters. We collected Metasychis specimens from the China Seas to report on Metasychis varicollaris sp. n. and Metasychis gotoi (Izuka, 1902) in greater detail. Geographic analysis of the potential distribution areas of M. gotoi indicates that it may be found in most coastal areas of China. The newly discovered species, M. varicollaris and M. gotoi, have an overlapping distribution in the northern South China Sea. Metasychis varicollaris sp. n. is characterized by a crenulated cephalic rim, complete collar on chaetiger 1, a packet-shaped anal funnel, and a spirally-fringed notochaetae with spiral pectinate bands imbricated over the main shaft. Our study provides a taxonomic key to all species of Metasychis. creator: Yueyun Wang creator: Xinzheng Li creator: Chunsheng Wang uri: https://doi.org/10.7717/peerj.10608 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Wang et al. title: QTL mapping of agronomic traits in wheat using the UK Avalon ×  Cadenza reference mapping population grown in Kazakhstan link: https://peerj.com/articles/10733 last-modified: 2021-02-18 description: BackgroundThe success of wheat production is largely dependent on local breeding projects that focus on the development of high-yielding cultivars with the use of novel molecular tools. One strategy for improving wheat productivity involves the deployment of diverse germplasms with a high potential yield. An important factor for achieving success involves the dissection of quantitative trait loci (QTLs) for complex agronomic traits, such as grain yield components, in targeted environments for wheat growth.MethodsIn this study, we tested the United Kingdom (UK) spring set of the doubled haploid (DH) reference population derived from the cross between two British cultivars, Avalon (winter wheat) and Cadenza (spring wheat), in the Northern, Central, and Southern regions (Karabalyk, Karaganda, Kyzylorda) of Kazakhstan over three years (2013–2015). The DH population has previously been genotyped by UK scientists using 3647 polymorphic DNA markers. The list of tested traits includes the heading time, seed maturation time, plant height, spike length, productive tillering, number of kernels per spike, number of kernels per meter, thousand kernel weight, and yield per square meter. Windows QTL Cartographer was applied for QTL mapping using the composite interval mapping method.ResultsIn total, 83 out of 232 QTLs were identified as stable QTLs from at least two environments. A literature survey suggests that 40 QTLs had previously been reported elsewhere, indicating that this study identified 43 QTLs that are presumably novel marker-trait associations (MTA) for these environments. Hence, the phenotyping of the DH population in new environments led to the discovery of novel MTAs. The identified SNP markers associated with agronomic traits in the DH population could be successfully used in local Kazakh breeding projects for the improvement of wheat productivity. creator: Akerke Amalova creator: Saule Abugalieva creator: Vladimir Chudinov creator: Grigoriy Sereda creator: Laura Tokhetova creator: Alima Abdikhalyk creator: Yerlan Turuspekov uri: https://doi.org/10.7717/peerj.10733 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Amalova et al. title: Effects of training frequency on muscular strength for trained men under volume matched conditions link: https://peerj.com/articles/10781 last-modified: 2021-02-18 description: BackgroundIn resistance training, the role of training frequency to increase maximal strength is often debated. However, the limited data available does not allow for clear training frequency “optimization” recommendations. The purpose of this study was to investigate the effects of training frequency on maximal muscular strength and rate of perceived exertion (RPE). The total weekly training volume was equally distributed between two and four sessions per muscle group.MethodsTwenty-one experienced resistance-trained male subjects (height: 1.85 ± 0.06 m, body mass: 85.3 ± 12.3 kg, age: 27.6 ± 7.6 years) were tested prior to and after an 8-week training period in one-repetition maximum (1RM) barbell back squat and bench press. Subjects were randomly assigned to a SPLIT group (n = 10), in which there were two training sessions of squats and lower-body exercises and two training sessions of bench press and upper-body exercises, or a FULLBODY group (n = 11), in which four sessions with squats, bench press and supplementary exercises were conducted every session. In each session, the subjects rated their RPE after barbell back squat, bench press, and the full session.ResultsBoth groups significantly increased 1RM strength in barbell back squat (SPLIT group: +13.25 kg; FULLBODY group: +14.31 kg) and bench press (SPLIT group: +7.75 kg; FULLBODY group: +8.86 kg) but training frequency did not affect this increase for squat (p = 0.640) or bench press (p = 0.431). Both groups showed a significant effect for time on RPE on all three measurements. The analyses showed only an interaction effect between groups on time for the RPE after the squat exercise (p = 0.002).ConclusionWe conclude that there are no additional benefits of increasing the training frequency from two to four sessions under volume-equated conditions, but it could be favorable to spread the total training volume into several training bouts through the week to avoid potential increases in RPE, especially after the squat exercise. creator: Emil Johnsen creator: Roland van den Tillaar uri: https://doi.org/10.7717/peerj.10781 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Johnsen and van den Tillaar title: Estimation of parameters for a humidity-dependent compartmental model of the COVID-19 outbreak link: https://peerj.com/articles/10790 last-modified: 2021-02-18 description: Building an effective and highly usable epidemiology model presents two main challenges: finding the appropriate, realistic enough model that takes into account complex biological, social and environmental parameters and efficiently estimating the parameter values with which the model can accurately match the available outbreak data, provide useful projections. The reproduction number of the novel coronavirus (SARS-CoV-2) has been found to vary over time, potentially being influenced by a multitude of factors such as varying control strategies, changes in public awareness and reaction or, as a recent study suggests, sensitivity to temperature or humidity changes. To take into consideration these constantly evolving factors, the paper introduces a time dynamic, humidity-dependent SEIR-type extended epidemiological model with range-defined parameters. Using primarily the historical data of the outbreak from Northern and Southern Italy and with the help of stochastic global optimization algorithms, we are able to determine a model parameter estimation that provides a high-quality fit to the data. The time-dependent contact rate showed a quick drop to a value slightly below 2. Applying the model for the COVID-19 outbreak in the northern region of Italy, we obtained parameters that suggest a slower shrinkage of the contact rate to a value slightly above 4. These findings indicate that model fitting and validation, even on a limited amount of available data, can provide useful insights and projections, uncover aspects that upon improvement might help mitigate the disease spreading. creator: Csaba Farkas creator: David Iclanzan creator: Boróka Olteán-Péter creator: Géza Vekov uri: https://doi.org/10.7717/peerj.10790 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Farkas et al. title: Microbiomes of different ages in Rendzic Leptosols in the Crimean Peninsula link: https://peerj.com/articles/10871 last-modified: 2021-02-18 description: Rendzic Leptosols are intrazonal soils formed on limestone bedrock. The specialty of these soils is that parent rock material is more influential in shaping soil characteristics than zonal factors such as climate, especially during soil formation. Unlike fast evolving Podzols due to their leaching regime, Leptosols do not undergo rapid development due to the nature of the limestone. Little is known how microbiome reflects this process, so we assessed microbiome composition of Rendzic Leptosols of different ages, arising from disruption and subsequent reclamation. The mountains and foothills that cover much of the Crimean Peninsula are ideal for this type of study, as the soils were formed on limestone and have been subjected to anthropogenic impacts through much of human history. Microbiomes of four soil sites forming a chronosequence, including different soil horizons, were studied using sequencing of 16S rRNA gene libraries and quantitative PCR. Dominant phyla for all soil sites were Actinobacteria, Proteobacteria, Acidobacteria, Bacteroidetes, Thaumarchaeota, Planctomycetes, Verrucomicrobia and Firmicutes. Alpha diversity was similar across sites and tended to be higher in topsoil. Beta diversity showed that microbiomes diverged according to the soil site and the soil horizon. The oldest and the youngest soils had the most similar microbiomes, which could have been caused by their geographic proximity. Oligotrophic bacteria from Chitinophagaceae, Blastocatellaceae and Rubrobacteriaceae dominated the microbiome of these soils. The microbiome of 700-year old soil was the most diverse. This soil was from the only study location with topsoil formed by plant litter, which provided additional nutrients and could have been the driving force of this differentiation. Consistent with this assumption, this soil was abundant in copiotrophic bacteria from Proteobacteria and Actinobacteria phyla. The microbiome of 50-year old Leptosol was more similar to the microbiome of benchmark soil than the microbiome of 700-year old soil, especially by weighted metrics. CCA analysis, in combination with PERMANOVA, linked differences in microbiomes to the joint change of all soil chemical parameters between soil horizons. Local factors, such as parent material and plant litter, more strongly influenced the microbiome composition in Rendzic Leptosols than soil age. creator: Anastasiia K. Kimeklis creator: Grigory V. Gladkov creator: Aleksei O. Zverev creator: Arina A. Kichko creator: Evgeny E. Andronov creator: Elena I. Ergina creator: Igor V. Kostenko creator: Evgeny V. Abakumov uri: https://doi.org/10.7717/peerj.10871 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Kimeklis et al. title: Pseudocnidae of ribbon worms (Nemertea): ultrastructure, maturation, and functional morphology link: https://peerj.com/articles/10912 last-modified: 2021-02-18 description: The fine structure of mature pseudocnidae of 32 species of nemerteans, representatives of 20 genera, six families, and two classes was investigated with scanning and transmission electron microscopy. Pseudocnidae are composed of four layers (cortex, medulla, precore layer, and core) in most species investigated, but the degree of development and position of each layer can vary between different species. The secretion products comprising immature pseudocnidae segregate into separate layers: a thin envelope, which subsequently separates into the cortex and medulla and an extensive internal layer. We distinguish two pseudocnida types: type I is characterized by a two-layered core and type II by a three-layered core. Type I pseudocnidae are present in archinemertean species, Carinoma mutabilis, and in all pilidiophoran species, except Heteronemertea sp. 5DS; type II pseudocnidae occur in all studied species of Tubulanidae and the basal Heteronemertea sp. 5DS. Based on the structure of the discharged pseudocnidae observed in eleven species of palaeonemerteans and in eight species of pilidiophorans, we distinguish three different mechanisms (1–3) of core extrusion/discharge with the following characteristics and distribution: (1) the outer core layer is everted simultaneously with the tube-like layer and occurs in type I pseudocnidae of most species; (2) the extruded core is formed by both eversion of the outer core layer and medullar layer, and occurs in type I pseudocnidae of Micrura cf. bella; (3) the eversion of the outer core layer begins together with the core rod and core rod lamina and occurs in type II pseudocnidae. Morpho-functional comparison with other extrusomes (cnidae, sagittocysts, rhabdtites, and paracnids) confirm that pseudocnidae are homologous structures that are unique to nemerteans. creator: Timur Yu Magarlamov creator: James M. Turbeville creator: Alexei V. Chernyshev uri: https://doi.org/10.7717/peerj.10912 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Magarlamov et al. title: The tetrapod fauna of the upper Permian Naobaogou Formation of China: 6. Turfanodon jiufengensis sp. nov. (Dicynodontia) link: https://peerj.com/articles/10854 last-modified: 2021-02-17 description: The dicynodont fossils from the Naobaogou Formation of Nei Mongol, China are abundant and diverse but poorly studied. In this article, one nearly complete skeleton and four cranial specimens from the Naobaogou Formation are referred to the dicynodontoid genus Turfanodon as a new species, T. jiufengensis. Previously, Turfanodon was known only from upper Permian sites in Xinjiang and Gansu. The new specimens are referred to Turfanodon based on the following characters: snout tall with steeply sloping profile, anterior tip of the snout squared off, facial region heavily pitted, nasal bosses present as paired swellings near the posterodorsal margin of the external nares, preparietal depressed, intertemporal bar long and narrow, premaxilla contacting frontal, palatal surface of premaxilla exposed in lateral view, and anterior pterygoid keel restricted to the anterior tip of the anterior ramus of the pterygoid. Turfanodon jiufengensis is differentiated from the type species, T. bogdaensis, by a contact of the lacrimal with the septomaxilla, discrete, raised nasal bosses, the dorsal edge of the erupted portion of the canine tusk slightly posterior to the anterior orbital margin, an anterior extension of the lacrimal distinctly shorter than that of the prefrontal, and a premaxillary dorsal surface with a median ridge. The holotype skeleton of T. jiufengensis includes a complete axial column with 50 vertebrae (six cervical, 23 dorsal, six sacral, and 15 caudal). Turfanodon represents the first confirmed tetrapod genus shared by the late Permian faunas of the Junggar and Ordos basins, and appears to be the first dicynodont genus distributed across both tropical and temperate zones (based on paleoclimate reconstructions). Based on tetrapod fossil content, the Naobaogou Formation can be roughly correlated to the Daptocephalus Assemblage Zone of South Africa (255–252 Ma in age). creator: Jun Liu uri: https://doi.org/10.7717/peerj.10854 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Liu title: Assessment of the impact of land use in an agricultural catchment area on water quality of lowland rivers link: https://peerj.com/articles/10564 last-modified: 2021-02-17 description: In several countries around the world, agricultural land area exceeds 70% (Uruguay 82.6%, Kazakhstan 80.4%, Turkmenistan 72.0%, Great Britain 71.7%, Ukraine 71.6% and others). This poses a serious risk of dissipating nitrates into the aquatic environment in agricultural catchments. The aim of this study was to assess the impact of land use on water quality parameters in an agricultural catchment area. It was decided to select for analysis the catchment of the Orla River (river length of 88 km, catchment area of 1,546 km2). The catchment area is predominantly agricultural in character and its entire area has been declared as an agricultural nitrate vulnerable zone (NVZ). A total of 27 survey sites were selected on the main watercourse and its tributaries. Analyses were conducted in the years 2010–2012 to determine physical and chemical parameters of water (pH reaction, conductivity, dissolved oxygen, total nitrogen, organic nitrogen, ammonia nitrogen, nitrates, total and reactive phosphorus) as well as six macrophyte metrics of ecological status assessment (MIR, IBMR, RMNI, MTR, TIM, RI). The average values of most physico-chemical parameters of water quality repeatedly exceeded limits of good ecological status, both in the Orla River and its tributaries. As many as 18 survey sites were classified as moderate ecological status, five sites as poor and only four as good ecological status. The results indicate the impact of land use in the catchment on water conductivity. Differences were observed in the concentrations of biotic components in the main watercourse and its tributaries, and in water quality in the southern part of the catchment in relation to the rest of the study area. This is probably connected with a greater share of forests and surface waters in that area. creator: Jerzy M. Kupiec creator: Ryszard Staniszewski creator: Szymon Jusik uri: https://doi.org/10.7717/peerj.10564 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Kupiec et al. title: Scoping review of distribution models for selected Amblyomma ticks and rickettsial group pathogens link: https://peerj.com/articles/10596 last-modified: 2021-02-17 description: The rising prevalence of tick-borne diseases in humans in recent decades has called attention to the need for more information on geographic risk for public health planning. Species distribution models (SDMs) are an increasingly utilized method of constructing potential geographic ranges. There are many knowledge gaps in our understanding of risk of exposure to tick-borne pathogens, particularly for those in the rickettsial group. Here, we conducted a systematic scoping review of the SDM literature for rickettsial pathogens and tick vectors in the genus Amblyomma. Of the 174 reviewed articles, only 24 studies used SDMs to estimate the potential extent of vector and/or pathogen ranges. The majority of studies (79%) estimated only tick distributions using vector presence as a proxy for pathogen exposure. Studies were conducted at different scales and across multiple continents. Few studies undertook original data collection, and SDMs were mostly built with presence-only datasets from public database or surveillance sources. The reliance on existing data sources, using ticks as a proxy for disease risk, may simply reflect a lag in new data acquisition and a thorough understanding of the tick-pathogen ecology involved. creator: Catherine A. Lippi creator: Holly D. Gaff creator: Alexis L. White creator: Sadie J. Ryan uri: https://doi.org/10.7717/peerj.10596 license: https://creativecommons.org/licenses/by-nc/4.0 rights: © 2021 Lippi et al. title: Density estimation of tiger and leopard using spatially explicit capture–recapture framework link: https://peerj.com/articles/10634 last-modified: 2021-02-17 description: The conservation of large carnivores often requires precise and accurate estimates of their populations. Being cryptic and occurring at low population densities, obtaining an unbiased population estimate is difficult in large carnivores. To overcome the uncertainties in the conventional capture–recapture (CR) methods used to estimate large carnivore densities, more robust methods such as spatially explicit capture-recapture (SECR) framework are now widely used. We modeled the CR data of tiger (Panthera tigris tigris) and leopard (Panthera pardus fusca) in the SECR framework with biotic and abiotic covariates likely believed to influence their densities. An effort of 2,211 trap nights resulted in the capture of 33 and 38 individual tigers and leopards. A total of 95 and 74 detections of tigers and leopards were achieved using 35 pairs of camera traps. Tiger and leopard density were estimated at 4.71 ± 1.20 (3.05–5.11) and 3.03 ± 0.78 (1.85–4.99) per 100 km2. Our results show that leopard density increased with high road density, high terrain ruggedness and habitats with high percentage of cropland and natural vegetation. The tiger density was positively influenced by the mosaic of cropland and natural vegetation. This study provides the first robust density estimates of tiger and leopard within the study area. Our results support the notion that large carnivores can attain moderate densities within human-dominated regions around protected areas relying on domestic livestock. Broader management strategies aimed at maintaining wild prey in the human-dominated areas around protected areas are necessary for large and endangered carnivores’ sustenance in the buffer zones around protected areas. creator: Tahir Ali Rather creator: Sharad Kumar creator: Jamal Ahmad Khan uri: https://doi.org/10.7717/peerj.10634 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Rather et al. title: Sleep disturbance and its association with quality of life among psychiatric nurses in China link: https://peerj.com/articles/10659 last-modified: 2021-02-17 description: BackgroundSleep disturbance is associated with a number of negative adverse outcomes. This study examined the prevalence of sleep disturbance and its association with demographic and clinical characteristics and quality of life (QOL) in psychiatric nurses in China.MethodsThis is a multi-center, cross-sectional study involving 11 psychiatric hospitals in China. Three types of sleep disturbance (difficulty initiating sleep (DIS), difficulty maintaining sleep (DMS) and early-morning awakening (EMA)) and QOL were measured by standardized questions or instruments.ResultsA total of 1,847 psychiatric nurses participated. The overall prevalence of at least one type of sleep disturbance was 71.5% (95% CI [69.3–73.5]); the prevalence of DIS, DMS and EMA was 58.5% (95% CI [56.2–60.8]), 53.7% (95% CI [51.4–56.0]) and 54.6% (95% CI [52.3–56.9]), respectively. Nurses with sleep disturbance had significantly lower QOL in physical (F(1, 1,846) = 219.12, P < 0.001), psychological (F(1, 1,846) = 72.18, P < 0.001), social (F(1, 1,846) = 37.57, P < 0.001) and environmental domains (F(1, 1,846) = 95.45, P < 0.001). Multivariable logistic regression analyses revealed that shift work (DIS, OR = 1.6, 95% CI [1.28–1.98]; DMS, OR = 1.2, 95% CI [1.001–1.54]; EMA, OR = 1.3, 95% CI [1.02–1.58]) and alcohol use (DIS, OR = 1.8, 95% CI [1.46–2.32]; DMS, OR = 1.8, 95% CI [1.43–2.23]; EMA, OR = 1.7, 95% CI [1.33–2.07]) were positively associated with sleep disturbance, while higher monthly income (DIS, OR = 0.5, 95% CI [0.38–0.75]; DMS, OR = 0.7, 95% CI [0.51–0.98]) was negatively associated with sleep disturbance.ConclusionSleep disturbance was common among nurses in psychiatric hospitals in China, particularly those on shifts and having alcohol use. Health authorities should develop effective measures to reduce risk of sleep disturbance in this population. creator: Li Lu creator: Ka-In Lok creator: Qinge Zhang creator: Ling Zhang creator: Yifan Xiang creator: Gabor S. Ungvari creator: Brian J. Hall creator: Feng-Rong An creator: Yu-Tao Xiang uri: https://doi.org/10.7717/peerj.10659 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Lu et al. title: Analysis in silico of the functional interaction between WNT5A and YAP/TEAD signaling in cancer link: https://peerj.com/articles/10869 last-modified: 2021-02-17 description: To date, most data regarding the crosstalk between the Wnt signaling pathway and the YAP/TAZ transcriptional coactivators focuses on the Wnt/β-catenin branch of the pathway. In contrast, the relationship between the non-canonical Wnt pathway and YAP/TAZ remains significantly less explored. Wnt5a is usually regarded as a prototypical non-canonical Wnt ligand, and its expression has been related to cancer progression. On the other hand, YAP/TAZ transcriptional coactivators act in concert with TEAD transcription factors to control gene expression. Although one article has shown previously that WNT5A is a YAP/TEAD target gene, there is a need for further evidence supporting this regulatory relationship, because a possible YAP/Wnt5a regulatory circuit might have profound implications for cancer biology. This article analyzes publicly available ChIP-Seq, gene expression, and protein expression data to explore this relationship, and shows that WNT5A might be a YAP/TEAD target gene in several contexts. Moreover, Wnt5a and YAP expression are significantly correlated in specific cancer types, suggesting that the crosstalk between YAP/TAZ and the Wnt pathway is more intricate than previously thought. creator: Pablo Astudillo uri: https://doi.org/10.7717/peerj.10869 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Astudillo title: Expression, purification, crystallization and preliminary X-ray crystallographic studies of a mitochondrial membrane-associated protein Cbs2 from Saccharomyces cerevisiae link: https://peerj.com/articles/10901 last-modified: 2021-02-17 description: BackgroundMitochondria are unique organelles that are found in most eukaryotic cells. The main role of the mitochondria is to produce ATP. The nuclear genome encoded proteins Cbs1 and Cbs2 are located at the mitochondrial inner membrane and are reported to be essential for the translation of mitochondrial cytochrome b mRNA. Genetic studies show that Cbs2 protein recognizes the 5′ untranslated leader sequence of mitochondrial cytochrome b mRNA. However, due to a lack of biochemical and structural information, this biological process remains unclear. To investigate the structural characteristics of how Saccharomyces cerevisiae (S. cerevisiae) Cbs2 tethers cytochrome b mRNA to the mitochondrial inner membrane, a preliminary X-ray crystallographic study was carried out and is reported here.MethodsThe target gene from S. cerevisiae was amplified by polymerase chain reaction. The PCR fragment was digested by the NdeI and XhoI restriction endonucleases and then inserted into expression vector p28. After sequencing, the plasmid was transformed into Escherichia coli C43 competent cells. The selenomethionine derivative Cbs2 protein was overexpressed using M9 medium based on a methionine-biosynthesis inhibition method. The protein was first purified to Ni2+-nitrilotriacetate affinity chromatography and then further purified by Ion exchange chromatography and Gel-filtration chromatography. The purified Se-Cbs2 protein was concentrated to 10 mg/mL. The crystallization trials were performed using the sitting-drop vapor diffusion method at 16 °C. The complete diffraction data was processed and scaled with the HKL2000 package and programs in the CCP4 package, respectively.ResultsCbs2 from S. cerevisiae was cloned, prokaryotic expressed and purified. The analysis of the size exclusion chromatography showed that the Cbs2 protein peaked at a molecular weight of approximately 90 KDa. The crystal belonged to the space group C2, with unit-cell parameters of a = 255.11, b = 58.10, c = 76.37, and β = 95.35°. X-ray diffraction data was collected at a resolution of 2.7 Å. The Matthews coefficient and the solvent content were estimated to be 3.22 Å 3 Da-1 and 61.82%, respectively.ConclusionsIn the present study Cbs2 from S. cerevisiae was cloned, expressed, purified, and crystallized for structural studies. The molecular weight determination results indicated that the biological assembly of Cbs2 may be a dimer.The preliminary X-ray crystallographic studies indicated the presence of two Cbs2 molecules in the asymmetric unit. This study will provide an experimental basis for exploring how Cbs2 protein mediates cytochrome b synthesis. creator: Dan Wu creator: Guanyu Zhu creator: Yufei Zhang creator: Yan Wu creator: Chunlei Zhang creator: Jiayi Shi creator: Xiaofeng Zhu creator: Xiaohuan Yuan uri: https://doi.org/10.7717/peerj.10901 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wu et al. title: Mental health status and quality of life in elderly patients with coronary heart disease link: https://peerj.com/articles/10903 last-modified: 2021-02-17 description: BackgroundCoronary heart disease (CHD) is the leading cause of morbidity and mortality among elderly individuals. Patients with CHD are at high risk for mental health disorders, and psychological issues may affect the quality of life (QoL) of these patients. Nevertheless, there is little evidence regarding the psychological issues and QoL of patients with CHD among the elderly population. This study aimed to investigate the relationship between comorbidities and mental status as well as QoL among elderly patients with CHD.MethodsOverall, 216 patients were included in this cross-sectional, observational, single-center study. The demographics and clinical manifestations of the patients were collected from electronic medical records. All patients were interviewed using the Chinese version of Symptom Checklist 90 (SCL-90) to assess the mental health status and the World Health Organization Quality of Life-BREF questionnaire (WHOQOL-BREF) to assess the QoL.ResultsIn total, 96 men and 120 women, with a mean age of 71.69 ±  8.30 years, were included. When controlling for the patients’ sex, marital status and stroke, multiple stepwise linear regression analyses suggested that for CHD patients, comorbid type 2 diabetes mellitus had the significant influence on average positive factors (Coef., 5.809; 95% CI [2.467–9.152] p = 0.001); when controlling for the patients’ sex, marital status and type 2 diabetes mellitus, multiple stepwise linear regression analyses suggested that for CHD patients, comorbid stroke had the significant influence on average positive factors (Coef., 8.680; 95% CI [4.826–12.534]; p < 0.001); when controlling for the patients’ sex, marital status, type 2 diabetes mellitus and stroke, multiple stepwise linear regression analyses suggested that for CHD patients, comorbid primary hypertension had the significant influence on phobic anxiety (Coef., 0.178; 95% CI [0.010–0.347]; p = 0.038).ConclusionsFor elderly CHD patients, comorbid type 2 diabetes mellitus and stroke were at risk for psychological problems and lower QoL. Our findings may guide patients and clinicians to make better decisions and achieve better outcomes. creator: Min Tang creator: Song-Hao Wang creator: Hui-Lin Li creator: Han Chen creator: Xin-Yi Sun creator: Wei-Wei Bian creator: Jing Sheng creator: Shao-Jun Ma uri: https://doi.org/10.7717/peerj.10903 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Tang et al. title: Biomarker identification of isolated compartments of the cell wall, cytoplasm and vacuole from the internodal cell of characean Nitellopsis obtusa link: https://peerj.com/articles/10930 last-modified: 2021-02-17 description: Cells of characean algae are attractive for plant cell physiologists because of their large size and their close relation to higher plant cells. The objective of our study was to evaluate the purity of the compartments (cell wall, cytoplasm with plastids, mitochondria, nuclei and endomembrane system, and vacuole) separated mechanically from the internodal cells of Nitellopsis obtusa using enzymatic markers. These included α-mannosidase and malate dehydrogenase, vacuolar and cytoplasmic enzymes, respectively. The biomarkers applied revealed the degree of compartment contamination with the material from unwanted cell parts. The cell wall was contaminated slightly by vacuole and cytoplasm residuals, respectively by 12.3 and 1.96% of corresponding biomarker activities. Relatively high activity of vacuolar marker in the cell wall could be associated with the cell vacuoles in the multicellular structure of the nodes. The biomarkers confirmed highly purified vacuolar (99.5%) and cytoplasmic (86.7%) compartments. Purity estimation of the cell fractions enabled reevaluating nCuO related Cu concentrations in the compartments of charophyte cell. The internalisation of CuO nanoparticles in N. obtusa cell occurred already after 0.5h. In general, the approach seems to be useful for assessing the accumulation and distribution of various xenobiotics and/or metabolites within plant cell. All this justifies N.obtusa internodal cells as a model organism for modern studies in cell biology and nanotoxicology. creator: Brigita Gylytė creator: Sigita Jurkonienė creator: Reda Cimmperman creator: Vaidevutis Šveikauskas creator: Levonas Manusadžianas uri: https://doi.org/10.7717/peerj.10930 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Gylytė et al. title: Comprehensive analysis of AHL gene family and their expression under drought stress and ABA treatment in Populus trichocarpa link: https://peerj.com/articles/10932 last-modified: 2021-02-17 description: The AT-hook motif nuclear-localized (AHL) family is a plant transcription factor family, which plays an important role in growth and development and stress responses. We identified and analyzed 37 AHL genes in poplar (Populus trichocarpa). Phylogenetic analysis classified the PtrAHL members into three subfamilies based on their conserved domain. All PtrAHL paralogous pairs evolved under purifying selection. The promoter analysis revealed the presence of stress-related and phytohormone-related cis-elements of the PtrAHL genes. Our analysis of the tissue-specific expression pattern of PtrAHL genes indicated their significance in tissue and organ development. Network-based prediction suggested that PtrAHL genes may interact with histone deacetylases (HDAC) and participate in the development of organs, such as roots. Drought negatively impacts plant growth and development. ABA is produced under osmotic stress condition, and it takes an important part in the stress response and tolerance of plants. Real-time quantitative PCR (qRT-PCR) showed that PtrAHL genes were induced by drought stress and ABA treatment. These insights into the expression of PtrAHL genes under stress provide a basis for PtrAHL gene functional analysis. Our study will help develop new breeding strategies to improve drought tolerance in poplar. creator: Hanzeng Wang creator: Xue Leng creator: Jia Yang creator: Mengqiu Zhang creator: Minzhen Zeng creator: Xuemei Xu creator: Fude Wang creator: Chenghao Li uri: https://doi.org/10.7717/peerj.10932 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: Phylogenetically-controlled correlates of primate blinking behaviour link: https://peerj.com/articles/10950 last-modified: 2021-02-17 description: Eye blinking is an essential maintenance behaviour for many terrestrial animals, but is also a risky behaviour as the animal is unable to scan the environment and detect hazards while its eyes are temporarily closed. It is therefore likely that the length of time that the eyes are closed and the length of the gap between blinks for a species may reflect aspects of the ecology of that species, such as its social or physical environment. An earlier published study conducted a comparative study linking blinking behaviour and ecology, and detailed a dataset describing the blinking behaviour of a large number of primate species that was collected from captive animals, but the analysis presented did not control for the nonindependence of the data due to common evolutionary history. In the present study, the dataset is reanalysed using phylogenetic comparative methods, after reconsideration of the parameters describing the physical and social environments of the species. I find that blink rate is best described by the locomotion mode of a species, where species moving through arboreal environments blink least, ground-living species blink most, and species that use both environments show intermediate rates. The duration of a blink was also related to locomotion mode, and positively correlated with both mean species group size and mean species body mass, although the increase in relation to group size is small. How a species moves through the environment therefore appears to be important for determining blinking behaviour, and suggests that complex arboreal environments may require less interruption to visual attention. Given that the data were collected with captive individuals, caution is recommended for interpreting the correlations found. creator: Sean A. Rands uri: https://doi.org/10.7717/peerj.10950 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Rands title: Comparative transcriptome analysis of differentially expressed genes related to the physiological changes of yellow-green leaf mutant of maize link: https://peerj.com/articles/10567 last-modified: 2021-02-16 description: Chlorophylls, green pigments in chloroplasts, are essential for photosynthesis. Reduction in chlorophyll content may result in retarded growth, dwarfism, and sterility. In this study, a yellow-green leaf mutant of maize, indicative of abnormity in chlorophyll content, was identified. The physiological parameters of this mutant were measured. Next, global gene expression of this mutant was determined using transcriptome analysis and compared to that of wild-type maize plants. The yellow-green leaf mutant of maize was found to contain lower contents of chlorophyll a, chlorophyll b and carotenoid compounds. It contained fewer active PSII centers and displayed lower values of original chlorophyll fluorescence parameters than the wild-type plants. The real-time fluorescence yield, the electron transport rate, and the net photosynthetic rate of the mutant plants showed reduction as well. In contrast, the maximum photochemical quantum yield of PSII of the mutant plants was similar to that of the wild-type plants. Comparative transcriptome analysis of the mutant plants and wild-type plants led to the identification of differentially expressed 1,122 genes, of which 536 genes were up-regulated and 586 genes down-regulated in the mutant. Five genes in the chlorophyll metabolism pathway, nine genes in the tricarboxylic acid cycle and seven genes related to the conversion of sucrose to starch displayed down-regulated expression. In contrast, genes encoding a photosystem II reaction center PsbP family protein and the PGR5-like protein 1A (PGRL1A) exhibited increased transcript abundance. creator: Tingchun Li creator: Huaying Yang creator: Yan Lu creator: Qing Dong creator: Guihu Liu creator: Feng Chen creator: Yingbing Zhou uri: https://doi.org/10.7717/peerj.10567 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Li et al. title: A comparison of fourteen fully characterized mammalian-associated Campylobacter fetus isolates suggests that loss of defense mechanisms contribute to high genomic plasticity and subspecies evolution link: https://peerj.com/articles/10586 last-modified: 2021-02-16 description: Campylobacter fetus is currently classified into three main subspecies, but only two of these, C. fetus subspecies fetus and C. fetus subsp. venerealis originate principally from ruminants where they inhabit different niches and cause distinct pathogenicity. Their importance as pathogens in international trade and reporting is also different yet the criteria defining these properties have never been fully substantiated nor understood. The situation is further compromised because the ability to differentiate between these two closely related C. fetus subspecies has traditionally been performed by phenotypic characterisation of isolates, methods which are limited in scope, time-consuming, tedious, and often yield inconsistent results, thereby leading to isolate misidentification. The development of robust genetic markers that could enable rapid discrimination between C. fetus subsp. fetus and subsp. venerealis has also been challenging due to limited differences in the gene complement of their genomes, high levels of sequence repetition, the small number of closed genome sequences available and the lack of standardisation of the discriminatory biochemical tests employed for comparative purposes. To yield a better understanding of the genomic differences that define these C. fetus strains, seven isolates were exhaustively characterised phenotypically and genetically and compared with seven previously well characterised isolates. Analysis of these 14 C. fetus samples clearly illustrated that adaption by C. fetus subsp. venerealis to the bovine reproductive tract correlated with increasing genome length and plasticity due to the acquisition and propagation of several mobile elements including prophages, transposons and plasmids harbouring virulence factors. Significant differences in the repertoire of the CRISPR (clustered regularly interspersed short palindromic repeats)-cas system of all C. fetus strains was also found. We therefore suggest that a deficiency in this adaptive immune system may have permitted the emergence of extensive genome plasticity and led to changes in host tropism through gene disruption and/or changes in gene expression. Notable differences in the sub-species complement of DNA adenine methylase genes may also have an impact. These data will facilitate future studies to better understand the precise genetic differences that underlie the phenotypic and virulence differences between these animal pathogens and may identify additional markers useful for diagnosis and sub-typing. creator: Susan A. Nadin-Davis creator: John Chmara creator: Catherine D. Carrillo creator: Kingsley Amoako creator: Noriko Goji creator: Marc-Olivier Duceppe creator: John Devenish uri: https://doi.org/10.7717/peerj.10586 license: http://www.nationalarchives.gov.uk/doc/open-government-licence/ rights: title: An integrated analysis of lncRNA and mRNA expression profiles in the kidneys of mice with lupus nephritis link: https://peerj.com/articles/10668 last-modified: 2021-02-16 description: Long noncoding RNAs (lncRNAs) are persistently expressed and have been described as potential biomarkers and therapeutic targets in various diseases. However, there is limited information regarding lncRNA expression in the tissue of kidney exhibiting lupus nephritis (LN)a serious complication of systemic lupus erythematosus (SLE). In this study, RNA sequencing (RNA-seq) was performed to characterize the lncRNA and mRNA expression in kidney tissues from LN (MRL/lpr) and control mice. We identified 12,979 novel lncRNAs in mouse. The expression profiles of both mRNAs and lncRNAs were differed significantly between LN and control mice. In particular, there were more upregulated lncRNAs and mRNAs than downregulated ones in the kidney tissues of LN mice. However, GO analysis showed that more downregulated genes were enriched in immune and inflammatory response-associated pathways. KEGG analysis showed that both downregulated and upregulated genes were enriched in a number of pathways, including the SLE pathway, and approximately half of these SLE-associated genes encoded inflammatory factors. Moreover, we observed that 2,181 DElncRNAs may have targeted and regulated the expression of 778 mRNAs in LN kidney tissues. The results of this study showed that 11 DElncRNAs targeted and were co-expressed with six immune and SLE-associated genes. qPCR analysis confirmed that lncRNA Gm20513 positively regulated the expression of the SLE-associated gene H2-Aa. In conclusion, the results of our study demonstrates that lncRNAs influence the progression of LN and provide some cues for further study of lncRNAs in LN. These results regarding the lncRNA-mRNAregulatory network may have important value in LN diagnosis and therapy. creator: Juan Wang creator: Xiongfei Wu creator: Yafang Tu creator: Jianzhong Dang creator: Zhitao Cai creator: Wenjing Liao creator: Weili Quan creator: Yaxun Wei uri: https://doi.org/10.7717/peerj.10668 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: Homologous recombination repair rathway and RAD54L in early-stage lung adenocarcinoma link: https://peerj.com/articles/10680 last-modified: 2021-02-16 description: ObjectiveThe current study aims to identify the dysregulated pathway involved in carcinogenesis and the essential survival-related dysregulated genes among this pathway in the early stage of lung adenocarcinoma (LUAD).Patients and MethodsData from The Cancer Genome Atlas (TCGA) including 526 tumor tissues of LUAD and 59 healthy lung tissues were analyzed to gain differentially expressed genes (DEGs). Gene ontology (GO) analysis was conducted with DAVID, while the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of DEGs was performed, followed by gene set enrichment analysis (GSEA) methods. Survival analysis was implemented in TCGA dataset and validated in Gene Expression Omnibus (GEO) cohort GSE50081, which includes 127 patients with stage I LUAD.ResultsGSEA enrichment analysis suggested that homologous recombination repair (HRR) pathway was significantly enriched. Subsequent KEGG pathway enrichment analysis indicated the significant up-regulation of HRR pathway in patients with T1 stage LUAD. Retrieved in Gene database, RAD54L is involved in HRR pathway and were recognized to be significantly differentially expressed in T1 stage LUAD in our study. The survival analysis indicated that high expression of RAD54L was significantly related to worse overall survival in patients with T1 stage LUAD (TCGA cohort: HR=2.10, 95% CI [1.47–2.98], P = 0.001; GSE50081 validation cohort: HR = 2.61, 95% CI [1.51–4.52], P = 0.002). Multivariate cox regression analysis indicated that RAD54L is an independent prognostic factor in the early-stage LUAD.ConclusionHRR pathway is up-regulated in LUAD, among which the expression of RAD54L was found to be significantly differentially expressed in T1 stage tumor tissue. Patients with high expression of RAD54L were associated with worse overall survival in the TCGA cohort and validation cohort. This study suggests a potential mechanism of lung cancer progression and provide a budding prognostic factor and treatment target in early-stage LUAD. creator: Shaopeng Zheng creator: Lintong Yao creator: Fasheng Li creator: Luyu Huang creator: Yunfang Yu creator: Zenan Lin creator: Hao Li creator: Jin Xia creator: Michael Lanuti creator: Haiyu Zhou uri: https://doi.org/10.7717/peerj.10680 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Zheng et al. title: Historical and current distribution ranges and loss of mega-herbivores and carnivores of Asia link: https://peerj.com/articles/10738 last-modified: 2021-02-16 description: Ecosystem functioning is dependent a lot on large mammals, which are, however, vulnerable and facing extinction risks due to human impacts mainly. Megafauna of Asia has been declining for a long, not only in numbers but also in their distribution ranges. In the current study, we collected information on past and current occurrence and distribution records of Asia’s megafauna species. We reconstructed the historical distribution ranges of the six herbivores and four carnivores for comparison with their present ranges, to quantify spatially explicit levels of mega-defaunation. Results revealed that historically the selected megafauna species were more widely distributed than at current. Severe range contraction was observed for the Asiatic lion, three rhino species, Asian elephant, tigers, and tapirs. Defaunation maps generated have revealed the vanishing of megafauna from parts of the East, Southeast, and Southwest Asia, even some protected Areas losing up to eight out of ten megafaunal species. These defaunation maps can help develop future conservation policies, to save the remaining distribution ranges of large mammals. creator: Tariq Mahmood creator: Tuong Thuy Vu creator: Ahimsa Campos-Arceiz creator: Faraz Akrim creator: Shaista Andleeb creator: Muhammad Farooq creator: Abdul Hamid creator: Nadeem Munawar creator: Muhammad Waseem creator: Abid Hussain creator: Hira Fatima creator: Muhammad Raza Khan creator: Sajid Mahmood uri: https://doi.org/10.7717/peerj.10738 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Mahmood et al. title: Determining novel candidate anti-hepatocellular carcinoma drugs using interaction networks and molecular docking between drug targets and natural compounds of SiNiSan link: https://peerj.com/articles/10745 last-modified: 2021-02-16 description: BackgroundSiNiSan (SNS) is an ancient traditional Chinese medicine (TCM) used to treat liver and spleen deficiencies. We studied the unique advantages of using SNS to treat hepatocellular carcinoma (HCC) with multiple components and targets to determine its potential mechanism of action.MethodsThe active compounds from the individual herbs in the SNS formula and their targets were mined from Traditional Chinese Medicine Systems Pharmacology Database (TCMSP). HCC-associated targets were collected from the TCGA and GEO databases and samples were collected from patients with stage III hepatocellular carcinoma. A compound-disease target network was constructed, visualized, and analyzed using Cytoscape software. We built a protein-protein interaction (PPI) network using the String database. We enriched and analyzed key targets using GSEA, GO, and KEGG in order to explore their functions. Autodock software was used to simulate the process of SNS molecules acting on HCC targets.ResultsA total of 113 candidate compounds were taken from SNS, and 64 of the same targets were chosen from HCC and SNS. The predominant targets genes were PTGS2, ESR1, CHEK1, CCNA2, NOS2 and AR; kaempferol and quercetin from SNS were the principal ingredients in HCC treatment. The compounds may work against HCC due to a cellular response to steroid hormones and histone phosphorylation. The P53 signaling pathway was significantly enriched in the gene set GSEA enrichment analysis and differential gene KEGG enrichment analysis.ConclusionsOur results showed that the SNS component has a large number of stage III HCC targets. Among the targets, the sex hormone receptors, the AR and ESR1 genes, are the core targets of SNS component and the most active proteins in the PPI network. In addition, quercetin, which has the most targets, can act on the main targets (BAX, CDK1, CCNB1, SERPINE1, CHEK2, and IGFBP3) of the P53 pathway to treat HCC. creator: Qin Zhang creator: Zhangying Feng creator: Mengxi Gao creator: Liru Guo uri: https://doi.org/10.7717/peerj.10745 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Zhang et al. title: Thick bark can protect trees from a severe ambrosia beetle attack link: https://peerj.com/articles/10755 last-modified: 2021-02-16 description: Thick bark has been shown to protect trees from wildfires, but can it protect trees from an ambrosia beetle attack? We addressed this question by examining the distribution of holes of the invasive Kuroshio Shot Hole Borer (KSHB, Euwallacea kuroshio; Coleoptera: Scolytinae) in the bark of Goodding’s black willow (Salix gooddingii), one of the KSHB’s most-preferred hosts. The study was conducted in the Tijuana River Valley, California, in 2016–17, during the peak of the KSHB infestation there. Using detailed measurements of bark samples cut from 27 infested trees, we tested and found support for two related hypotheses: (1) bark thickness influences KSHB attack densities and attack locations, i.e., the KSHB bores abundantly through thin bark and avoids boring through thick bark; and (2) bark thickness influences KSHB impacts, i.e., the KSHB causes more damage to thinner-barked trees than to thicker-barked trees. Our results indicate that thick bark protects trees because it limits the density of KSHB entry points and thereby limits internal structural damage to low, survivable levels. This is the first study to identify bark thickness as a factor that influences the density of KSHB—or any ambrosia beetle—in its host tree, and the first to link bark thickness to rates of host tree mortality. creator: John M. Boland creator: Deborah L. Woodward uri: https://doi.org/10.7717/peerj.10755 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Boland and Woodward title: Lower Locus Coeruleus MRI intensity in patients with late-life major depression link: https://peerj.com/articles/10828 last-modified: 2021-02-16 description: BackgroundThe locus coeruleus (LC) is the major noradrenergic source in the central nervous system. Structural alterations in the LC contribute to the pathophysiology of different neuropsychiatric disorders, which may increase to a variable extent the likelihood of developing neurodegenerative conditions. The characterization of such alterations may therefore help to predict progression to neurodegenerative disorders. Despite the LC cannot be visualized with conventional magnetic resonance imaging (MRI), specific MRI sequences have been developed to infer its structural integrity.MethodsWe quantified LC signal Contrast Ratios (LCCRs) in late-life major depressive disorder (MDD) (n = 37, 9 with comorbid aMCI), amnestic Mild Cognitive Impairment (aMCI) (n = 21, without comorbid MDD), and healthy controls (HCs) (n = 31), and also assessed the putative modulatory effects of comorbidities and other clinical variables.ResultsLCCRs were lower in MDD compared to aMCI and HCs. While no effects of aMCI comorbidity were observed, lower LCCRs were specifically observed in patients taking serotonin/norepinephrine reuptake inhibitors (SNRIs).ConclusionOur results do not support the hypothesis that lower LCCRs characterize the different clinical groups that may eventually develop a neurodegenerative disorder. Conversely, our results were specifically observed in patients with late-life MDD taking SNRIs. Further research with larger samples is warranted to ascertain whether medication or particular clinical features of patients taking SNRIs are associated with changes in LC neurons. creator: Andrés Guinea-Izquierdo creator: Mónica Giménez creator: Ignacio Martínez-Zalacaín creator: Inés del Cerro creator: Pol Canal-Noguer creator: Gerard Blasco creator: Jordi Gascón creator: Ramon Reñé creator: Inmaculada Rico creator: Angels Camins creator: Carlos Aguilera creator: Mikel Urretavizcaya creator: Isidre Ferrer creator: José Manuel Menchón creator: Virginia Soria creator: Carles Soriano-Mas uri: https://doi.org/10.7717/peerj.10828 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Guinea-Izquierdo et al. title: Influence of periodontal treatment on blood microbiotas: a clinical trial link: https://peerj.com/articles/10846 last-modified: 2021-02-16 description: ObjectiveTo investigate the effects of periodontal treatment on the abundance and diversity of blood microbiota.Methods and MaterialsTwenty-seven periodontitis patients were randomly allocated to a control group (A) and two test groups (B1 and B2). Group A patients received full-mouth scaling and root planing (SRP), group B1 patients received subgingival glycine air polishing (GAP) right after SRP, and group B2 patients received subgingival glycine air polishing right before SRP. Peripheral blood samples were obtained at the baseline, the day after periodontal treatment, and 6 weeks after treatment and evaluated using nested polymerase chain reaction and 16SrRNA Gene Sequencing (Miseq platform).ResultsAll participants exhibited significant improvements in the clinical parameters evaluated at the 6-week follow-up visit compared to the values at the baseline, but no significant differences were observed between the three groups. The total bacterial count was lowest in group B2. The bacterial species diversity (α-diversity) in group B1 was significantly higher (Chao-1 index, P = 0.03) and Porphyromonas and Pantoea were the dominant genera (linear discriminant analysis (LDA > 2)) in this group the day after treatment compared to the baseline. No significant difference was detected in the relative abundance and α-diversity of blood microbiota between the baseline and 6 weeks after treatment.ConclusionLocal periodontal treatment merely disrupts the stability of blood microbiota in the short term. Periodontitis treatment using full-mouth SRP followed by adjunctive GAP is a promising approach to reduce the introduction of bacteria into the bloodstream during the procedure. creator: Wenyi Zhang creator: Yang Meng creator: Jin Jing creator: Yingtao Wu creator: Shu Li uri: https://doi.org/10.7717/peerj.10846 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Zhang et al. title: Tissue inhibitor of metalloproteinase-1 (TIMP-1) as a prognostic biomarker in gastrointestinal cancer: a meta-analysis link: https://peerj.com/articles/10859 last-modified: 2021-02-16 description: BackgroundTissue inhibitor of metalloproteinase 1 (TIMP-1) has recently been shown to be dependent on or independent of Matrix metalloproteinases (MMPs) in its roles in tumorigenesis and progression. This appreciation has prompted various studies assessing the prognostic value of TIMP-1 in patients with gastrointestinal cancer, however, the conclusions were still inconsistent. The aim of this study was to assess the prognostic value of TIMP-1-immunohistochemistry (IHC) staining and pretreatment serum/plasma TIMP-1 level in gastrointestinal cancer survival as well as the association between TIMP-1 and clinicopathologic features.MethodsThe meta-analysis was registered in the International Prospective Register of Systematic Reviews (PROSPERO; Registration NO. CRD42020185407) and followed the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) statement. A highly sensitive literature search was performed in electronic databases including PubMed, EMBASE and the Cochrane Library. Heterogeneity analysis was conducted using both chi-square-based Q statistics and the I2 test. The pooled hazard ratios (HRs) with 95% confidence intervals (CIs) were calculated to assess the prognostic value of TIMP-1 using the fixed-effects model. Odds ratios (ORs) with 95% CIs were calculated to evaluate the associations between TIMP-1 and clinicopathological characteristics. The meta-analysis was conducted using STATA 12.0 software.ResultsA total of 3,958 patients from twenty-two studies were included in the meta-analysis. Elevated TIMP-1 levels were significantly associated with poor survival in gastrointestinal cancer (TIMP-1-IHC staining: HR = 2.04, 95% CI [1.59–2.61], I2 = 35.7%, PQ = 0.156; pretreatment serum/plasma TIMP-1 levels: HR = 2.02, 95% CI [1.80–2.28], I2 = 0%, PQ = 0.630). Moreover, clinicopathological parameter data analysis showed that elevated TIMP-1 levels were significantly associated with lymph node metastasis (N1/N2/N3 vs N0: OR = 2.92, 95% CI [1.95–4.38]) and higher TNM stages (III/IV vs I/II: OR = 2.73, 95% CI [1.23–6.04]).ConclusionBoth TIMP-1-positive IHC staining and high serum/plasma TIMP-1 levels are poor prognostic factors for the survival of gastrointestinal cancer. In addition, TIMP-1 overexpression was correlated with more advanced clinicopathological features. creator: Lili Qin creator: Yueqi Wang creator: Na Yang creator: Yangyu Zhang creator: Tianye Zhao creator: Yanhua Wu creator: Jing Jiang uri: https://doi.org/10.7717/peerj.10859 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Qin et al. title: Sugarcane/peanut intercropping system improves physicochemical properties by changing N and P cycling and organic matter turnover in root zone soil link: https://peerj.com/articles/10880 last-modified: 2021-02-16 description: BackgroundThe sugarcane/peanut intercropping system is a specific and efficient cropping pattern in South China. Intercropping systems change the bacterial diversity of soils and decrease disease rates. It can not only utilized light, heat, water and land resources efficiently, but also increased yield and economic benefits of farmers.MethodsWe determined soil nutrients, enzymes and microbes in sugarcane/peanut intercropping system, and analyzed relevance of the soil physicochemical properties and the genes involved in N and P cycling and organic matter turnover by metagenome sequencing.ResultsThe results showed that sugarcane/peanut intercropping significantly boosted the content of total nitrogen, available phosphorus, total potassium, organic matter, pH value and bacteria and enhanced the activity of acid phosphatase compared to monocropping. Especially the content of available nitrogen, available phosphorus and organic matter increased significantly by 20.1%, 65.3% and 56.0% in root zone soil of IP2 treatment than monocropping treatment. The content of available potassium and microbial biomass carbon, as well as the activity of catalase, sucrase and protease, significantly decreased in intercropping root zone soil. Intercropping resulted in a significant increase by 7.8%, 16.2% and 23.0% in IS, IP1 and IP2, respectively, of the acid phosphatase content relative to MS. Metagenomic analysis showed that the pathways involved in carbohydrate and amino acid metabolism were dominant and more abundant in intercropping than in monocropping. Moreover, the relative abundances of genes related to N cycling (glnA, GLUD1_2, nirK), P cycling (phoR, phoB) and organic matter turnover (PRDX2_4) were higher in the intercropping soil than in the monocropping soil. The relative abundance of GLUD1_2 and phoR were 25.5% and 13.8% higher in the IP2 treatment respectively,and bgIX was higher in IS treatment compared to the monocropping treatment. Genes that were significantly related to phosphorus metabolism and nitrogen metabolism (TREH, katE, gudB) were more abundant in intercropping than in monocropping.ConclusionThe results of this study indicate that the intercropping system changed the numbers of microbes as well as enzymes activities, and subsequently regulate genes involved in N cycling, P cycling and organic matter turnover. Finally, it leads to the increase of nutrients in root zone soil and improved the soil environment. creator: Xiumei Tang creator: Yixin Zhang creator: Jing Jiang creator: Xiuzhen Meng creator: Zhipeng Huang creator: Haining Wu creator: Liangqiong He creator: Faqian Xiong creator: Jing Liu creator: Ruichun Zhong creator: Zhuqiang Han creator: Ronghua Tang uri: https://doi.org/10.7717/peerj.10880 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Tang et al. title: Predicting lung adenocarcinoma disease progression using methylation-correlated blocks and ensemble machine learning classifiers link: https://peerj.com/articles/10884 last-modified: 2021-02-16 description: Applying the knowledge that methyltransferases and demethylases can modify adjacent cytosine-phosphorothioate-guanine (CpG) sites in the same DNA strand, we found that combining multiple CpGs into a single block may improve cancer diagnosis. However, survival prediction remains a challenge. In this study, we developed a pipeline named “stacked ensemble of machine learning models for methylation-correlated blocks” (EnMCB) that combined Cox regression, support vector regression (SVR), and elastic-net models to construct signatures based on DNA methylation-correlated blocks for lung adenocarcinoma (LUAD) survival prediction. We used methylation profiles from the Cancer Genome Atlas (TCGA) as the training set, and profiles from the Gene Expression Omnibus (GEO) as validation and testing sets. First, we partitioned the genome into blocks of tightly co-methylated CpG sites, which we termed methylation-correlated blocks (MCBs). After partitioning and feature selection, we observed different diagnostic capacities for predicting patient survival across the models. We combined the multiple models into a single stacking ensemble model. The stacking ensemble model based on the top-ranked block had the area under the receiver operating characteristic curve of 0.622 in the TCGA training set, 0.773 in the validation set, and 0.698 in the testing set. When stratified by clinicopathological risk factors, the risk score predicted by the top-ranked MCB was an independent prognostic factor. Our results showed that our pipeline was a reliable tool that may facilitate MCB selection and survival prediction. creator: Xin Yu creator: Qian Yang creator: Dong Wang creator: Zhaoyang Li creator: Nianhang Chen creator: De-Xin Kong uri: https://doi.org/10.7717/peerj.10884 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Yu et al. title: Quantile-dependent expressivity of serum C-reactive protein concentrations in family sets link: https://peerj.com/articles/10914 last-modified: 2021-02-16 description: Background“Quantile-dependent expressivity” occurs when the effect size of a genetic variant depends upon whether the phenotype (e.g., C-reactive protein, CRP) is high or low relative to its distribution. We have previously shown that the heritabilities (h2) of coffee and alcohol consumption, postprandial lipemia, lipoproteins, leptin, adiponectin, adiposity, and pulmonary function are quantile-specific. Whether CRP heritability is quantile-specific is currently unknown.MethodsSerum CRP concentrations from 2,036 sibships and 6,144 offspring-parent pairs were analyzed from the Framingham Heart Study. Quantile-specific heritability from full-sib (βFS, h2 ={(1 + 8rspouseβFS)0.5 − 1}/(2rspouse)) and offspring-parent regression slopes (βOP, h2 = 2βOP/(1 + rspouse)) were estimated robustly by quantile regression with nonparametric significance determined from 1,000 bootstrap samples.ResultsQuantile-specific h2 (±SE) increased with increasing percentiles of the offspring’s age- and sex-adjusted CRP distribution when estimated from βOP (Ptrend = 0.0004): 0.02 ± 0.01 at the 10th, 0.04 ± 0.01 at the 25th, 0.10 ± 0.02 at the 50th, 0.20 ± 0.05 at the 75th, and 0.33 ± 0.10 at the 90th percentile, and when estimated from βFS (Ptrend = 0.0008): 0.03±0.01 at the 10th, 0.06 ± 0.02 at the 25th, 0.14 ± 0.03 at the 50th, 0.24 ± 0.05 at the 75th, and 0.53 ± 0.21 at the 90th percentile.ConclusionHeritability of serum CRP concentration is quantile-specific, which may explain or contribute to the inflated CRP differences between CRP (rs1130864, rs1205, rs1800947, rs2794521, rs3091244), FGB (rs1800787), IL-6 (rs1800795, rs1800796), IL6R (rs8192284), TNF-α (rs1800629) and APOE genotypes following CABG surgery, stroke, TIA, curative esophagectomy, intensive periodontal therapy, or acute exercise; during acute coronary syndrome or Staphylococcus aureus bacteremia; or in patients with chronic rheumatoid arthritis, diabetes, peripheral arterial disease, ankylosing spondylitis, obesity or inflammatory bowel disease or who smoke. creator: Paul T. Williams uri: https://doi.org/10.7717/peerj.10914 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Williams title: Immune function of miR-214 and its application prospects as molecular marker link: https://peerj.com/articles/10924 last-modified: 2021-02-16 description: MicroRNAs are a class of evolutionary conserved non-coding small RNAs that play key regulatory roles at the post-transcriptional level. In recent years, studies have shown that miR-214 plays an important role in regulating several biological processes such as cell proliferation and differentiation, tumorigenesis, inflammation and immunity, and it has become a hotspot in the miRNA field. In this review, the regulatory functions of miR-214 in the proliferation, differentiation and functional activities of immune-related cells, such as dendritic cells, T cells and NK cells, were briefly reviewed. Also, the mechanisms of miR-214 involved in tumor immunity, inflammatory regulation and antivirus were discussed. Finally, the value and application prospects of miR-214 as a molecular marker in inflammation and tumor related diseases were analyzed briefly. We hope it can provide reference for further study on the mechanism and application of miR-214. creator: Qiuyuan Wang creator: Yang Liu creator: Yiru Wu creator: Jie Wen creator: Chaolai Man uri: https://doi.org/10.7717/peerj.10924 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: A kingdom in decline: Holocene range contraction of the lion (Panthera leo) modelled with global environmental stratification link: https://peerj.com/articles/10504 last-modified: 2021-02-15 description: AimWe use ecological niche models and environmental stratification of palaeoclimate to reconstruct the changing range of the lion (Panthera leo) during the late Pleistocene and Holocene.LocationThe modern (early 21st century) range of the lion extends from southern Africa to the western Indian Subcontinent, yet through the 20th century this range has been drastically reduced in extent and become increasingly fragmented as a result of human impacts.MethodsWe use Global Environmental Stratification with MaxEnt ecological niche models to map environmental suitability of the lion under current and palaeoclimatic scenarios. By examining modelled lion range in terms of categorical environmental strata, we characterise suitable bioclimatic conditions for the lion in a descriptive manner.ResultsWe find that lion habitat suitability has reduced throughout the Holocene, controlled by pluvial/interpluvial cycles. The aridification of the Sahara  6ka dramatically reduced lion range throughout North Africa. The association of Saharan aridification with the development of pastoralism and the growth of sedentary communities, who practised animal husbandry, would have placed additional and lasting anthropogenic pressures on the lion.Main ConclusionsThis research highlights the need to integrate the full effects of the fluctuating vegetation and desiccation of the Sahara into palaeoclimatic models, and provides a starting point for further continental-scale analyses of shifting faunal ranges through North Africa and the Near East during the Holocene. This scale of ecological niche modelling does not explain the current pattern of genetic variation in the lion, and we conclude that narrow but substantial physical barriers, such as rivers, have likely played a major role in population vicariance throughout the Late Pleistocene. creator: David M. Cooper creator: Andrew J. Dugmore creator: Andrew C. Kitchener creator: Marc J. Metzger creator: Antonio Trabucco uri: https://doi.org/10.7717/peerj.10504 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Cooper et al. title: Magnetic resonance diffusion tensor imaging applied to rat model of contrast-induced acute kidney injury link: https://peerj.com/articles/10620 last-modified: 2021-02-15 description: ObjectivesIn this preclinical investigation, the feasibility of using diffusion tensor imaging (DTI) to study contrast-induced acute kidney injury (CIAKI) is explored, comparing radiographic outcomes with histopathologic and immunohistochemical findings after repeated animal exposures to iodinated contrast agent.Materials and MethodsForty-five male wistar rats were allocated to three groups (n = 15 each), each receiving two separate injections 1 day apart: group 1 (iodixanol then saline); group 2 (iodixanol twice); and control group (saline twice). Five rats were then randomly selected from each group at three separate time points (1 h, 24 h, and 120 h) for magnetic resonance imaging (MRI). Upon MRI completion, the animals were sacrificed, examining renal tissue and serum creatinine level. DTI data served to calculate fractional anisotropy (FA) and apparent diffusion coefficient (ADC).ResultsFA values were significantly lower in group 2 than in the others. Compared with controls, FA assessments at 1 h, 24 h, and 120 h after injections commenced were significantly lower in group 2; and ADC was significantly more pronounced at 24 h. Serum creatinine levels at 24 h were markedly elevated in both groups 1 and 2. Pearson correlation analysis revealed significant negative correlations between FA (r = −0.730; p < 0.05) or ADC (r = −0.827; p < 0.05) and tubular injury and between FA (r = −0.563; p < 0.05) or ADC (r = −0.805; p < 0.05) and hypoxia-inducible factor-1α.ConclusionsAnalytic approaches to DTI with better reproducibility should aid in monitoring the early pathophysiologic derangements of CIAKI, thus facilitating timely reversal of the detrimental effects. creator: Bin Wang creator: Junjie Li creator: Yongfang Wang uri: https://doi.org/10.7717/peerj.10620 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Wang et al. title: New data on tail lengths and variation along the caudal series in the non-avialan dinosaurs link: https://peerj.com/articles/10721 last-modified: 2021-02-15 description: The tails of non-avialan dinosaurs varied considerably in terms of overall length, total number of vertebrae, and gross form and function. A new dataset confirms that there is little or no consistent relationship between tail length and snout-sacrum length. Consequently, attempts to estimate one from the other are likely to be very error-prone. Patterns of changes in centra lengths across the caudal series vary among non-avian dinosaurs. However, some overarching patterns do emerge. A number of taxa show (anterior to posterior) a series of short centra, followed by a series of longer centra, with the remainder of the tail consisting of a long series of centra tapering in length. This pattern is consistent with functional constraints, and the anterior series of longer centra are coincident with the major attachments of femoral musculature. This pattern is not present in many basal taxa and may have evolved independently in different dinosaurian groups, further suggesting functional importance. creator: David W.E. Hone creator: W. Scott Persons creator: Steven C. Le Comber uri: https://doi.org/10.7717/peerj.10721 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Hone et al. title: Evaluation of full-length nanopore 16S sequencing for detection of pathogens in microbial keratitis link: https://peerj.com/articles/10778 last-modified: 2021-02-15 description: BackgroundMicrobial keratitis is a leading cause of preventable blindness worldwide. Conventional sampling and culture techniques are time-consuming, with over 40% of cases being culture-negative. Nanopore sequencing technology is portable and capable of generating long sequencing reads in real-time. The aim of this study is to evaluate the potential of nanopore sequencing directly from clinical samples for the diagnosis of bacterial microbial keratitis.MethodsUsing full-length 16S rRNA amplicon sequences from a defined mock microbial community, we evaluated and benchmarked our bioinformatics analysis pipeline for taxonomic assignment on three different 16S rRNA databases (NCBI 16S RefSeq, RDP and SILVA) with clustering at 97%, 99% and 100% similarities. Next, we optimised the sample collection using an ex vivo porcine model of microbial keratitis to compare DNA recovery rates of 12 different collection methods: 21-gauge needle, PTFE membrane (4 mm and 6 mm), Isohelix™ SK-2S, Sugi® Eyespear, Cotton, Rayon, Dryswab™, Hydraflock®, Albumin-coated, Purflock®, Purfoam and Polyester swabs. As a proof-of-concept study, we then used the sampling technique that provided the highest DNA recovery, along with the optimised bioinformatics pipeline, to prospectively collected samples from patients with suspected microbial keratitis. The resulting nanopore sequencing results were then compared to standard microbiology culture methods.ResultsWe found that applying alignment filtering to nanopore sequencing reads and aligning to the NCBI 16S RefSeq database at 100% similarity provided the most accurate bacterial taxa assignment. DNA concentration recovery rates differed significantly between the collection methods (p < 0.001), with the Sugi® Eyespear swab providing the highest mean rank of DNA concentration. Then, applying the optimised collection method and bioinformatics pipeline directly to samples from two patients with suspected microbial keratitis, sequencing results from Patient A were in agreement with culture results, whilst Patient B, with negative culture results and previous antibiotic use, showed agreement between nanopore and Illumina Miseq sequencing results.ConclusionWe have optimised collection methods and demonstrated a novel workflow for identification of bacterial microbial keratitis using full-length 16S nanopore sequencing. creator: Liying Low creator: Pablo Fuentes-Utrilla creator: James Hodson creator: John D. O’Neil creator: Amanda E. Rossiter creator: Ghazala Begum creator: Kusy Suleiman creator: Philip I. Murray creator: Graham R. Wallace creator: Nicholas J. Loman creator: Saaeha Rauz uri: https://doi.org/10.7717/peerj.10778 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Low et al. title: Prevalence of Xanthomonas euvesicatoria (formally X. perforans) associated with bacterial spot severity in Capsicum annuum crops in South Central Chihuahua, Mexico link: https://peerj.com/articles/10913 last-modified: 2021-02-15 description: BackgroundXanthomonas spp. causes bacterial spot disease, which reduces quality and yield of pepper crops in Mexico. Identification of phytopathogen species is necessary to implement more effective control strategies.ObjectiveThe aim of this study was to isolate and identify infecting Xanthomonas species in South Central Chihuahua pepper-producing areas.MethodsDiseased plants were collected from 30 cultivation lots and bacteria were isolated from damaged tissues. Potential causative agents were isolated, identified, and characterized by biochemical and molecular analysis. Pathogenicity tests from each isolate were then performed on 30-d-old pepper seedlings, exposing five leaves to 10 µL of 1 × 108 CFU/mL bacterial suspensions of each isolate, using sterile distilled water as a control. Disease severity was determined after 10 d by calculating leaf damage percentage. Furthermore, we evaluated the susceptibility of the highest bacterial spot severity-causing isolates (13 isolates) to copper sulphate (CuS), copper gluconate (CuG), copper oxychloride + oxytetracycline hydrochloride (Cu + Ox), gentamicin + oxytetracycline hydrochloride (Gen + Ox), and gentamicin sulphate (GenS). Copper-resistance genes (copLAB) were detected by PCR analysis among isolates.ResultsThirty-seven foliage isolates were identified as Xanthomonas euvesicatoria (14%), which were associated with bacterial spot disease in jalapeño pepper. Tested Xanthomonas isolates were resistant to Cu-based compounds, but susceptible to Cu + Ox. All isolates were susceptible to Gen + Ox and GenS. CopLAB genes were detected in all but one strain.ConclusionsX. euvesicatoria (formally X. perforans) may be considered as an emerging pathogen of bacterial spot pepper in Mexico. Among disease management strategies, alternatives to copper should be taken into consideration. creator: Jared Hernández-Huerta creator: Patricia Tamez-Guerra creator: Ricardo Gomez-Flores creator: Ma. Carmen E. Delgado-Gardea creator: Margarita S. García-Madrid creator: Loreto Robles-Hernández creator: Rocio Infante-Ramirez uri: https://doi.org/10.7717/peerj.10913 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Hernández-Huerta et al. title: Effect of incubation temperature and substrate moisture on embryonic development, hatchling phenotypes and post-hatching growth in the Reeves’ Turtle, Mauremys reevesii link: https://peerj.com/articles/10553 last-modified: 2021-02-12 description: BackgroundReeves’ Turtles (Mauremys reevesii) are economically important in aquaculture in China. Understanding the effects of incubation temperature and substrate moisture on embryos and hatchlings is of great significance for improving the artificial culture of M. reevesii. However, available studies have not yet determined the thermal and hydric optima for M. reevesii eggs, and the potential interaction between the two factors.MethodsIn this study, eggs of M. reevesii were incubated at five temperature levels (23, 26, 29, 32 and 35 °C, fluctuation range ± 0.5 °C). In each temperature level, there were three substrate moisture levels (1:0.5, 1:0.9 and 1:1.2, weight ratio of vermiculite to water). Thus, a total of 15 combinations of temperature and moisture were used to examine the effects of incubation temperature and substrate moisture on incubation duration, hatching success, hatchling phenotypes, post-hatching growth and hatchling survival.ResultsSubstrate moisture did not significantly affect most development parameters (except incubation duration and carapace width of hatchlings). Eggs incubated at low moisture level (1:0.5) had a longer incubation duration and produced hatchlings with smaller carapace widths than those incubated at medium (1:0.9) or high (1:1.2) moisture levels. Incubation temperature had a significant effect on incubation duration, hatching success, hatchling phenotypes and hatchling survival. Incubation duration decreased as incubation temperature increased. Eggs incubated at 23, 26 and 29 °C showed higher hatching success than those incubated at 32 and 35 °C. Hatchlings incubated at 32 °C were smaller in body size and mass than those incubated at 23, 26 and 29 °C. At 12 months of age, incubation temperature had no long-lasting effect on body mass, but hatchlings incubated at 23 and 35 °C had lower survival rates than those incubated at 26, 29 and 32 °C. For the development of embryos and hatchlings, the interaction between incubation temperature and substrate moisture was not significant.ConclusionsOur results indicate that incubation temperature has a significant influence on the development of embryos and hatchlings of M. reevesii, while substrate moisture only significantly affects the incubation duration and carapace width of hatchlings. The combination of an incubation temperature of 29 ± 0.5 °C and a substrate moisture level of 1:1.2 represented optimal incubation conditions in this experiment. Such incubation conditions are helpful in obtaining higher hatching success, shorter incubation duration and higher survival rates for this aquaculture species. creator: Yufeng Wei creator: Yangchun Gao creator: Dainan Cao creator: Yan Ge creator: Haitao Shi creator: Shiping Gong uri: https://doi.org/10.7717/peerj.10553 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Wei et al. title: COVID-19 incidence and mortality in Nigeria: gender based analysis link: https://peerj.com/articles/10613 last-modified: 2021-02-12 description: BackgroundCoronavirus Disease 2019 (COVID-19) has been surging globally. Risk strata in medical attention are of dynamic significance for apposite assessment and supply distribution. Presently, no known cultured contrivance is available to fill this gap of this pandemic. The aim of this study is to develop a predictive model based on vector autoregressive moving average (VARMA) model of various orders for gender based daily COVID-19 incidence in Nigeria. This study also aims to proffer empirical evidence that compares incidence between male and female for COVID-19 risk factors.MethodsWilcoxon signed-rank test is employed to investigate the significance of the difference in the gender distributions of the daily incidence. A VARMA model of various orders is formulated for the gender based daily COVID-19 incidence in Nigeria. The optimal VARMA model is identified using Bayesian information criterion. Also, a predictive model based on univariate autoregressive moving average model is formulated for the daily death cases in Nigeria. Fold change is estimated based on crude case-fatality risk to investigate whether there is massive underreporting and under-testing of COVID-19 cases in Nigeria.ResultsDaily incidence is higher in males on most days from 11 April 2020 to 12 September 2020. Result of Wilcoxon signed-rank test shows that incidence among male is significantly higher than female (p-value < 2.22 × 10−16). White neural network test shows that daily female incidence is not linear in mean (p-value = 0.00058746) while daily male incidence is linear in mean (p-value = 0.4257). McLeod-Li test shows that there is autoregressive conditional heteroscedasticity in the female incidence (Maximum p-value = 1.4277 × 10−5) and male incidence (Maximum p-value = 9.0816 × 10−14) at 5% level of significance. Ljung-Box test (Tsay, 2014) shows that the daily incidence cases are not random (p-value=0.0000). The optimal VARMA model for male and female daily incidence is VARMA (0,1). The optimal model for the Nigeria’s daily COVID-19 death cases is identified to be ARIMA (0,1,1). There is no evidence of massive underreporting and under-testing of COVID-19 cases in Nigeria.ConclusionsComparison of the observed incidence with fitted data by gender shows that the optimal VARMA and ARIMA models fit the data well. Findings highlight the significant roles of gender on daily COVID-19 incidence in Nigeria. creator: Olubukola O. Olusola-Makinde creator: Olusola S. Makinde uri: https://doi.org/10.7717/peerj.10613 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Olusola-Makinde and Makinde title: Gut bacterial communities in the freshwater snail Planorbella trivolvis and their modification by a non-herbivorous diet link: https://peerj.com/articles/10716 last-modified: 2021-02-12 description: The freshwater pulmonate snail Planorbella trivolvis is a common species in various bodies of water but is not native to China. Planorbella trivolvis usually live on diets with high fiber content, such as water grasses, algae and fallen leaves. These snails can attach to the wall of a water tank or to water grass and can be transported overseas to China through the ornamental fish trade. There are few studies investigating the intestinal microbiota of freshwater snails. In this study, using culture-independent molecular analysis, we assessed for the first time the complexity of bacterial communities in the intestines of reared snails. The intestinal microbiota in the snails fed different diets, that is, herbivorous feed (HV) with high cellulose and non-herbivorous feed (NHV) with low cellulose, were analyzed by Illumina sequencing. The results showed that the NHV-based diet significantly increased the body mass, shell diameter and specific growth rate of the snails after 60 days of rearing (P < 0.05). Histological experiments showed that the fat droplets in the epithelium columnar cells of the intestines of the NHV snails increased, and the cilia on these cells fell off. The sequencing results identified 486 and 195 OTUs in HV and NHV, respectively. Lots of bacteria were not reported previously in snails. The intestinal microbiota diversity index (Shannon, Simpson, Ace and Chao) in the NHV snails was significantly lower than that in the HV snails. The gut microbiota in the HV snails were predominantly Proteobacteria (52.97%) and Bacteroidetes (28.75%), while the gut microbiota in NHV snails were predominantly Proteobacteria (95.23%). At the genus level, Cloacibacterium (24.60%), Pseudomonas (4.47%), OM6ON (6.12%), and Rhodobacter (5.79%) were observed to be abundant in HV snails. However, Aeromonas (85.4%) was determined to be predominant in NHV snails. Functional prediction of the gut microbiome in snails by PICRUSt demonstrated a significant difference between the two groups, and the HV snails exhibited higher lignocellulose enzyme activity than did the NHV snails. This study represents a first step in characterizing the gut microbiota of the freshwater snail. Most of these microbes can process plant biomass and digest cellulose and lignocellulose, and the enzymes of these bacteria may have potential biotechnological applications in a variety of industrial processes. creator: Zongfu Hu creator: Qing Tong creator: Jie Chang creator: Jianhua Yu creator: Shuguo Li creator: Huaxin Niu creator: Deying Ma uri: https://doi.org/10.7717/peerj.10716 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Hu et al. title: Effects of dry-wet cycles on nitrous oxide emissions in freshwater sediments: a synthesis link: https://peerj.com/articles/10767 last-modified: 2021-02-12 description: BackgroundSediments frequently exposed to dry-wet cycles are potential biogeochemical hotspots for greenhouse gas (GHG) emissions during dry, wet and transitional phases. While the effects of drying and rewetting on carbon fluxes have been studied extensively in terrestrial and aquatic systems, less is known about the effects of dry-wet cycles on N2O emissions from aquatic systems. As a notable part of lotic systems are temporary, and small lentic systems can substantially contribute to GHG emissions, dry-wet cycles in these ecosystems can play a major role on N2O emissions.MethodologyThis study compiles literature focusing on the effects of drying, rewetting, flooding, and water level fluctuations on N2O emissions and related biogeochemical processes in sediments of lentic and lotic ecosystems.ResultsN2O pulses were observed following sediment drying and rewetting events. Moreover, exposed sediments during dry phases can be active spots for N2O emissions. The general mechanisms behind N2O emissions during dry-wet cycles are comparable to those of soils and are mainly related to physical mechanisms and enhanced microbial processing in lotic and lentic systems. Physical processes driving N2O emissions are mainly regulated by water fluctuations in the sediment. The period of enhanced microbial activity is driven by increased nutrient availability. Higher processing rates and N2O fluxes have been mainly observed when nitrification and denitrification are coupled, under conditions largely determined by O2 availability.ConclusionsThe studies evidence the driving role of dry-wet cycles leading to temporarily high N2O emissions in sediments from a wide array of aquatic habitats. Peak fluxes appear to be of short duration, however, their relevance for global emission estimates as well as N2O emissions from dry inland waters has not been quantified. Future research should address the temporal development during drying-rewetting phases in more detail, capturing rapid flux changes at early stages, and further explore the functional impacts of the frequency and intensity of dry-wet cycles. creator: Renata Pinto creator: Gabriele Weigelhofer creator: António Guerreiro Brito creator: Thomas Hein uri: https://doi.org/10.7717/peerj.10767 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Pinto et al. title: RNA sequencing-based exploration of the effects of far-red light on lncRNAs involved in the shade-avoidance response of D. officinale link: https://peerj.com/articles/10769 last-modified: 2021-02-12 description: Dendrobium officinale (D. officinale) is a valuable medicinal plant with a low natural survival rate, and its shade-avoidance response to far-red light is as an important strategy used by the plant to improve its production efficiency. However, the lncRNAs that play roles in the shade-avoidance response of D. officinale have not yet been investigated. This study found that an appropriate proportion of far-red light can have several effects, including increasing the leaf area and accelerating stem elongation, in D. officinale. The effects of different far-red light treatments on D. officinale were analysed by RNA sequencing technology, and a total of 69 and 78 lncRNAs were differentially expressed in experimental group 1 (FR1) versus the control group (CK) (FR1-CK) and in experimental group 4 (FR4) versus the CK (FR4-CK), respectively. According to GO and KEGG analyses, most of the differentially expressed lncRNA targets are involved in the membrane, some metabolic pathways, hormone signal transduction, and O-methyltransferase activity, among other functions. Physiological and biochemical analyses showed that far-red light promoted the accumulation of flavonoids, alkaloids, carotenoids and polysaccharides in D. officinale. The effect of far-red light on D. officinalemight be closely related to the cell membrane and Ca2+ transduction. Based on a Cytoscape analysis and previous research, this study also found that MSTRG.38867.1, MSTRG.69319.1, and MSTRG.66273.1, among other components, might participate in the far-red light signalling network through their targets and thus regulate the shade-avoidance response of D. officinale. These findings will provide new insights into the shade-avoidance response of D. officinale. creator: Hansheng Li creator: Wei Ye creator: Yaqian Wang creator: Xiaohui Chen creator: Yan Fang creator: Gang Sun uri: https://doi.org/10.7717/peerj.10769 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Li et al. title: The effect of ethanol concentration on the morphological and molecular preservation of insects for biodiversity studies link: https://peerj.com/articles/10799 last-modified: 2021-02-12 description: Traditionally, insects collected for scientific purposes have been dried and pinned, or preserved in 70% ethanol. Both methods preserve taxonomically informative exoskeletal structures well but are suboptimal for preserving DNA for molecular biology. Highly concentrated ethanol (95–100%), preferred as a DNA preservative, has generally been assumed to make specimens brittle and prone to breaking. However, systematic studies on the correlation between ethanol concentration and specimen preservation are lacking. Here, we tested how preservative ethanol concentration in combination with different sample handling regimes affect the integrity of seven insect species representing four orders, and differing substantially in the level of sclerotization. After preservation and treatments (various levels of disturbance), we counted the number of appendages (legs, wings, antennae, or heads) that each specimen had lost. Additionally, we assessed the preservation of DNA after long-term storage by comparing the ratio of PCR amplicon copy numbers to an added artificial standard. We found that high ethanol concentrations indeed induce brittleness in insects. However, the magnitude and nature of the effect varied strikingly among species. In general, ethanol concentrations at or above 90% made the insects more brittle, but for species with robust, thicker exoskeletons, this did not translate to an increased loss of appendages. Neither freezing the samples nor drying the insects after immersion in ethanol had a negative effect on the retention of appendages. However, the morphology of the insects was severely damaged if they were allowed to dry. We also found that DNA preserves less well at lower ethanol concentrations when stored at room temperature for an extended period. However, the magnitude of the effect varies among species; the concentrations at which the number of COI amplicon copies relative to the standard was significantly decreased compared to 95% ethanol ranged from 90% to as low as 50%. While higher ethanol concentrations positively affect long-term DNA preservation, there is a clear trade-off between preserving insects for morphological examination and genetic analysis. The optimal ethanol concentration for the latter is detrimental for the former, and vice versa. These trade-offs need to be considered in large insect biodiversity surveys and other projects aiming to combine molecular work with traditional morphology-based characterization of collected specimens. creator: Daniel Marquina creator: Mateusz Buczek creator: Fredrik Ronquist creator: Piotr Łukasik uri: https://doi.org/10.7717/peerj.10799 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Marquina et al. title: Prognostic implications of cell division cycle protein 45 expression in hepatocellular carcinoma link: https://peerj.com/articles/10824 last-modified: 2021-02-12 description: BackgroundThe overall prognosis of hepatocellular carcinoma (HCC) is poor and novel prognostic biomarkers might better monitor the progression of HCC. Cell division cycle protein 45 (CDC45) plays a key role in DNA replication and considered to be involved in tumorigenesis. This study investigated CDC45 expression in tumour tissues and defined its prognostic value in HCC patients.MethodsWe used immunohistochemistry (IHC) staining to examine the expression of CDC45 in tumour tissue specimens and compare them with adjacent normal tissue specimens using a constructed tissue microarray (TMA) and analyzed how clinical features are related to HCC prognosis. Functional enrichment analyses were used to describe significantly involved hallmark pathways of differentially expressed genes (DEGs, which were screened out according to the high or low expression of CDC45 in tumour tissues).ResultsOur findings showed that the proteome expression of CDC45 was evidently downregulated in HCC tissues compared with matched normal tissues (P < 0.0001). Although we did not find any differences in terms of vascular invasion, metastasis, lymphatic infiltration, or Edmondson grade between patients with high and low CDC45 expression, low CDC45 expression was significantly correlated with microvascular invasion (P = 0.046). Multivariate analysis indicated that CDC45 expression (P = 0.035) was an independent prognostic factor for the overall survival (OS) rate of HCC patients. Patients with CDC45 expression was positively correlated with OS rates among HCC patients (P < 0.05). Functional annotations indicated that CDC45 is involved in the most significant pathways, including the cell cycle, DNA replication, chemical carcinogenesis and drug metabolism–cytochrome P450 pathways.DiscussionOur findings showed that low proteomic level of CDC45 was associated with a poor prognosis in HCC patients, indicating that CDC45 might be a novel prognostic marker. creator: Chen Yang creator: shufang Xie creator: Yi Wu creator: Guoqing Ru creator: Xianglei He creator: Hong-yin Pan creator: Shibing Wang creator: Xiangmin Tong uri: https://doi.org/10.7717/peerj.10824 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Yang et al. title: Effects of pharmaceutically active compounds (PhACs) on fish body and scale shape in natural waters link: https://peerj.com/articles/10642 last-modified: 2021-02-11 description: BackgroundIn recent years, there are growing concerns about pharmaceutically active compounds (PhACs) in natural ecosystems. These compounds have been found in natural waters and in fish tissues worldwide. Regarding their growing distribution and abundance, it is becoming clear that traditionally used risk assessment methodologies and ecotoxicological studies have limitations in several respects. In our study a new, combined approach of environmental impact assesment of PhACs has been used.MethodsIn this study, the constant watercourses of the suburban region of the Hungarian capital (Budapest) were sampled, and the body shape and scale shape of three fish species (roach Rutilus rutilus, chub Squalius cephalus, gibel carp Carassius gibelio) found in these waters were analyzed, based on landmark-based geometric morphometric methods. Possible connections were made between the differences in body shape and scale shape, and abiotic environmental variables (local- and landscape-scale) and measured PhACs.ResultsSignificant connections were found between shape and PhACs concentrations in several cases. Despite the relatively large number of compounds (54) detected, citalopram, propranolol, codeine and trimetazidine significantly affected only fish body and scale shape, based on their concentrations. These four PhACs were shown to be high (citalopram), medium (propranolol and codeine), and low (trimetazidine) risk levels during the environmental risk assessment, which were based on Risk Quotient calculation. Furthermore, seven PhACs (diclofenac, Estrone (E1), tramadol, caffeine 17α-Ethinylestradiol (EE2), 17α-Estradiol (aE2), Estriol (E3)) were also categorized with a high risk level. However, our morphological studies indicated that only citalopram was found to affect fish phenotype amongst the PhACs posing high risk. Therefore, our results revealed that the output of (traditional) environmental/ecological risk assessment based on ecotoxicological data of different aquatic organisms not necessarily show consistency with a “real-life” situation; furthermore, the morphological investigations may also be a good sub-lethal endpoint in ecotoxicological assessments. creator: Adam Staszny creator: Peter Dobosy creator: Gabor Maasz creator: Zoltan Szalai creator: Gergely Jakab creator: Zsolt Pirger creator: Jozsef Szeberenyi creator: Eva Molnar creator: Lilianna Olimpia Pap creator: Vera Juhasz creator: Andras Weiperth creator: Bela Urbanyi creator: Attila Csaba Kondor creator: Arpad Ferincz uri: https://doi.org/10.7717/peerj.10642 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Staszny et al. title: Freshwater sponge hosts and their green algae symbionts: a tractable model to understand intracellular symbiosis link: https://peerj.com/articles/10654 last-modified: 2021-02-11 description: In many freshwater habitats, green algae form intracellular symbioses with a variety of heterotrophic host taxa including several species of freshwater sponge. These sponges perform important ecological roles in their habitats, and the poriferan:green algae partnerships offers unique opportunities to study the evolutionary origins and ecological persistence of endosymbioses. We examined the association between Ephydatia muelleri and its chlorophyte partner to identify features of host cellular and genetic responses to the presence of intracellular algal partners. Chlorella-like green algal symbionts were isolated from field-collected adult E. muelleri tissue harboring algae. The sponge-derived algae were successfully cultured and subsequently used to reinfect aposymbiotic E. muelleri tissue. We used confocal microscopy to follow the fate of the sponge-derived algae after inoculating algae-free E. muelleri grown from gemmules to show temporal patterns of symbiont location within host tissue. We also infected aposymbiotic E. muelleri with sponge-derived algae, and performed RNASeq to study differential expression patterns in the host relative to symbiotic states. We compare and contrast our findings with work in other systems (e.g., endosymbiotic Hydra) to explore possible conserved evolutionary pathways that may lead to stable mutualistic endosymbioses. Our work demonstrates that freshwater sponges offer many tractable qualities to study features of intracellular occupancy and thus meet criteria desired for a model system. creator: Chelsea Hall creator: Sara Camilli creator: Henry Dwaah creator: Benjamin Kornegay creator: Christie Lacy creator: Malcolm S. Hill creator: April L. Hill uri: https://doi.org/10.7717/peerj.10654 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Hall et al. title: Taxonomic revision of the threatened African genus Pseudohydrosme Engl. (Araceae), with P. ebo, a new, critically endangered species from Ebo, Cameroon link: https://peerj.com/articles/10689 last-modified: 2021-02-11 description: This is the first revision in more than 100 years of the African genus Pseudohydrosme, formerly considered endemic to Gabon. Closely related to Anchomanes, Pseudohydrosme is distinct from Anchomanes because of its 2-3-locular ovary (vs. unilocular), peduncle concealed by cataphylls at anthesis and far shorter than the spathe (vs. exposed, far exceeding the spathe), stipitate fruits and viviparous (asexually reproductive) roots (vs. sessile, roots non-viviparous), lack of laticifers (vs. laticifers present) and differences in spadix: spathe proportions and presentation. However, it is possible that a well sampled molecular phylogenetic analysis might show that one of these genera is nested inside the other. In this case the synonymisation of Pseudohydrosme will be required. Three species, one new to science, are recognised, in two sections. Although doubt has previously been cast on the value of recognising Pseudohydrosme buettneri, of Gabon, it is here accepted and maintained as a distinct species in the monotypic section, Zyganthera. However, it is considered to be probably globally extinct. Pseudohydrosme gabunensis, type species of the genus, also Gabonese but probably extending to Congo, is maintained in Sect. Pseudohydrosme together with Pseudohydrosme ebo sp.nov. of the Ebo Forest, Littoral Region, Cameroon, the first addition to the genus since the nineteenth century, and which extends the range of the genus 450 km north from Gabon, into the Cross-Sanaga biogeographic area. The discovery of Pseudohydrosme ebo resulted from a series of surveys for conservation management in Cameroon, and triggered this article. All three species are morphologically characterised, their habitat and biogeography discussed, and their extinction risks are respectively assessed as Critically Endangered (Possibly Extinct), Endangered and Critically Endangered using the IUCN standard. Clearance of forest habitat for logging, followed by agriculture or urbanisation are major threats. Pseudohydrosme gabunensis may occur in a formally protected area and is also cultivated widely but infrequently in Europe, Australia and the USA for its spectacular inflorescences. creator: Martin Cheek creator: Barthélemy Tchiengué creator: Xander van der Burgt uri: https://doi.org/10.7717/peerj.10689 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Cheek et al. title: Rapid relative increase of crustose coralline algae following herbivore exclusion in a reef of El Salvador link: https://peerj.com/articles/10696 last-modified: 2021-02-11 description: The Eastern Tropical Pacific (ETP) is one of the most isolated and least studied regions in the world. This particularly applies to the coast of El Salvador, where the only reef between Guatemala and Nicaragua, called Los Cóbanos reef, is located. There is very little published information about the reef’s biodiversity, and to our knowledge, no research on its ecology and responses to anthropogenic impacts, such as overfishing, has been conducted. The present study, therefore, described the benthic community of Los Cóbanos reef, El Salvador, using the Line-Point-Intercept-Transect method and investigated changes in the benthic community following the exclusion of piscine macroherbivores over a period of seven weeks. Results showed high benthic algae cover (up to 98%), dominated by turf and green algae, and low coral cover (0–4%). Porites lobata was the only hermatypic coral species found during the surveys. Surprisingly, crustose coralline algae (CCA) showed a remarkable total cover increase by 58%, while turf algae cover decreased by 82%, in experimental plots after seven weeks of piscine macroherbivore exclusion. These findings apparently contradict the results of most previous similar studies. While it was not possible to ascertain the exact mechanisms leading to these drastic community changes, the most likely explanation is grazing on turf by small grazing macroherbivores that had access to the cages during the experiment and clearing of CCA initially covered by epiphytes and sediments. A higher CCA cover would promote the succesful settlement by corals and prevent further erosion of the reef framework. Therefore it is crucial to better understand algal dynamics, herbivory, and implications of overfishing at Los Cóbanos to avoid further reef deterioration. This could be achieved through video surveys of the fish community, night-time observations of the macroinvertebrate community, exclusion experiments that also keep out herbivorous macroinvertebrates, and/or experimental assessments of turf algae/CCA interactions. creator: Xochitl E. Elías Ilosvay creator: Johanna Segovia creator: Sebastian Ferse creator: Walter Ernesto Elias creator: Christian Wild uri: https://doi.org/10.7717/peerj.10696 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Elías Ilosvay et al. title: A new elasmosaurid (Sauropterygia: Plesiosauria) from the non-marine to paralic Dinosaur Park Formation of southern Alberta, Canada link: https://peerj.com/articles/10720 last-modified: 2021-02-11 description: Elasmosaurid plesiosaurian remains have been documented from non-marine to paralic (fluvial to estuarine) sediments of the upper Campanian Dinosaur Park Formation (DPF) of southern Alberta since 1898. Despite this long collection history, this material has received relatively little research attention, largely due to the highly fragmentary nature of most recovered specimens. However, this assemblage is significant, as it constitutes a rare occurrence of plesiosaurian remains in a non-marine depositional environment. This study reports on a recently collected and prepared specimen, which represents the most complete elasmosaurid yet collected from the DPF. This specimen preserves the trunk region, the base of the neck and tail, a partial fore and hind limb, and tooth, and is sufficiently complete to be assigned as the holotype of a new genus and species. This new taxon is diagnosed by a distinctive character state combination including a boomerang-shaped clavicular arch with acute anterior process, convex anterolateral margin, deeply embayed posterior margin, and pronounced ventral keel, together with the presence of 22 dorsal vertebrae, and the anterior dorsal centra bearing a ventral notch. The DPF plesiosaurian fossils were recovered from both estuarine/bay and fluvial palaeochannel sediments. The holotype skeleton represents an osteologically mature individual with an estimated body length of around 5 m, although the largest referred DPF elasmosaurid might have been closer to 7 m, which is considerably larger than other plesiosaurians reported from non-marine deposits. This suggests small-body lengths relative to typical elasmosaurids from marine settings, but is consistent with other plesiosaurians recovered from non-marine sediments. The identification of a distinct elasmosaurid taxon in the DPF might be evidence of niche-partitioning among the predominantly oceanic members of the ubiquitous plesiosaurian clade. creator: James A. Campbell creator: Mark T. Mitchell creator: Michael J. Ryan creator: Jason S. Anderson uri: https://doi.org/10.7717/peerj.10720 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Campbell et al. title: Transcriptomic analysis of nonylphenol effect on Saccharomyces cerevisiae link: https://peerj.com/articles/10794 last-modified: 2021-02-11 description: Nonylphenol (NP) is a bioaccumulative environmental estrogen that is widely used as a nonionic surfactant. We have previously examined short-term effects of NP on yeast cells using microarray technology. In the present study, we investigated the adaptive response of Saccharomyces cerevisiae BY4742 cells to NP exposure by analyzing genome-wide transcriptional profiles using RNA-sequencing. We used 2 mg/L NP concentration for 40 days of exposure. Gene expression analysis showed that a total of 948 genes were differentially expressed. Of these, 834 genes were downregulated, while 114 genes were significantly upregulated. GO enrichment analysis revealed that 369 GO terms were significantly affected by NP exposure. Further analysis showed that many of the differentially expressed genes were associated with oxidative phosphorylation, iron and copper acquisition, autophagy, pleiotropic drug resistance and cell cycle progression related processes such as DNA and mismatch repair, chromosome segregation, spindle checkpoint activity, and kinetochore organization. Overall, these results provide considerable information and a comprehensive understanding of the adaptive response to NP exposure at the gene expression level. creator: Ceyhun Bereketoglu creator: Gozde Nacar creator: Tugba Sari creator: Bulent Mertoglu creator: Ajay Pradhan uri: https://doi.org/10.7717/peerj.10794 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Bereketoglu et al. title: Low pH alleviated salinity stress of ginger seedlings by enhancing photosynthesis, fluorescence, and mineral element contents link: https://peerj.com/articles/10832 last-modified: 2021-02-11 description: We investigated the effects of low pH on the photosynthesis, chlorophyll fluorescence, and mineral contents of the leaves of ginger plants under salt stress. This experiment involved four treatments: T1 (pH 6, 0 salinity), T2 (pH 4, 0 salinity), T3 (pH 6, 100 mmol L−1 salinity) and T4 (pH 4, 100 mmol L−1 salinity). This study showed that photosynthesis (Pn, Gs, WUE and Tr) and chlorophyll fluorescence (qP, Φ PSII, and Fv/Fm) significantly decreased under salt stress; however, all the parameters of the ginger plants under the low-pH treatment and salt stress recovered. Moreover, low pH reduced the content of Na and enhanced the contents of K, Mg, Fe and Zn in the leaves of ginger plants under salt stress. Taken together, these results suggest that low pH improves photosynthesis efficiency and nutrient acquisition and reduces the absorption of Na, which could enhance the salt tolerance of ginger. creator: Fengman Yin creator: Shanying Zhang creator: Bili Cao creator: Kun Xu uri: https://doi.org/10.7717/peerj.10832 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Yin et al. title: Using natural history collections to investigate changes in pangolin (Pholidota: Manidae) geographic ranges through time link: https://peerj.com/articles/10843 last-modified: 2021-02-11 description: Pangolins, often considered the world’s most trafficked wild mammals, have continued to experience rapid declines across Asia and Africa. All eight species are classed as either Vulnerable, Endangered or Critically Endangered by the International Union for Conservation of Nature (IUCN) Red List. Alongside habitat loss, they are threatened mainly by poaching and/or legal hunting to meet the growing consumer demand for their meat and keratinous scales. Species threat assessments heavily rely on changes in species distributions which are usually expensive and difficult to monitor, especially for rare and cryptic species like pangolins. Furthermore, recent assessments of the threats to pangolins focus on characterising their trade using seizure data which provide limited insights into the true extent of global pangolin declines. As the consequences of habitat modifications and poaching/hunting on species continues to become apparent, it is crucial that we frequently update our understanding of how species distributions change through time to allow effective identification of geographic regions that are in need of urgent conservation actions. Here we show how georeferencing pangolin specimens from natural history collections can reveal how their distributions are changing over time, by comparing overlap between specimen localities and current area of habitat maps derived from IUCN range maps. We found significant correlations in percentage area overlap between species, continent, IUCN Red List status and collection year, but not ecology (terrestrial or arboreal/semi-arboreal). Human population density (widely considered to be an indication of trafficking pressure) and changes in primary forest cover, were weakly correlated with percentage overlap. Our results do not suggest a single mechanism for differences among historical distributions and present-day ranges, but rather show that multiple explanatory factors must be considered when researching pangolin population declines as variations among species influence range fluctuations. We also demonstrate how natural history collections can provide temporal information on distributions and discuss the limitations of collecting and using historical data. creator: Emily Buckingham creator: Jake Curry creator: Charles Emogor creator: Louise Tomsett creator: Natalie Cooper uri: https://doi.org/10.7717/peerj.10843 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Buckingham et al. title: Arousal modulates the motor interference effect stimulated by pictures of threatening animals link: https://peerj.com/articles/10876 last-modified: 2021-02-11 description: Previous research related to the motor interference effect from dangerous objects indicated that delayed responses to dangerous objects were associated with more positive parietal P3 amplitudes, suggesting that great attentional resources were allocated to evaluate the level of danger (i.e., negative valence). However, arousal covaried with valence in this research. Together with previous studies in which the P3 amplitude was found to be increased along with a higher arousal level in the parietal lobe, we raised the issue that more positive parietal P3 amplitudes might also be affected by a high arousal level. To clarify whether valence or arousal impacted the motor interference effect, this study used a motor priming paradigm mixed with a Go/NoGo task and manipulated the valence (negative, neutral and positive) and arousal (medium and high) of target stimuli. Analysis of the behavioral results identified a significant motor interference effect (longer reaction times (RTs) in the negative valence condition than in the neutral valence condition) at the medium arousal level and an increased effect size (increment of RT difference) at the high arousal level. The results indicated that negative valence stimuli may interfere with the prime elicited motor preparation more strongly at the high arousal level than at the medium arousal level. The ERP results identified larger centroparietal P3 amplitudes for the negative valence condition than for the neutral valence condition at a high arousal level. However, the inverse result, i.e., lower centroparietal P3 amplitudes for the negative valence condition than for the neutral valence condition, was observed at a medium arousal level. The ERP results further indicated that the effect size of the behavioral motor interference effect increased because subjects are more sensitive to the negative valence stimuli at the high arousal level than at the medium arousal level. Furthermore, the motor interference effect is related to the negative valence rather than emotionality of the target stimuli because different result patterns emerged between the positive and negative valence conditions. Detailed processes underlying the interaction between valence and arousal effects are discussed. creator: Gai Cao creator: Peng Liu uri: https://doi.org/10.7717/peerj.10876 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Cao and Liu title: Characterization and complete genome sequence of Privateer, a highly prolate Proteus mirabilis podophage link: https://peerj.com/articles/10645 last-modified: 2021-02-10 description: The Gram-negative bacterium Proteus mirabilis causes a large proportion of catheter-associated urinary tract infections, which are among the world’s most common nosocomial infections. Here, we characterize P. mirabilis bacteriophage Privateer, a prolate podophage of the C3 morphotype isolated from Texas wastewater treatment plant activated sludge. Basic characterization assays demonstrated Privateer has a latent period of ~40 min and average burst size around 140. In the 90.7 kb Privateer genome, 43 functions were assigned for the 144 predicted protein-coding genes. Genes encoding DNA replication proteins, DNA modification proteins, four tRNAs, lysis proteins, and structural proteins were identified. Cesium-gradient purified Privateer particles analyzed via LC-MS/MS verified the presence of several predicted structural proteins, including a longer, minor capsid protein apparently produced by translational frameshift. Comparative analysis demonstrated Privateer shares 83% nucleotide similarity with Cronobacter phage vB_CsaP_009, but low nucleotide similarity with other known phages. Predicted structural proteins in Privateer appear to have evolutionary relationships with other prolate podophages, in particular the Kuraviruses creator: James E. Corban creator: Jolene Ramsey uri: https://doi.org/10.7717/peerj.10645 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Corban and Ramsey title: A validation of Illumina EPIC array system with bisulfite-based amplicon sequencing link: https://peerj.com/articles/10762 last-modified: 2021-02-10 description: The Illumina Infinium® MethylationEPIC BeadChip system (hereafter EPIC array) is considered to be the current gold standard detection method for assessing DNA methylation at the genome-wide level. EPIC arrays are often used for hypothesis generation or pilot studies, the natural conclusion to which is to validate methylation candidates and expand these in a larger cohort, in a targeted manner. As such, an accurate smaller-scale, targeted technique, that generates data at the individual CpG level that is equivalent to the EPIC array, is needed. Here, we tested an alternative DNA methylation detection technique, known as bisulfite-based amplicon sequencing (BSAS), to determine its ability to validate CpG sites detected in EPIC array studies. BSAS was able to detect differential DNA methylation at CpG sites to a degree which correlates highly with the EPIC array system at some loci. However, BSAS correlated less well with EPIC array data in some instances, and most notably, when the magnitude of change via EPIC array was greater than 5%. Therefore, our data suggests that BSAS can be used to validate EPIC array data, but each locus must be compared on an individual basis, before being taken forward into large scale screening. Further, BSAS does offer advantages compared to the probe-based EPIC array; BSAS amplifies a region of the genome (∼500 bp) around a CpG of interest, allowing analyses of other CpGs in the region that may not be present on the EPIC array, aiding discovery of novel CpG sites and differentially methylated regions of interest. We conclude that BSAS offers a valid investigative tool for specific regions of the genome that are currently not contained on the array system. creator: Alexandra J. Noble creator: John F. Pearson creator: Joseph M. Boden creator: L. John Horwood creator: Neil J. Gemmell creator: Martin A. Kennedy creator: Amy J. Osborne uri: https://doi.org/10.7717/peerj.10762 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Noble et al. title: Comparative analysis of the complete plastid genomes of Mangifera species and gene transfer between plastid and mitochondrial genomes link: https://peerj.com/articles/10774 last-modified: 2021-02-10 description: Mango is an important commercial fruit crop belonging to the genus Mangifera. In this study, we reported and compared four newly sequenced plastid genomes of the genus Mangifera, which showed high similarities in overall size (157,780–157,853 bp), genome structure, gene order, and gene content. Three mutation hotspots (trnG-psbZ, psbD-trnT, and ycf4-cemA) were identified as candidate DNA barcodes for Mangifera. These three DNA barcode candidate sequences have high species identification ability. We also identified 12 large fragments that were transferred from the plastid genome to the mitochondrial genome, and found that the similarity was more than 99%. The total size of the transferred fragment was 35,652 bp, accounting for 22.6% of the plastid genome. Fifteen intact chloroplast genes, four tRNAs and numerous partial genes and intergenic spacer regions were identified. There are many of these genes transferred from mitochondria to the chloroplast in other species genomes. Phylogenetic analysis based on whole plastid genome data provided a high support value, and the interspecies relationships within Mangifera were resolved well. creator: Yingfeng Niu creator: Chengwen Gao creator: Jin Liu uri: https://doi.org/10.7717/peerj.10774 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Niu et al. title: Somatic determinants of changes in selected body posture parameters in younger school-age children link: https://peerj.com/articles/10821 last-modified: 2021-02-10 description: BackgroundThe aim of this study was to describe changes in selected parameters of body posture in children between 5 and 9 years old with diversified somatic structures.MethodsThe study was carried out in 2015 and then repeated in 2018 among 67 participants who had previously been observed to have scoliotic posture. Basic body weight and height measurements were taken, which were then used to calculate the body mass index. Posture tests were conducted using the photogrammetric method.ResultsGirls and boys were not significantly different in body dimensions. With age, the number of overweight boys and children with normal growth-weight proportions increased. Temporary differences in posture variables indicating abnormalities were small.ConclusionsThere were no significant differences in somatic parameters between the girls and the boys. Those children with a slender body structure had the most abnormalities in the coronal plane. On the other hand, changes in spinal position in the sagittal plane were more frequent in overweight children. Relations were noted between the compensation index in the sagittal plane and deviation of the trunk inclination, the maximum deviation of the line of the spinous processes, and the angle of the shoulder line in the coronal plane and body mass index values were noted. creator: Marta Kinga Labecka creator: Krystyna Górniak creator: Małgorzata Lichota uri: https://doi.org/10.7717/peerj.10821 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Labecka et al. title: Marine amphipods as a new live prey for ornamental aquaculture: exploring the potential of Parhyale hawaiensis and Elasmopus pectenicrus link: https://peerj.com/articles/10840 last-modified: 2021-02-10 description: Marine amphipods are gaining attention in aquaculture as a natural live food alternative to traditional preys such as brine shrimps (Artemia spp.). The use of Artemia is convenient for the culture of many marine species, but often problematic for some others, such as seahorses and other marine ornamental species. Unlike Artemia, marine amphipods are consumed by fish in their natural environment and show biochemical profiles that better match the nutritional requirements of marine fish, particularly of polyunsaturated fatty acids (PUFA), including eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids. Despite their potentially easy culture, there are no established culture techniques and a deeper knowledge on the reproductive biology, nutritional profiles and culture methodologies is still needed to potentiate the optimization of mass production. The present study assessed, for the first time, the aquaculture potential of Parhyale hawaiensis and Elasmopus pectenicrus, two cosmopolitan marine gammarids (as per traditional schemes of classification) that naturally proliferate in the wild and in aquaculture facilities. For that purpose, aspects of the population and reproductive biology of the species were characterized and then a series of laboratory-scale experiments were conducted to determine amphipod productivity, the time needed to reach sexual maturity by hatchlings (generation time), cannibalism degree, the effects of sex ratio on fecundity and the effects of diet (shrimp diet, plant-based diet and commercial fish diet) on fecundity and juvenile growth. P. hawaiensis, unlike E. pectenicrus, was easily maintained and propagated in laboratory conditions. P. hawaiensis showed a higher total length (9.3 ± 1.3 mm), wet weight (14.4 ± 6.2 mg), dry weight (10.5 ± 4.4 mg), females/males sex ratio (2.24), fecundity (12.8 ± 5.7 embryos per female), and gross energy content (16.71 ± 0.67 kJ g-1) compared to E. pectenicrus (7.9 ± 1.2 mm total length; 8.4 ± 4.3 mg wet weight; 5.7 ± 3.2 mg dry weight; 1.34 females/males sex ratio; 6.5 ± 3.9 embryos per female; 12.86 ± 0.82 kJ g−1 gross energy content). P. hawaiensis juvenile growth showed a small, but significant, reduction by the use of a plant-based diet compared to a commercial shrimp and fish diet; however, fecundity was not affected, supporting the possible use of inexpensive diets to mass produce amphipods as live or frozen food. Possible limitations of P. hawaiensis could be their quite long generation times (50.9 ± 5.8 days) and relatively low fecundity levels (12.8 ± 5.7 embryos per female). With an observed productivity rate of 0.36 ± 0.08 juveniles per amphipod couple per day, P. hawaiensis could become a specialty feed for species that cannot easily transition to a formulated diet such as seahorses and other highly priced marine ornamental species. creator: Jorge Arturo Vargas-Abúndez creator: Humberto Ivan López-Vázquez creator: Maite Mascaró creator: Gemma Leticia Martínez-Moreno creator: Nuno Simões uri: https://doi.org/10.7717/peerj.10840 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Vargas-Abúndez et al. title: Genome size versus geographic range size in birds link: https://peerj.com/articles/10868 last-modified: 2021-02-10 description: Why do some species occur in small, restricted areas, while others are distributed globally? Environmental heterogeneity increases with area and so does the number of species. Hence, diverse biotic and abiotic conditions across large ranges may lead to specific adaptations that are often linked to a species’ genome size and chromosome number. Therefore, a positive association between genome size and geographic range is anticipated. Moreover, high cognitive ability in organisms would be favored by natural selection to cope with the dynamic conditions within large geographic ranges. Here, we tested these hypotheses in birds—the most mobile terrestrial vertebrates—and accounted for the effects of various confounding variables, such as body mass, relative brain mass, and geographic latitude. Using phylogenetic generalized least squares and phylogenetic confirmatory path analysis, we demonstrated that range size is positively associated with bird genome size but probably not with chromosome number. Moreover, relative brain mass had no effect on range size, whereas body mass had a possible weak and negative effect, and range size was larger at higher geographic latitudes. However, our models did not fully explain the overall variation in range size. Hence, natural selection may impose larger genomes in birds with larger geographic ranges, although there may be additional explanations for this phenomenon. creator: Beata Grzywacz creator: Piotr Skórka uri: https://doi.org/10.7717/peerj.10868 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Grzywacz and Skórka title: Exploring the occurrence of thioflavin-T-positive insulin amyloid aggregation intermediates link: https://peerj.com/articles/10918 last-modified: 2021-02-10 description: The aggregation of proteins is considered to be the main cause of several neurodegenerative diseases. Despite much progress in amyloid research, the process of fibrillization is still not fully understood, which is one of the main reasons why there are still very few effective treatments available. When the aggregation of insulin, a model amyloidogenic protein, is tracked using thioflavin-T (ThT), an amyloid specific dye, there is an anomalous occurrence of double-sigmoidal aggregation kinetics. Such an event is likely related to the formation of ThT-positive intermediates, which may affect the outcome of both aggregation kinetic data, as well as final fibril structure. In this work we explore insulin fibrillization under conditions, where both normal and double-sigmoidal kinetics are observed and show that, despite their dye-binding properties and random occurrence, the ThT-positive intermediates do not significantly alter the overall aggregation process. creator: Mantas Ziaunys creator: Andrius Sakalauskas creator: Kamile Mikalauskaite creator: Vytautas Smirnovas uri: https://doi.org/10.7717/peerj.10918 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Ziaunys et al. title: Bacterial community diversity, lignocellulose components, and histological changes in composting using agricultural straws for Agaricus bisporus production link: https://peerj.com/articles/10452 last-modified: 2021-02-09 description: Agricultural straws (AS) may serve as potential base-substances in the production of Agaricus bisporus. Six AS that occur across China were investigated in a two-stage composting experiment; lignocellulose components, AS morphology, and the effects of different AS on mushroom yields from 2015–2017 were examined. In addition, microbial biodiversity and their impact on substrate degradation were studied using 16S gene sequenc based on six different AS on the 3rd (I.F), 6th (I.S), and 10th (I.T) day of Phase I, and Phase II (II). Results showed that the six different AS exhibited differences in the progression of degradation under the same compost condition; the wheat straw, rice straw, and cotton straw induced a significantly higher mushroom yield than did the others (P < 0.05); Thermobispora, Thermopolyspora, and Vulgatibacter genera may play an important role in the different AS degradations. According to our experiments, we can adjust formulations and compost methods to obtain high-yield mushroom compost based on different AS in the future. creator: Tingting Song creator: Yingyue Shen creator: Qunli Jin creator: Weilin Feng creator: Lijun Fan creator: Guangtian Cao creator: Weiming Cai uri: https://doi.org/10.7717/peerj.10452 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Song et al. title: A risk score model with five long non-coding RNAs for predicting prognosis in gastric cancer: an integrated analysis combining TCGA and GEO datasets link: https://peerj.com/articles/10556 last-modified: 2021-02-09 description: BackgroundGastric cancer (GC) is one of the most common carcinomas of the digestive tract, and the prognosis for these patients may be poor. There is evidence that some long non-coding RNAs(lncRNAs) can predict the prognosis of patients with GC. However, few lncRNA signatures have been used to predict prognosis. Herein, we aimed to construct a risk score model based on the expression of five lncRNAs to predict the prognosis of patients with GC and provide new potential therapeutic targets.MethodsWe performed differentially expressed and survival analyses to identify differentially expressed survival-ralated lncRNAs by using GC patient expression profile data from The Cancer Genome Atlas (TCGA) database. We then established a formula including five lncRNAs to predict the prognosis of patients with GC. In addition, to verify the prognostic value of this risk score model, two independent Gene Expression Omnibus (GEO) datasets, GSE62254 (N = 300) and GSE15459 (N = 200), were employed as validation groups.ResultsBased on the characteristics of five lncRNAs, patients with GC were divided into high or low risk subgroups. The prognostic value of the risk score model with five lncRNAs was confirmed in both TCGA and the two independent GEO datasets. Furthermore, stratification analysis results showed that this model had an independent prognostic value in patients with stage II–IV GC. We constructed a nomogram model combining clinical factors and the five lncRNAs to increase the accuracy of prognostic prediction. Enrichment analysis based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) suggested that the five lncRNAs are associated with multiple cancer occurrence and progression-related pathways.ConclusionThe risk score model including five lncRNAs can predict the prognosis of patients with GC, especially those with stage II-IV, and may provide potential therapeutic targets in future. creator: Yiguo Wu creator: Junping Deng creator: Shuhui Lai creator: Yujuan You creator: Jing Wu uri: https://doi.org/10.7717/peerj.10556 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wu et al. title: Evaluation of resistance to wheat stem rust and identification of resistance genes in wheat lines from Heilongjiang province link: https://peerj.com/articles/10580 last-modified: 2021-02-09 description: Wheat stem rust, caused by Puccinia graminis f. sp. tritici, (Pgt) is a devastating disease in wheat production. The disease has been effectively controlled since the 1970s due to the widespread use of the Sr31 resistance gene. However, Sr31 has lost its effectiveness following the emergence and spread of the Ug99 race variants. Therefore, there is an urgent global effort to identify new germplasm resources effective against those races. In this study, the resistance to Pgt of 95 wheat advance lines from Heilongjiang Province was evaluated using three predominant races of Pgt, 21C3CTTTM, 34C0MKGSM, and 34C3MTGQM, in China at the seedling and adult plant stage. The presence of 6 Sr genes (Sr2, Sr24, Sr25, Sr26, Sr31, and Sr38) was evaluated using linked molecular markers. The results showed that 86 (90.5%) wheat lines had plant stage resistance to all three races. Molecular marker analysis showed that 24 wheat lines likely carried Sr38, 15 wheat lines likely carried Sr2, 11 wheat lines likely carried Sr31, while none of the wheat lines carried Sr24, Sr25, or Sr26. Furthermore, six out of the 95 wheat lines tested carried both Sr2 and Sr38, three contained both Sr31 and Sr38, and two wheat lines contained both Sr2 and Sr31. Wheat lines with known Sr genes may be used as donor parents for further breeding programs to provide resistance to stem rust. creator: Qiujun Lin creator: Yue Gao creator: Xianxin Wu creator: Xinyu Ni creator: Rongzhen Chen creator: Yuanhu Xuan creator: Tianya Li uri: https://doi.org/10.7717/peerj.10580 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Lin et al. title: Transcriptome analysis reveals the mechanism of improving erect-plant-type peanut yield by single-seeding precision sowing link: https://peerj.com/articles/10616 last-modified: 2021-02-09 description: BackgroundIn China, double-seed (DS) sowing (i.e., sowing two seeds per hole) has been conventionally performed towards the erect-plant-type peanuts to increase the low germination rate due to poor seed preservation conditions. However, the corresponding within-hole plant competition usually limits the subsequent plant growth and the final yield. We developed a high-yield cultivation system of single-seed (SS) precision sowing to solve this paradox, saving 20% of seeds and increasing yields by more than 10% relative to the conventional DS sowing.MethodsTo explore the mechanisms of these two different cropping patterns in peanut yields, we conducted transcriptomic and physiological comparisons in the seeding plant leaf and root tissues between SS precision sowing and standard DS sowing treatments.ResultsAfter assembly, each library contained an average of 43 million reads and generated a total of 523,800, 338 clean reads. After GO and Kyoto Encyclopedia of Genes and Genomes pathway analysis, we found the key genes for biotic and abiotic stress showed higher expression in roots of plants grown under the SS precision sowing treatment, including genes encoding disease resistance, oxidation-reduction, hormone related, and stress response transcription factors and signaling regulation proteins. In particular, the resveratrol synthesis genes related to stress and disease resistance appeared induced in roots under the SS sowing treatment.ConclusionThese data indicated that Aspergillus flavus resistance and stress tolerance in roots under SS precision sowing were enhanced compared with roots under the DS sowing treatment. This work benefits the development of underground pods and thus increasing peanut yields. creator: Sha Yang creator: Jialei Zhang creator: Yun Geng creator: Zhaohui Tang creator: Jianguo Wang creator: Feng Guo creator: Jingjing Meng creator: Quan Wang creator: Shubo Wan creator: Xinguo Li uri: https://doi.org/10.7717/peerj.10616 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Yang et al. title: The potential for income improvement and biodiversity conservation via specialty coffee in Ethiopia link: https://peerj.com/articles/10621 last-modified: 2021-02-09 description: The specialty coffee sector represents opportunities for coffee farmers and other actors due to increased value within the supply chain, driven by elevated consumer purchase prices. We investigated these relationships up to the export stage, as well as the potential for specialty coffee to improve stakeholder participation in biodiversity conservation. Household data was collected from a sample of 272 coffee farmers belonging to five primary cooperatives, in the heavily forested area of Yayu, in the Illubabor administrative zone of Oromia regional state in south-western Ethiopia, for the 2017/2018 harvest season. Qualitative and quantitative data, gathered between 2015 and 2019, from focus group discussions, was used to supplement the survey and to explain the empirical findings. We show that the income from coffee (and thus household income) can be increased, in some cases substantially, via participation in the specialty coffee market. The unit price for coffee was much higher than standard market prices and those including certification premiums. Whilst quality is a key factor for specialty coffee, income increases via the specialty market are caveat bound. In particular, there is a critical requirement for efficient and effective cooperative management, and we provide suggestions for improvements. In the long-term, more focus is needed to increase the financial and human capacities of Ethiopian coffee cooperatives, to make them more effective partners in the specialty coffee value chain. The high conversion rates from harvested to clean exportable specialty coffee (8:1, or higher) is identified as a major constraint on profitability. We show that specialty coffee can bring about positive environmental benefits. Elevating coffee prices and farm profitability to bring household income levels to around or above the global poverty line may ensure the continuation of forest coffee production, and thus the retention of forest. The increases in income via specialty coffee production, reported in the study, were achieved without increasing land use or the application of environmentally costly inputs. Moreover, analysis of satellite data shows that a large proportion of the coffee farms surveyed retain a level of forest cover and quality approaching primary (undisturbed) forest, and that the coffee production area at Yayu has not experienced any significant deforestation (since 2000). We recommend that coffee premiums linked to environmental benefit should demonstrate clearly defined and appropriate metrics, as we have demonstrated here for forest (canopy) quality and coverage (area). creator: Pascale Schuit creator: Justin Moat creator: Tadesse Woldemariam Gole creator: Zeleke Kebebew Challa creator: Jeremy Torz creator: Steven Macatonia creator: Graciano Cruz creator: Aaron P. Davis uri: https://doi.org/10.7717/peerj.10621 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Schuit et al. title: The Slavcleft: a three-center study of the outcome of treatment of cleft lip and palate. Nasolabial appearance link: https://peerj.com/articles/10631 last-modified: 2021-02-09 description: BackgroundThere is a multitude of protocols of treatment of cleft lip and palate (CLP) worldwide differing in number of operations, surgical techniques, and timings of surgeries. Despite, facial appearance in subjects with CLP is rarely ideal and residual stigmata are easy to notice in many patients irrespective of the protocol. The prospective controlled investigations are optimal for comparing effectiveness of treatment protocols. Because prospective studies are very challenging to perform in CLP field, it is reasonable to retrospectively assess different surgical protocols to identify the promising ones and then to test them in a prospective way.MethodsOur objective was to assess the nasolabial appearance in a preadolescent Slavic population with unilateral cleft lip and palate (UCLP) by using the 0–200 numeric scale with reference photographs. Patients treated in Warsaw, Poland (n = 32), Prague, Czech Republic (n = 26) and Bratislava, Slovakia (n = 17) were included in this retrospective study. Each cleft center used a unique surgical protocol. Two panels of professional raters (n = 7) and laypeople (n = 10) scored blindly the nasolabial esthetics on cropped frontal and profile images with cropped reference photograph present on the same slide. Intra- and inter-rater agreement was assessed with Cronbach’s alpha, intraclass correlation coefficients, t-tests, and Bland–Altman plots. Inter-group differences were evaluated with one-way ANOVA and regression analysis.ResultsThe agreement within and between raters was acceptable. We found that patients treated in Warsaw, Prague, and Bratislava showed comparable nasolabial appearance on frontal and profile photographs when judged by both professional raters (p > 0.05) and laypeople (p > 0.05). Regression analysis did not identify influence of gender, group (i.e., Warsaw, Prague, and Bratislava), age at lip repair, surgeon, and age at photographic assessment on esthetic outcome (p > 0.05).ConclusionThis study showed that none of the surgical protocols showed superiority to produce good nasolabial appearance. creator: Adam Stebel creator: Wanda Urbanová creator: Irena Klimova creator: Andrzej Brudnicki creator: Ivana Dubovska creator: Petra Polackova creator: Daniela Kroupová creator: Magdalena Koťová creator: Piotr S. Fudalej uri: https://doi.org/10.7717/peerj.10631 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Stebel et al. title: TET2 mutations in acute myeloid leukemia: a comprehensive study in patients of Sindh, Pakistan link: https://peerj.com/articles/10678 last-modified: 2021-02-09 description: BackgroundThe tet oncogene family member 2 (TET2) gene has been reported to be involved in DNA methylation and epigenetic regulation in acute myeloid leukemia (AML). Various studies have proven functional role of TET2 mutations in AML. We herein studied the frequency and genotype-phenotype correlation of TET2 gene in AML patients in Sindh, Pakistan.Patients and methodsThe current study was carried out at Liaquat University of Medical & Health Sciences, Jamshoro, Pakistan, in collaboration with National Institute of Blood Disease & Bone Marrow Transplant, Karachi, Pakistan, during the period from June 2019 to June 2020. A total of 130 patients diagnosed with AML were screened for TET2 mutations. Whole exome sequencing of 14 individuals was carried out to find the genetic variants in TET2 gene. The pathogenicity of the variants was predicted by SIFT, PolyPhen2, Mutation Taster and CADD Phred scores. The allele frequency of the variants was compared with global population using 1000 genomes project and Exome Aggregation Consortium (ExAC). Furthermore, exon 3 and exon 5 of the TET2 gene were sequenced by using Sanger sequencing. The findings were correlated with subtypes of AML and corresponding karyotypes.ResultsThrough the exome sequencing, 17 genetic variants (13 SNPs and four indels) were identified in 14 individuals. Of these, four variants that is, one frameshift deletion, one frameshift insertion and two nonsense variants were novel and not present in dbSNP151 database. Three novel variants were found in exon 3 including two frameshift variants that is, p.T395fs and G494fs, predicted as deleterious by CADD Phred scores, and one stop-gain variant (p.G898X) predicted as deleterious by Mutation Taster and CADD Phred scores. One novel non sense variant (p.Q1191X) was found in the exon 5 predicted as deleterious by SIFT, Mutation Taster and CADD Phred scores. Sanger sequencing analysis revealed one novel deletion at g105233851: del.TAGATAGA, and one novel SNP g;105233861 T>G identified in the TET2 gene. Majority of the exon 3 mutations were seen in the patients diagnosed with AML with maturation, and had a normal karyotype.ConclusionTET2 mutations were identified in around 16% of the total patients of our study indicating other mechanisms being involved in pathophysiology of AML in this cohort. The TET2 mutations provide a prognostic value in determining AML classification. creator: Abdul Rehman Khalil Shaikh creator: Ikram Ujjan creator: Muhammad Irfan creator: Arshi Naz creator: Tahir Shamsi creator: Muhammad Tariq Masood Khan creator: Muhammad Shakeel uri: https://doi.org/10.7717/peerj.10678 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Shaikh et al. title: It’s all about time: precision and accuracy of Emotiv event-marking for ERP research link: https://peerj.com/articles/10700 last-modified: 2021-02-09 description: BackgroundThe use of consumer-grade electroencephalography (EEG) systems for research purposes has become more prevalent. In event-related potential (ERP) research, it is critical that these systems have precise and accurate timing. The aim of the current study was to investigate the timing reliability of event-marking solutions used with Emotiv commercial EEG systems.MethodWe conducted three experiments. In Experiment 1 we established a jitter threshold (i.e. the point at which jitter made an event-marking method unreliable). To do this, we introduced statistical noise to the temporal position of event-marks of a pre-existing ERP dataset (recorded with a research-grade system, Neuroscan SynAmps2 at 1,000 Hz using parallel-port event-marking) and calculated the level at which the waveform peaks differed statistically from the original waveform. In Experiment 2 we established a method to identify ‘true’ events (i.e. when an event should appear in the EEG data). We did this by inserting 1,000 events into Neuroscan data using a custom-built event-marking system, the ‘Airmarker’, which marks events by triggering voltage spikes in two EEG channels. We used the lag between Airmarker events and events generated by Neuroscan as a reference for comparisons in Experiment 3. In Experiment 3 we measured the precision and accuracy of three types of Emotiv event-marking by generating 1,000 events, 1 s apart. We measured precision as the variability (standard deviation in ms) of Emotiv events and accuracy as the mean difference between Emotiv events and true events. The three triggering methods we tested were: (1) Parallel-port-generated TTL triggers; (2) Arduino-generated TTL triggers; and (3) Serial-port triggers. In Methods 1 and 2 we used an auxiliary device, Emotiv Extender, to incorporate triggers into the EEG data. We tested these event-marking methods across three configurations of Emotiv EEG systems: (1) Emotiv EPOC+ sampling at 128 Hz; (2) Emotiv EPOC+ sampling at 256 Hz; and (3) Emotiv EPOC Flex sampling at 128 Hz.ResultsIn Experiment 1 we found that the smaller P1 and N1 peaks were attenuated at lower levels of jitter relative to the larger P2 peak (21 ms, 16 ms, and 45 ms for P1, N1, and P2, respectively). In Experiment 2, we found an average lag of 30.96 ms for Airmarker events relative to Neuroscan events. In Experiment 3, we found some lag in all configurations. However, all configurations exhibited precision of less than a single sample, with serial-port-marking the most precise when paired with EPOC+ sampling at 256 Hz.ConclusionAll Emotiv event-marking methods and configurations that we tested were precise enough for ERP research as the precision of each method would provide ERP waveforms statistically equivalent to a research-standard system. Though all systems exhibited some level of inaccuracy, researchers could easily account for these during data processing. creator: Nikolas S. Williams creator: Genevieve M. McArthur creator: Nicholas A. Badcock uri: https://doi.org/10.7717/peerj.10700 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Williams et al. title: Effect of vitamin D supplementation in patients with chronic hepatitis C after direct-acting antiviral treatment: a randomized, double-blind, placebo-controlled trial link: https://peerj.com/articles/10709 last-modified: 2021-02-09 description: BackgroundReplacement of vitamin D (VD) among patients with chronic hepatitis C (CHC) before viral eradication has demonstrated a protective effect on serum markers associated with hepatic fibrogenesis. We therefore hypothesized that VD may facilitate further fibrosis amelioration following curative treatment with direct-acting antivirals (DAA).MethodsThis study was a randomized, double-blind, placebo-controlled trial conducted between February 2018 and August 2018. Patients with CHC and VD deficiency were randomized in a 1:1 ratio to either receive ergicalciferol or placebo over 6 weeks. Biochemical analysis indicators, including 25-hydroxyvitamin D (25(OH)D), fibrogenic markers [(transforming growth factor beta 1 (TGF-β1) and tissue inhibitors of matrix metalloproteinases 1 (TIMP-1)], and fibrolytic markers [matrix metalloproteinase 9 (MMP-9) and amino terminal type III procollagen peptide (P3NP)], were assessed at baseline and at 6 weeks. Serum 25(OH)D was analyzed by a chemiluminescence immunoassay. Serum hepatic fibrogenesis markers were measured using a quantitative sandwich enzyme-linked immunosorbent assay.ResultsSeventy-five patients with CHC and VD deficiency were randomly assigned to VD (n = 37) and placebo (n = 38) groups. At the end of the study, the mean serum 25(OH)D level had risen to a normal level in the VD group, but was still deficient in the placebo group (41.8 ±   9.1 vs. 18.1 ±  4.6 ng/mL, p < 0.001). Upon restoration of the VD level, there were no significant mean differences in the change from baseline for TGF-β1 (−0.6 ng/mL (95% confidence interval (95% CI) [−2.8–1.7]), p = 0.63), TIMP-1 (−5.5 ng/mL (95% CI [−26.4 –15.3]), p = 0.60), MMP-9 (122.9 ng/mL (95% CI [−69.0 –314.8]), p = 0.21), and P3NP (−0.1 ng/mL (95% CI [−2.4 –2.2]), p = 0.92) between the VD and placebo groups.ConclusionShort-term VD supplementation after DAA treatment in patients with CHC does not improve serum fibrogenesis markers and may not expedite the residual liver fibrosis healing process. Future studies are warranted to evaluate the long-term effect of VD supplementation on hepatic fibrosis regression. creator: Supachaya Sriphoosanaphan creator: Kessarin Thanapirom creator: Stephen J. Kerr creator: Sirinporn Suksawatamnuay creator: Panarat Thaimai creator: Sukanya Sittisomwong creator: Kanokwan Sonsiri creator: Nunthiya Srisoonthorn creator: Nicha Teeratorn creator: Natthaporn Tanpowpong creator: Bundit Chaopathomkul creator: Sombat Treeprasertsuk creator: Yong Poovorawan creator: Piyawat Komolmit uri: https://doi.org/10.7717/peerj.10709 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Sriphoosanaphan et al. title: Synopsis of Ecuadorian Pterichis (Orchidaceae) link: https://peerj.com/articles/10807 last-modified: 2021-02-09 description: A taxonomic synopsis of the orchid genus Pterichis in Ecuador is presented. All national representatives of this genus are characterized and their floral segments are illustrated. Four new species and two new varieties are described. An updated key to Ecuadorian Pterichis is provided. Plants of the genus are growing mostly as terrestrial herbs at the altitude of 2,300–4,110 m. Often two or more species co-occur in the area of 25 × 25 km. Their occurrence was reported from three ecoregions—the Eastern Cordillera real montane forests, the Northern Andean páramo and the Northwestern Andean montane forests. Seven Ecuadorian Pterichis are endemic. creator: Marta Kolanowska creator: Dariusz L. Szlachetko creator: Sławomir Nowak uri: https://doi.org/10.7717/peerj.10807 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Kolanowska et al. title: modelBuildR: an R package for model building and feature selection with erroneous classifications link: https://peerj.com/articles/10849 last-modified: 2021-02-09 description: BackgroundModel building is a crucial part of omics based biomedical research to transfer classifications and obtain insights into underlying mechanisms. Feature selection is often based on minimizing error between model predictions and given classification (maximizing accuracy). Human ratings/classifications, however, might be error prone, with discordance rates between experts of 5–15%. We therefore evaluate if a feature pre-filtering step might improve identification of features associated with true underlying groups.MethodsData was simulated for up to 100 samples and up to 10,000 features, 10% of which were associated with the ground truth comprising 2–10 normally distributed populations. Binary and semi-quantitative ratings with varying error probabilities were used as classification. For feature preselection standard cross-validation (V2) was compared to a novel heuristic (V1) applying univariate testing, multiplicity adjustment and cross-validation on switched dependent (classification) and independent (features) variables. Preselected features were used to train logistic regression/linear models (backward selection, AIC). Predictions were compared against the ground truth (ROC, multiclass-ROC). As use case, multiple feature selection/classification methods were benchmarked against the novel heuristic to identify prognostically different G-CIMP negative glioblastoma tumors from the TCGA-GBM 450 k methylation array data cohort, starting from a fuzzy umap based rough and erroneous separation.ResultsV1 yielded higher median AUC ranks for two true groups (ground truth), with smaller differences for true graduated differences (3–10 groups). Lower fractions of models were successfully fit with V1. Median AUCs for binary classification and two true groups were 0.91 (range: 0.54–1.00) for V1 (Benjamini-Hochberg) and 0.70 (0.28–1.00) for V2, 13% (n = 616) of V2 models showed AUCs < = 50% for 25 samples and 100 features. For larger numbers of features and samples, median AUCs were 0.75 (range 0.59–1.00) for V1 and 0.54 (range 0.32–0.75) for V2. In the TCGA-GBM data, modelBuildR allowed best prognostic separation of patients with highest median overall survival difference (7.51 months) followed a difference of 6.04 months for a random forest based method.ConclusionsThe proposed heuristic is beneficial for the retrieval of features associated with two true groups classified with errors. We provide the R package modelBuildR to simplify (comparative) evaluation/application of the proposed heuristic (http://github.com/mknoll/modelBuildR). creator: Maximilian Knoll creator: Jennifer Furkel creator: Juergen Debus creator: Amir Abdollahi uri: https://doi.org/10.7717/peerj.10849 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Knoll et al. title: Detection and monitoring of insect traces in bioaerosols link: https://peerj.com/articles/10862 last-modified: 2021-02-09 description: Studies on bioaerosols have primarily focused on their chemical and biological compositions and their impact on public health and the ecosystem. However, most bioaerosol studies have only focused on viruses, bacteria, fungi, and pollen. To assess the diversity and composition of airborne insect material in particulate matter (PM) for the first time, we attempted to detect DNA traces of insect origin in dust samples collected over a two-year period. These samples were systematically collected at one-month intervals and categorized into two groups, PM2.5 and PM10, based on the aerodynamic diameter of the aerosol particles. Cytochrome-c oxidase I (COI) was the barcoding region used to identify the origins of the extracted DNA. The airborne insect community in these samples was analyzed using the Illumina MiSeq platform. The most abundant insect sequences belonged to the order Hemiptera (true bugs), whereas order Diptera were also detected in both PM2.5 and PM10 samples. Additionally, we inferred the presence of particulates of insect origin, such as brochosomes and integument particles, using scanning electron microscopy (SEM). This provided additional confirmation of the molecular results. In this study, we demonstrated the benefits of detection and monitoring of insect information in bioaerosols for understanding the source and composition. Our results suggest that the PM2.5 and PM10 groups are rich in insect diversity. Lastly, the development of databases can improve the identification accuracy of the analytical results. creator: Panyapon Pumkaeo creator: Junko Takahashi creator: Hitoshi Iwahashi uri: https://doi.org/10.7717/peerj.10862 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Pumkaeo et al. title: Osmotic stress in banana is relieved by exogenous nitric oxide link: https://peerj.com/articles/10879 last-modified: 2021-02-09 description: Drought is one of the severe environmental stresses threatening agriculture around the globe. Nitric oxide plays diverse roles in plant growth and defensive responses. Despite a few studies supporting the role of nitric oxide in plants under drought responses, little is known about its pivotal molecular amendment in the regulation of stress signaling. In this study, a label-free nano-liquid chromatography-mass spectrometry approach was used to determine the effects of sodium nitroprusside (SNP) on polyethylene glycol (PEG)-induced osmotic stress in banana roots. Plant treatment with SNP improved plant growth and reduced the percentage of yellow leaves. A total of 30 and 90 proteins were differentially identified in PEG+SNP against PEG and PEG+SNP against the control, respectively. The majority of proteins differing between them were related to carbohydrate and energy metabolisms. Antioxidant enzyme activities, such as superoxide dismutase and ascorbate peroxidase, decreased in SNP-treated banana roots compared to PEG-treated banana. These results suggest that the nitric oxide-induced osmotic stress tolerance could be associated with improved carbohydrate and energy metabolism capability in higher plants. creator: Muhammad Asyraf Mohd Amnan creator: Teen-Lee Pua creator: Su-Ee Lau creator: Boon Chin Tan creator: Hisateru Yamaguchi creator: Keisuke Hitachi creator: Kunihiro Tsuchida creator: Setsuko Komatsu uri: https://doi.org/10.7717/peerj.10879 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Mohd Amnan et al. title: Validity of the Polar Vantage M watch when measuring heart rate at different exercise intensities link: https://peerj.com/articles/10893 last-modified: 2021-02-09 description: BackgroundThe use of wrist worn wearable fitness trackers has been growing rapidly over the last decade. The growing popularity can be partly attributed to the improvements in technology, making activity trackers more affordable, comfortable and convenient for use in different fitness and environmental applications. Fitness trackers typically monitor activity level, track steps, distance, heart rate (HR), sleep, peripheral capillary oxygen saturation and more, as the technology continuously is advancing. In terms of measuring HR, photoplethysmography (PPG) is a relatively new technology utilised in wearables. PPG estimates HR through an optical technique that monitors changes in blood volume beneath the skin. With these new products becoming available it is important that the validity of these devices be evaluated. Therefore, the aim of this study was to assess the validity of the Polar Vantage M (PVM) watch to measure HR compared to medical grade ECG on a healthy population during a range of treadmill exercise intensities.MethodsA total of 30 healthy participants (n = 17 males, n = 13 females) were recruited for this study. The validity of the PVM watch to measure HR was compared against the gold standard 5-lead ECG. The study was conducted on 2 separate testing days with 24–48 h between sessions. Participants completed the Bruce Treadmill Protocol, and HR was measured every 30 s. Validation of the PVM watch in comparison to the ECG was measured with an Intraclass Correlation Coefficient (ICC) and associated 95% confidence intervals (CI) and levels of agreement were identified with Bland–Altman plots with 90% limits of agreement. Linear regression analysis was performed to calculate the value of r2 computing the variation of HR obtained by the PVM watch and ECG.ResultsIn total, 30 participants completed the protocol, with data from 28 participants utilised for statistical analysis (16 males, 14 females, 26.10 ± 3.39 years, height 52.36 m ± 7.40 cm, mass 73.59 ± 11.90 kg). A strong and significant correlation was found between the PVM watch and ECG, demonstrating good criterion validity (p < 0.05, r2 = 0.87). Good validity was seen for day 1 and day 2 for stage 0 (ICC = 0.83; 95% CI [0.63–0.92], ICC = 0.74; 95% CI [0.37–0.88]), stage 1 (ICC = 0.78; 95% CI [0.52–0.90], ICC = 0.88; 95% CI [0.74–0.95]), and stage 2 (ICC = 0.88; 95% CI [0.73–0.94], ICC = 0.80; 95% CI [0.40–0.92]). Poor validity was demonstrated on day 1 and day 2 for stages 3–5 (ICC < 0.50).ConclusionThis study revealed that the PVM watch had a strong correlation with the ECG throughout the entire Bruce Protocol, however the level of agreement (LoA) becomes widely dispersed as exercise intensities increased. Due to the large LoA between the ECG and PVM watch, it is not advisable to use this device in clinical populations in which accurate HR measures are essential for patient safety; however, the watch maybe used in settings where less accurate HR is not critical to an individual’s safety while exercising. creator: Tricia Shumate creator: Magdalen Link creator: James Furness creator: Kevin Kemp-Smith creator: Vini Simas creator: Mike Climstein uri: https://doi.org/10.7717/peerj.10893 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Shumate et al. title: Incomplete lineage sorting and ancient admixture, and speciation without morphological change in ghost-worm cryptic species link: https://peerj.com/articles/10896 last-modified: 2021-02-09 description: Morphologically similar species, that is cryptic species, may be similar or quasi-similar owing to the deceleration of morphological evolution and stasis. While the factors underlying the deceleration of morphological evolution or stasis in cryptic species remain unknown, decades of research in the field of paleontology on punctuated equilibrium have originated clear hypotheses. Species are expected to remain morphologically identical in scenarios of shared genetic variation, such as hybridization and incomplete lineage sorting, or in scenarios where bottlenecks reduce genetic variation and constrain the evolution of morphology. Here, focusing on three morphologically similar Stygocapitella species, we employ a whole-genome amplification method (WGA) coupled with double-digestion restriction-site associated DNA sequencing (ddRAD) to reconstruct the evolutionary history of the species complex. We explore population structure, use population-level statistics to determine the degree of connectivity between populations and species, and determine the most likely demographic scenarios which generally reject for recent hybridization. We find that the combination of WGA and ddRAD allowed us to obtain genomic-level data from microscopic eukaryotes (∼1 millimetre) opening up opportunities for those working with population genomics and phylogenomics in such taxa. The three species share genetic variance, likely from incomplete lineage sorting and ancient admixture. We speculate that the degree of shared variation might underlie morphological similarity in the Atlantic species complex. creator: José Cerca creator: Angel G. Rivera-Colón creator: Mafalda S. Ferreira creator: Mark Ravinet creator: Michael D. Nowak creator: Julian M. Catchen creator: Torsten H. Struck uri: https://doi.org/10.7717/peerj.10896 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Cerca et al. title: Is there a link between pre-existing antibodies acquired due to childhood vaccinations or past infections and COVID-19? A case control study link: https://peerj.com/articles/10910 last-modified: 2021-02-09 description: BackgroundThere is growing evidence indicating that children are less affected from COVID-19. Some authors speculate that childhood vaccinations may provide some cross-protection against COVID-19. In this study, our aim was to compare the circulating antibody titers for multiple childhood vaccine antigens, as an indicator of the state of immune memory between patients with COVID-19 and healthy controls, with a specific aim to identify the association between disease severity and antibody titrations which may indicate a protective function related to vaccine or disease induced memory.MethodsThis study is a case-control study including 53 patients with COVID-19 and 40 healthy volunteers. COVID-19 severity was divided into three groups: asymptomatic, mild and severe. We measured the same set of antibody titers for vaccine antigens, and a set of biochemical and infection markers, in both the case and control groups.ResultsRubella (p = 0.003), pneumococcus (p = 0.002), and Bordetella pertussis (p < 0.0001) titers were found to be significantly lower in the case group than the control group. There was a significant decline in pneumococcus titers with severity of disease (p = 0.021) and a significant association with disease severity for Bordetella pertussis titers (p = 0.014) among COVID patients. Levels of AST, procalcitonin, ferritin and D-dimer significantly increased with the disease severity.DiscussionOur study supports the hypothesis that pre-existing immune memory, as monitored using circulating antibodies, acquired from childhood vaccinations, or past infections confer some protection against COVID-19. Randomized controlled studies are needed to support a definitive conclusion. creator: Bilge Sumbul creator: Hilmi Erdem Sumbul creator: Ramazan Azim Okyay creator: Erdinç Gülümsek creator: Ahmet Rıza Şahin creator: Baris Boral creator: Burhan Fatih Koçyiğit creator: Mostafa Alfishawy creator: Jeffrey Gold creator: ALİ Muhittin Tasdogan uri: https://doi.org/10.7717/peerj.10910 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Sumbul et al. title: Rapid coral reef assessment using 3D modelling and acoustics: acoustic indices correlate to fish abundance, diversity and environmental indicators in West Papua, Indonesia link: https://peerj.com/articles/10761 last-modified: 2021-02-08 description: BackgroundProviding coral reef systems with the greatest chance of survival requires effective assessment and monitoring to guide management at a range of scales from community to government. The development of rapid monitoring approaches amenable to collection at community level, yet recognised by policymakers, remains a challenge. Technologies can increase the scope of data collection. Two promising visual and audio approaches are (i) 3D habitat models, generated through photogrammetry from video footage, providing assessment of coral cover structural metrics and (ii) audio, from which acoustic indices shown to correlate to vertebrate and invertebrate diversity, can be extracted.MethodsWe collected audio and video imagery using low cost underwater cameras (GoPro Hero7™) from 34 reef samples from West Papua (Indonesia). Using photogrammetry one camera was used to generate 3D models of 4 m2 reef, the other was used to estimate fish abundance and collect audio to generate acoustic indices. We investigated relationships between acoustic metrics, fish abundance/diversity/functional groups, live coral cover and reef structural metrics.ResultsGeneralized linear modelling identified significant but weak correlations between live coral cover and structural metrics extracted from 3D models and stronger relationships between live coral and fish abundance. Acoustic indices correlated to fish abundance, species richness and reef functional metrics associated with overfishing and algal control. Acoustic Evenness (1,200–11,000 Hz) and Root Mean Square RMS (100–1,200 Hz) were the best individual predictors overall suggesting traditional bioacoustic indices, providing information on sound energy and the variability in sound levels in specific frequency bands, can contribute to reef assessment.ConclusionAcoustics and 3D modelling contribute to low-cost, rapid reef assessment tools, amenable to community-level data collection, and generate information for coral reef management. Future work should explore whether 3D models of standardised transects and acoustic indices generated from low cost underwater cameras can replicate or support ‘gold standard’ reef assessment methodologies recognised by policy makers in marine management. creator: Mika Peck creator: Ricardo F. Tapilatu creator: Eveline Kurniati creator: Christopher Rosado uri: https://doi.org/10.7717/peerj.10761 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Peck et al. title: Identification of SNPs associated with magnesium and sodium uptake and the effect of their accumulation on micro and macro nutrient levels in Vitis vinifera link: https://peerj.com/articles/10773 last-modified: 2021-02-08 description: Macro and micro nutrient accumulation affects all stages of plant growth and development. When nutrient deficiencies or excesses occur, normal plant growth is altered resulting in symptoms such as leaf chlorosis, plant stunting or death. In grapes, few genomic regions associated with nutrient accumulation or deficiencies have been identified. Our study evaluated micro and macro nutrient concentrations in Vitis vinifera L. to identify associated SNPs using an association approach with genotype by sequencing data. Nutrient concentrations and foliar symptoms (leaf chlorosis and stunting) were compared among 249 F1Vitis vinifera individuals in 2015 and 2016. Foliar symptoms were consistent (≥90%) between years and correlated with changes in nutrient concentrations of magnesium (r = 0.65 and r = 0.38 in 2015 and 2016, respectively), aluminum (r = 0.24 and r = 0.49), iron (r = 0.21 and r = 0.49), and sodium (r = 0.32 and r = 0.21). Single nucleotide polymorphisms associated with symptoms, sodium, and magnesium were detected on each chromosome with the exception of 5, 7 and 17 depending on the trait and genome used for analyses explaining up to 40% of the observed variation. Symptoms and magnesium concentration were primarily associated with SNPs on chromosome 3, while SNPs associated with increased sodium content were primarily found on chromosomes 11 and 18. Mean concentrations for each nutrient varied between years in the population between symptomatic and asymptomatic plants, but relative relationships were mostly consistent. These data suggest a complex relationship among foliar symptoms and micro and macro nutrients accumulating in grapevines. creator: Rachel P. Naegele creator: Jason P. Londo creator: Cheng Zou creator: Peter Cousins uri: https://doi.org/10.7717/peerj.10773 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Naegele et al. title: Abnormal expression of HOXD11 promotes the malignant behavior of glioma cells and leads to poor prognosis of glioma patients link: https://peerj.com/articles/10820 last-modified: 2021-02-08 description: BackgroundHomeobox D11 (HOXD11) plays an important role in a variety of cancers, but its precise role in gliomas remains unclear. This study aimed to explore the relationship between HOXD11 and gliomas by combining bioinformatics methods with basic experimental validation.Materials and methodsObtain gene expression information and clinical information of glioma and non-tumor brain tissue samples from multiple public databases such as TCGA (666 glioma samples), CGGA (749 glioma samples), GEPIA(163 glioblastoma samples and 207 normal control samples), GEO (GSE4290 and GSE15824). Nine cases of glioma tissue and five cases of normal control brain tissue were collected from the clinical department of Henan Provincial People’s Hospital for further verification. A series of bioinformatic analysis methods were used to confirm the relationship between HOXD11 expression and overall survival and clinical molecular characteristics of patients with glioma. RT-qPCR was used to verify the change of expression level of HOXD11 in glioma cells and tissues. MTT assay, colony formation assay, wound-healing assay, immunofluorescence staining, flow cytometry and western blotting were used to detect the effect of HOXD11 on the biological behavior of glioma cell line U251.ResultsThe high expression of HOXD11 was significantly related to age, World Health Organization (WHO) grade, chemotherapy status, histological type, and even 1p19q codeletion data and isocitrate dehydrogenase (IDH) mutation. HOXD11, as an independent risk factor, reduces the overall survival of glioma patients and has diagnostic value for the prognosis of glioma. Gene Set Enrichment Analysis (GSEA) showed that HOXD11 was significantly enriched in cell signaling pathway such as cell cycle, DNA replication and so on. Finally, we confirmed that the knockout of HOXD11 can inhibit the proliferation and invasion of U251 glioma cells, and change the biological behavior of tumor cells by preventing the progression of cell cycle.ConclusionsHOXD11 may be used as a candidate biomarker for the clinical application of targeted drug and prognostic assessment treatment of glioma. In addition, This study will help to explore the pathological mechanism of glioma. creator: Jialin Wang creator: Zhendong Liu creator: Cheng Zhang creator: Hongbo Wang creator: Ang Li creator: Binfeng Liu creator: Xiaoyu Lian creator: Zhishuai Ren creator: Wang Zhang creator: Yanbiao Wang creator: Bo Zhang creator: Bo Pang creator: Yanzheng Gao uri: https://doi.org/10.7717/peerj.10820 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: Identification of a metabolic-related gene signature predicting the overall survival for patients with stomach adenocarcinoma link: https://peerj.com/articles/10908 last-modified: 2021-02-08 description: BackgroundThe reprogramming of energy metabolism and consistently altered metabolic genes are new features of cancer, and their prognostic roles remain to be further studied in stomach adenocarcinoma (STAD).MethodsMessenger RNA (mRNA) expression profiles and clinicopathological data were downloaded from The Cancer Genome Atlas (TCGA) and the GSE84437 databases from the Gene Expression Omnibus (GEO) database. A univariate Cox regression analysis and the least absolute shrinkage and selection operator (LASSO) Cox regression model established a novel metabolic signature based on TCGA. The area under the receiver operating characteristic (ROC) curve (AUROC) and a nomogram were calculated to assess the predictive accuracy.ResultsA novel metabolic-related signature (including acylphosphatase 1, RNA polymerase I subunit A, retinol dehydrogenase 12, 5-oxoprolinase, ATP-hydrolyzing, malic enzyme 1, nicotinamide N-methyltransferase, gamma-glutamyl transferase 5, deoxycytidine kinase, galactosidase alpha, DNA polymerase delta 3, glutathione S-transferase alpha 2, N-acyl sphingosine amidohydrolase 1, and N-acyl sphingosine amidohydrolase 1) was identified. In both TCGA and GSE84437, patients in the high-risk group showed significantly poorersurvival than the patients in the low-risk group. A good predictive value was shown by the AUROC and nomogram. Furthermore, gene set enrichment analyses (GSEAs) revealed several significantly enriched pathways, which may help in explaining the underlying mechanisms.ConclusionsA novel robust metabolic-related signature for STAD prognosis prediction was conducted. The signature may reflect the dysregulated metabolic microenvironment and can provided potential biomarkers for metabolic therapy in STAD. creator: Yuan Nie creator: Linxiang Liu creator: Qi Liu creator: Xuan Zhu uri: https://doi.org/10.7717/peerj.10908 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Nie et al. title: Stage 1 registered report: spatiotemporal patterns of the COVID-19 epidemic in Mexico at the municipality level link: https://peerj.com/articles/10622 last-modified: 2021-02-05 description: In this stage 1 registered report, we propose an analysis of the spatio-temporal patterns of the COVID-19 epidemic in Mexico using the georeferenced confirmed cases aggregated at the municipality level. We will compute weekly Moran index to assess spatial autocorrelation over time and identify clusters of the disease using the “flexibly shaped spatial scan” approach. Finally, different distance models will be compared to select the best suited to predict inter-municipality contagion. This study will help us understand the spread of the epidemic over the Mexican territory and give insights to model and predict the epidemic behavior. creator: Jean-François Mas uri: https://doi.org/10.7717/peerj.10622 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Mas title: The relationship between (sub)tropical climates and the incidence of COVID-19 link: https://peerj.com/articles/10655 last-modified: 2021-02-05 description: This work explores (non)linear associations between relative humidity and temperature and the incidence of COVID-19 among 27 Brazilian state capital cities in (sub)tropical climates, measured daily from summer through winter. Previous works analyses have shown that SARS-CoV-2, the virus that causes COVID-19, finds stability by striking a certain balance between relative humidity and temperature, which indicates the possibility of surface contact transmission. The question remains whether seasonal changes associated with climatic fluctuations might actively influence virus survival. Correlations between climatic variables and infectivity rates of SARS-CoV-2 were applied by the use of a Generalized Additive Model (GAM) and the Locally Estimated Scatterplot Smoothing LOESS nonparametric model. Tropical climates allow for more frequent outdoor human interaction, making such areas ideal for studies on the natural transmission of the virus. Outcomes revealed an inverse relationship between subtropical and tropical climates for the spread of the novel coronavirus and temperature, suggesting a sensitivity behavior to climates zones. Each 1 °C rise of the daily temperature mean correlated with a −11.76% (t = −5.71, p < 0.0001) decrease and a 5.66% (t = 5.68, p < 0.0001) increase in the incidence of COVID-19 for subtropical and tropical climates, respectively. creator: David Prata creator: Waldecy Rodrigues creator: Paulo Henrique De Souza Bermejo creator: Marina Moreira creator: Wainesten Camargo creator: Marcelo Lisboa creator: Geovane Rossone Reis creator: Humberto Xavier de Araujo uri: https://doi.org/10.7717/peerj.10655 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Prata et al. title: Serum BDNF levels and the antidepressant effects of electroconvulsive therapy with ketamine anaesthesia: a preliminary study link: https://peerj.com/articles/10699 last-modified: 2021-02-05 description: ObjectiveTo firstly examine the relationship between serum brain-derived neurotrophic factor (BDNF) levels and antidepressant response to ketamine as an anaesthesia in electroconvulsive therapy (ECT) in Chinese patients with treatment-refractory depression (TRD).MethodsThirty patients with TRD were enrolled and underwent eight ECT sessions with ketamine anaesthesia (0.8 mg/kg) alone. Depression severity, response and remission were evaluated using the 17-item Hamilton Depression Rating Scale (HAMD-17). Enzyme-linked immunosorbent assay (ELISA) was applied to examine serum BDNF levels in patients with TRD at baseline and after the second, fourth and eighth ECT sessions. Baseline serum samples were also collected for 30 healthy controls.ResultsNo significant differences were observed in serum BDNF levels between patients with TRD and healthy controls at baseline (p > 0.05). The remission rate was 76.7% (23/30) after the last ECT treatment, although all patients with TRD obtained antidepressant response criteria. Serum BDNF levels were not altered compared to baseline, even between remitters and nonremitters (all p > 0.05), despite the significant reduction in HAMD-17 and Brief Psychiatric Rating Scale (BPRS) scores after ECT with ketamine anaesthesia (all p < 0.05). The antidepressant effects of ECT with ketamine anaesthesia were not correlated with changes in serum BDNF levels (all p > 0.05).ConclusionThis preliminary study indicated that serum BDNF levels do not appear to be a reliable biomarker to determine the antidepressant effects of ketamine as an anaesthesia in ECT for patients with TRD. Further studies with larger sample sizes are warranted to confirm these findings. creator: Wei Zheng creator: Qiaomei Cen creator: Sha Nie creator: Minyi Li creator: Rong Zeng creator: Sumiao Zhou creator: Dongbin Cai creator: Miaoling Jiang creator: Xiong Huang uri: https://doi.org/10.7717/peerj.10699 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Zheng et al. title: Quantifying upper limb motor impairment in people with Parkinson’s disease: a physiological profiling approach link: https://peerj.com/articles/10735 last-modified: 2021-02-05 description: BackgroundUpper limb motor impairments, such as slowness of movement and difficulties executing sequential tasks, are common in people with Parkinson’s disease (PD).ObjectiveTo evaluate the validity of the upper limb Physiological Profile Assessment (PPA) as a standard clinical assessment battery in people with PD, by determining whether the tests, which encompass muscle strength, dexterity, arm stability, position sense, skin sensation and bimanual coordination can (a) distinguish people with PD from healthy controls, (b) detect differences in upper limb test domains between “off” and “on” anti-Parkinson medication states and (c) correlate with a validated measure of upper limb function.MethodsThirty-four participants with PD and 68 healthy controls completed the upper limb PPA tests within a single session.ResultsPeople with PD exhibited impaired performance across most test domains. Based on validity, reliability and feasibility, six tests (handgrip strength, finger-press reaction time, 9-hole peg test, bimanual pole test, arm stability, and shirt buttoning) were identified as key tests for the assessment of upper limb function in people with PD.ConclusionsThe upper limb PPA provides a valid, quick and simple means of quantifying specific upper limb impairments in people with PD. These findings indicate clinical assessments should prioritise tests of muscle strength, unilateral movement and dexterity, bimanual coordination, arm stability and functional tasks in people with PD as these domains are the most commonly and significantly impaired. creator: Lewis A. Ingram creator: Vincent K. Carroll creator: Annie A. Butler creator: Matthew A. Brodie creator: Simon C. Gandevia creator: Stephen R. Lord uri: https://doi.org/10.7717/peerj.10735 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Ingram et al. title: Syncmers are more sensitive than minimizers for selecting conserved k‑mers in biological sequences link: https://peerj.com/articles/10805 last-modified: 2021-02-05 description: Minimizers are widely used to select subsets of fixed-length substrings (k-mers) from biological sequences in applications ranging from read mapping to taxonomy prediction and indexing of large datasets. The minimizer of a string of w consecutive k-mers is the k-mer with smallest value according to an ordering of all k-mers. Syncmers are defined here as a family of alternative methods which select k-mers by inspecting the position of the smallest-valued substring of length s < k within the k-mer. For example, a closed syncmer is selected if its smallest s-mer is at the start or end of the k-mer. At least one closed syncmer must be found in every window of length (k − s) k-mers. Unlike a minimizer, a syncmer is identified by its sequence alone, and is therefore synchronized in the following sense: if a given k-mer is selected from one sequence, it will also be selected from any other sequence. Also, minimizers can be deleted by mutations in flanking sequence, which cannot happen with syncmers. Experiments on minimizers with parameters used in the minimap2 read mapper and Kraken taxonomy prediction algorithm respectively show that syncmers can simultaneously achieve both lower density and higher conservation compared to minimizers. creator: Robert Edgar uri: https://doi.org/10.7717/peerj.10805 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Edgar title: Assessing red deer hunting management in the Iberian Peninsula: the importance of longitudinal studies link: https://peerj.com/articles/10872 last-modified: 2021-02-05 description: Understanding the dynamics of a wildlife population in relation to hunting strategies is essential to achieve sustainable management. We used monitoring data over 25 years from two red deer (Cervus elaphus) populations with different management (with and without supplemental feeding) in South Central Spain to: (i) characterise the density dependence of population dynamics under contrasted management, and (ii) provide the basis for sustainable extraction by considering the theoretical maximum sustainable yield (MSYt) as the reference. The red deer population displayed a typical management reactive culling approach (‘saw-tooth-like’ curves), with occasional strong annual harvests but not occurring on a regular basis. Interestingly, we found reduced population growth at high densities in both populations, indicating that density-mediated factors determined population growth even when artificial feeding was provided. However, no effects of sex not age class of the extracted population on the population growth rate were determined. The total number of animals hunted was only slightly above those predicted by MSYt (i.e. K50%) in both populations, despite high densities close to theoretical K, being consistent throughout the study period. The extraction rates (30.3 and 34.0%, for supplemented and unsupplemented populations, respectively) were 13.3% and 10.2% lower compared to the MSYt situation in the unsupplemented and supplemented populations, respectively. Long term population monitoring data provided feasible and suitable baseline values to optimise the sustainable exploitation of red deer populations in the Mediterranean ecosystem under these contrasting management scenarios. Adaptive management, involving objective-driven decision making informed by data on red deer population dynamic, can contribute (i) to maximising the total extraction over the long term while (ii) reducing the ecological impact of high population densities. creator: Antonio José Carpio Camargo creator: Jose Barasona creator: Pelayo Acevedo creator: Yolanda Fierro creator: Christian Gortazar creator: Carlos Vigal creator: Ángel Moreno creator: Joaquin Vicente uri: https://doi.org/10.7717/peerj.10872 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Carpio Camargo et al. title: Jasmonic acid biosynthesis by fungi: derivatives, first evidence on biochemical pathways and culture conditions for production link: https://peerj.com/articles/10873 last-modified: 2021-02-05 description: Jasmonic acid (JA) and its derivatives called jasmonates (JAs) are lipid-derived signalling molecules that are produced by plants and certain fungi. Beside this function, JAs have a great variety of applications in flavours and fragrances production. In addition, they may have a high potential in agriculture. JAs protect plants against infections. Although there is much information on the biosynthesis and function of JA concerning plants, knowledge on these aspects is still scarce for fungi. Taking into account the practical importance of JAs, the objective of this review is to summarize knowledge on the occurrence of JAs from fungal culture media, their biosynthetic pathways and the culture conditions for optimal JA production as an alternative source for the production of these valuable metabolites. creator: Felipe Eng creator: Jorge Erick Marin creator: Krzysztof Zienkiewicz creator: Mariano Gutiérrez-Rojas creator: Ernesto Favela-Torres creator: Ivo Feussner uri: https://doi.org/10.7717/peerj.10873 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Eng et al. title: What are the limits on whale ear bone size? Non-isometric scaling of the cetacean bulla link: https://peerj.com/articles/10882 last-modified: 2021-02-05 description: The history of cetaceans demonstrates dramatic macroevolutionary changes that have aided their transformation from terrestrial to obligate aquatic mammals. Their fossil record shows extensive anatomical modifications that facilitate life in a marine environment. To better understand the constraints on this transition, we examined the physical dimensions of the bony auditory complex, in relation to body size, for both living and extinct cetaceans. We compared the dimensions of the tympanic bulla, a conch-shaped ear bone unique to cetaceans, with bizygomatic width—a proxy for cetacean body size. Our results demonstrate that cetacean ears scale non-isometrically with body size, with about 70% of variation explained by increases in bizygomatic width. Our results, which encompass the breadth of the whale fossil record, size diversity, and taxonomic distribution, suggest that functional auditory capacity is constrained by congruent factors related to cranial morphology, as opposed to allometrically scaling with body size. creator: Sabrina L. Groves creator: Carlos Mauricio Peredo creator: Nicholas D. Pyenson uri: https://doi.org/10.7717/peerj.10882 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Groves et al. title: Genetic diversity and structure of the narrow endemic Seseli farrenyi (Apiaceae): implications for translocation link: https://peerj.com/articles/10521 last-modified: 2021-02-04 description: Seseli farrenyi (Apiaceae) is an extremely narrow endemic plant, which is considered as one of the species of most conservation concern in Catalonia (NW Mediterranean Basin). Given the accelerated fragmentation and reduction of population size (of over 90%), the environmental agency of Catalonia is currently preparing a recovery plan that includes reinforcements of the extant populations. The present study is aimed at providing the necessary knowledge to carry out genetically-informed translocations, by using microsatellites as genetic markers. Fourteen microsatellites have been specifically developed for S. farrenyi, of which nine have been used. Besides the extant natural populations, the three ex situ collections that are known to exist of this species have also been studied, as they would be the donor sources for translocation activities. Our main finding is that levels of genetic diversity in the natural populations of S. farrenyi are still high (He = 0.605), most likely as a result of a predominantly outcrossing mating system in combination with the limited time elapsed since the population decline. However, population fragmentation is showing the first genetic signs, as the values of genetic differentiation are relatively high, and two well-differentiated genetic lineages have been found even in such a narrow geographic range. These genetic results provide important information when designing conservation management measures. creator: Núria Garcia-Jacas creator: Jèssica Requena creator: Sergi Massó creator: Roser Vilatersana creator: Cèsar Blanché creator: Jordi López-Pujol uri: https://doi.org/10.7717/peerj.10521 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Garcia-Jacas et al. title: The natural product biosynthetic potential of Red Sea nudibranch microbiomes link: https://peerj.com/articles/10525 last-modified: 2021-02-04 description: BackgroundAntibiotic resistance is a growing problem that can be ameliorated by the discovery of novel drug candidates. Bacterial associates are often the source of pharmaceutically active natural products isolated from marine invertebrates, and thus, important targets for drug discovery. While the microbiomes of many marine organisms have been extensively studied, microbial communities from chemically-rich nudibranchs, marine invertebrates that often possess chemical defences, are relatively unknown.MethodsWe applied both culture-dependent and independent approaches to better understand the biochemical potential of microbial communities associated with nudibranchs. Gram-positive microorganisms isolated from nudibranchs collected in the Red Sea were screened for antibacterial and antitumor activity. To assess their biochemical potential, the isolates were screened for the presence of natural product biosynthetic gene clusters, including polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS) genes, using PCR. The microbiomes of the nudibranchs were investigated by high-throughput sequencing of 16S rRNA amplicons.ResultsIn screens against five model microorganisms, 51% of extracts displayed antimicrobial activity against more than one organism, and 19% exhibited antitumor activity against Ehrlich’s ascites carcinoma. Sixty-four percent of isolates contained PKS and NRPS genes, suggesting their genomes contain gene clusters for natural product biosynthesis. Thirty-five percent were positive for more than one class of biosynthetic gene. These strains were identified as belonging to the Firmicutes and Actinobacteria phyla via 16S rRNA gene sequencing. In addition, 16S rRNA community amplicon sequencing revealed all bacterial isolates were present in the uncultured host-associated microbiome, although they were a very small percentage of the total community. Taken together, these results indicate that bacteria associated with marine nudibranchs are potentially a rich source of bioactive compounds and natural product biosynthetic genes. creator: Samar M. Abdelrahman creator: Nastassia V. Patin creator: Amro Hanora creator: Akram Aboseidah creator: Shimaa Desoky creator: Salha G. Desoky creator: Frank J. Stewart creator: Nicole B. Lopanik uri: https://doi.org/10.7717/peerj.10525 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Abdelrahman et al. title: Vitamin C supplementation reduces expression of circulating miR-451a in subjects with poorly controlled type 2 diabetes mellitus and high oxidative stress link: https://peerj.com/articles/10776 last-modified: 2021-02-04 description: BackgroundVitamin C is an essential element required for normal metabolic function. We investigated the effect of vitamin C supplementation on circulating miRNA (miR) expression in subjects with poorly controlled type 2 diabetes mellitus (T2DM). Changes in miR expression were also correlated with clinical measures of disease.MethodsPre- and post-vitamin C supplementation samples from five participants who had increased vitamin C levels, improved oxidative status and polymorphonuclear (PMN) function after receiving 1,000 mg of vitamin C daily for six weeks were screened for miRNA expression using the NanoString miRNA assay. Differences in miRNA expression identified from the miRNA screen were validated by qRT-PCR.ResultsFour miRNAs showed significantly different expression post-vitamin C supplementation relative to baseline, including the down-regulation of miR-451a (−1.72 fold change (FC), p = 0.036) and up-regulation of miR-1253 (0.62 FC, p = 0.027), miR-1290 (0.53 FC, p = 0.036) and miR-644a (0.5 FC, p = 0.042). The validation study showed only miR-451a expression was significantly different from baseline with vitamin C supplementation. MiR-451a expression was negatively correlated with vitamin C levels (r =  − 0.497, p = 0.049) but positively correlated with levels of malondialdehyde (MDA) (r = 0.584, p = 0.017), cholesterol (r = 0.564, p = 0.022) and low-density lipoproteins (LDL) (r = 0.522, p = 0.037). Bioinformatics analysis of the putative target genes of miR-451a indicated gene functions related to signaling pathways involved in cellular processes, such as the mammalian target of rapamycin (mTOR) signaling pathway.ConclusionsVitamin C supplementation altered circulating miR-451a expression. The results from this pilot study suggest that miRNAs could be used as biomarkers to indicate oxidative status in subjects with T2DM and with poor glycemic control and could lead to a novel molecular strategy to reduce oxidative stress in T2DM. creator: Laongthip Ruknarong creator: Chongchira Boonthongkaew creator: Nisa Chuangchot creator: Amonrat Jumnainsong creator: Naruemon Leelayuwat creator: Apinya Jusakul creator: Silvana Gaudieri creator: Chanvit Leelayuwat uri: https://doi.org/10.7717/peerj.10776 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Ruknarong et al. title: DNA methylome signatures as epigenetic biomarkers of hexanal associated with lung toxicity link: https://peerj.com/articles/10779 last-modified: 2021-02-04 description: BackgroundNumerous studies have investigated the relationship of environmental exposure, epigenetic effects, and human diseases. These linkages may contribute to the potential toxicity mechanisms of environmental chemicals. Here, we investigated the epigenetic pulmonary response of hexanal, a major indoor irritant, following inhalation exposure in F-344 rats.MethodsBased on DNA methylation profiling in gene promoter regions, we identified hexanal-characterized methylated sites and target genes using an unpaired t-test with a fold-change cutoff of ≥ 3.0 and a p-value < 0.05. We also conducted an integrated analysis of DNA methylation and mRNA expression data to identify core anti-correlated target genes of hexanal exposure. To further investigate the potential key biological processes and pathways of core DNA methylated target genes, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were performed.ResultsThirty-six dose-dependent methylated genes and anti-correlated target genes of DNA methylation and mRNA in lung tissue of hexanal exposed F-344 rats were identified. These genes were involved in diverse biological processes such as neuroactive ligand-receptor interaction, protein kinase cascade, and intracellular signaling cascade associated with pulmonary toxicity. These results suggest that novel DNA methylation-based epigenetic biomarkers of exposure to hexanal and elucidate the potential pulmonary toxicological mechanisms of action of hexanal. creator: Yoon Cho creator: Mi-Kyung Song creator: Jae-Chun Ryu uri: https://doi.org/10.7717/peerj.10779 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Cho et al. title: Genome-wide identification, characterisation, and evolution of ABF/AREB subfamily in nine Rosaceae species and expression analysis in mei (Prunus mume) link: https://peerj.com/articles/10785 last-modified: 2021-02-04 description: Rosaceae is an important family containing some of the highly evolved fruit and ornamental plants. Abiotic stress responses play key roles in the seasonal growth and development of plants. However, the molecular basis of stress responses remains largely unknown in Rosaceae. Abscisic acid (ABA) is a stress hormone involving abiotic stress response pathways. The ABRE-binding factor/ABA-responsive element-binding protein (ABF/AREB) is a subfamily of the basic domain/leucine zipper (bZIP) transcription factor family. It plays an important role in the ABA-mediated signaling pathway. Here, we analyzed the ABF/AREB subfamily genes in nine Rosaceae species. A total of 64 ABF/AREB genes were identified, including 18, 28, and 18 genes in the Rosoideae, Amygdaloideae, and Maloideae traditional subfamilies, respectively. The evolutionary relationship of the ABF/AREB subfamily genes was studied through the phylogenetic analysis, the gene structure and conserved motif composition, Ka/Ks values, and interspecies colinearity. These gene sets were clustered into four groups. In the Prunus ABF/AREB (PmABF) promoters, several cis-elements related to light, hormone, and abiotic stress response were predicted. PmABFs expressed in five different tissues, except PmABF5, which expressed only in buds. In the dormancy stages, PmABF1, 2, 5 and 7 showed differential expression. The expression of PmABF3, 4 and 6 was positively correlated with the ABA concentration. Except for PmABF5, all the PmABFs were sensitive to ABA. Several ABRE elements were contained in the promoters of PmABF1, 3, 6, 7. Based on the findings of our study, we speculate that PmABFs may play a role in flower bud dormancy in P. mume. creator: Xue Yong creator: Tangchun Zheng creator: Xiaokang Zhuo creator: Sagheer Ahmad creator: Lulu Li creator: Ping Li creator: Jiayao Yu creator: Jia Wang creator: Tangren Cheng creator: Qixiang Zhang uri: https://doi.org/10.7717/peerj.10785 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Yong et al. title: The recognition of gaming disorder in China: a case series of 223 patients link: https://peerj.com/articles/10827 last-modified: 2021-02-04 description: BackgroundInternet gaming disorder (IGD) was listed in the appendix of the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition (DSM-5) as a condition requiring further study in 2013, and gaming disorder (GD) was considered a mental disorder and listed in the 11th International Classification of Diseases Manual (ICD-11) in 2018. The study aims to obtain preliminary knowledge of the recognition of GD in China.MethodsA total of 223 Chinese patients who met both the ICD-11 and DSM-5 criteria for GD participated in the study, and a self-administered questionnaire was used to collect demographic information, gaming use characteristics, and previous diagnoses.ResultsThe average age of patients with GD was 20.5 years, and 71.3% were male. Most patients were diagnosed with emotion-related disorders at their first psychiatric visit: mood disorders (59.2%), bipolar affective disorder (18.4%), depressive episode (12.6%), and anxiety disorder (4.9%). Among the adolescent patients with a first diagnosis of mood disorders, 71.2% and 33.3% were diagnosed with bipolar affective disorder and personality disorders, respectively, at subsequent visits. Overall, after the first visit, the most common diagnosis was bipolar affective disorder (61.0%), followed by personality disorders (34.1%), mood disorders (17.0%), depressive episode (17.0%), and other disorders. Only three patients had Internet overuse.ConclusionThe identification rate of GD is extremely low in routine psychiatric clinical practice in China. Most patients with GD were previously misdiagnosed with emotion-related disorders. Psychiatrists should be trained to improve their ability to recognize and manage GD. creator: Tianli Shao creator: Xinxin Chen creator: Shucai Huang creator: Zhenjiang Liao creator: Shuhong Lin creator: Jing Qi creator: Yi Cai creator: Qiuping Huang creator: Hongxian Shen uri: https://doi.org/10.7717/peerj.10827 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Shao et al. title: Fungal and bacterial communities of ‘Pinot noir’ must: effects of vintage, growing region, climate, and basic must chemistry link: https://peerj.com/articles/10836 last-modified: 2021-02-04 description: BackgroundThe geographic and temporal distributions of bacterial and fungal populations are poorly understood within the same wine grape cultivar. In this work, we describe the microbial composition from ‘Pinot noir’ must with respect to vintage, growing region, climate, and must chemistry across the states of California and Oregon, USA.Materials and MethodsWe sampled ‘Pinot noir’ clone 667 clusters from 15 vineyards existing in a latitudinal gradient spanning nearly 1,200 km in California and Oregon for two vintages (2016 and 2017). Regions included five American Viticultural Areas (AVA). In order from southern California to Oregon, these AVAs were Santa Barbara, Monterey, Sonoma, Mendocino, and Willamette Valley. Uninoculated grape musts were subjected to 16S rRNA gene and ITS-1 amplicon sequencing to assess composition of microbial communities. We also measured grape maturity metrics. Finally, to describe regions by precipitation and growing degree days, we queried the Parameter-elevation Regressions on Independent Slopes Model (PRISM) spatial climate dataset.ResultsMost of the dominant bacterial taxa in must samples were in the family Enterobacteriaceae, notably the lactic acid bacteria or the acetic acid bacteria groups, but some, like the betaproteobacterial genus Massilia, belonged to groups not commonly found in grape musts. Fungal communities were dominated by Hanseniaspora uvarum (Saccharomycetaceae). We detected relationships between covariates (e.g., vintage, precipitation during the growing season, pH, titratable acidity, and total soluble solids) and bacterial genera Gluconobacter and Tatumella in the family Enterobacteraceae, Sphingomonas (Sphingomonodaceae), Lactobacillus (Lactobacillaceae), and Massilia (Oxalobacteraceae), as well as fungal genera in Hanseniaspora, Kazachstania, Lachancea, Torulaspora in the family Saccharomycetaceae, as well as Alternaria (Pleosporaceae), Erysiphe (Erysiphaceae), and Udeniomyces (Cystofilobasidiaceae). Fungal community distances were significantly correlated with geographic distances, but this was not observed for bacterial communities. Climate varied across regions and vintages, with growing season precipitation ranging from 11 mm to 285 mm and growing degree days ranging from 1,245 to 1,846.DiscussionWe determined that (1) bacterial beta diversity is structured by growing season precipitation, (2) fungal beta diversity reflects growing season precipitation and growing degree days, and (3) microbial differential abundances of specific genera vary with vintage, growing season precipitation, and fruit maturity metrics. Further, the correlation between fungal community dissimilarities and geographic distance suggests dispersal limitation and the vineyard as a source for abundant fungal taxa. Contrasting this observation, the lack of correlation between bacterial community dissimilarity and geographic distance suggests that environmental filtering is shaping these communities. creator: Kerri L. Steenwerth creator: Ian Morelan creator: Ruby Stahel creator: Rosa Figueroa-Balderas creator: Dario Cantu creator: Jungmin Lee creator: Ron C. Runnebaum creator: Amisha T. Poret-Peterson uri: https://doi.org/10.7717/peerj.10836 license: http://creativecommons.org/publicdomain/zero/1.0/ rights: title: healthcareCOVID: a national cross-sectional observational study identifying risk factors for developing suspected or confirmed COVID-19 in UK healthcare workers link: https://peerj.com/articles/10891 last-modified: 2021-02-04 description: ObjectiveTo establish the prevalence, risk factors and implications of suspected or confirmed coronavirus disease 2019 (COVID-19) infection among healthcare workers in the United Kingdom (UK).DesignCross-sectional observational study.SettingUK-based primary and secondary care.ParticipantsHealthcare workers aged ≥18 years working between 1 February and 25 May 2020.Main outcome measuresA composite endpoint of laboratory-confirmed diagnosis of SARS-CoV-2, or self-isolation or hospitalisation due to suspected or confirmed COVID-19.ResultsOf 6,152 eligible responses, the composite endpoint was present in 1,806 (29.4%) healthcare workers, of whom 49 (0.8%) were hospitalised, 459 (7.5%) tested positive for SARS-CoV-2, and 1,776 (28.9%) reported self-isolation. Overall, between 11,870 and 21,158 days of self-isolation were required by the cohort, equalling approximately 71 to 127 working days lost per 1,000 working days. The strongest risk factor associated with the presence of the primary composite endpoint was increasing frequency of contact with suspected or confirmed COVID-19 cases without adequate personal protective equipment (PPE): ‘Never’ (reference), ‘Rarely’ (adjusted odds ratio 1.06, (95% confidence interval: [0.87–1.29])), ‘Sometimes’ (1.7 [1.37–2.10]), ‘Often’ (1.84 [1.28–2.63]), ‘Always’ (2.93, [1.75–5.06]). Additionally, several comorbidities (cancer, respiratory disease, and obesity); working in a ‘doctors’ role; using public transportation for work; regular contact with suspected or confirmed COVID-19 patients; and lack of PPE were also associated with the presence of the primary endpoint. A total of 1,382 (22.5%) healthcare workers reported lacking access to PPE items while having clinical contact with suspected or confirmed COVID-19 cases.ConclusionsSuspected or confirmed COVID-19 was more common in healthcare workers than in the general population and is associated with significant workforce implications. Risk factors included inadequate PPE, which was reported by nearly a quarter of healthcare workers. Governments and policymakers must ensure adequate PPE is available as well as developing strategies to mitigate risk for high-risk healthcare workers during future COVID-19 waves. creator: Justin Kua creator: Reshma Patel creator: Eveliina Nurmi creator: Sarah Tian creator: Harpreet Gill creator: Danny J.N. Wong creator: Calvin Moorley creator: Dmitri Nepogodiev creator: Imran Ahmad creator: Kariem El-Boghdadly uri: https://doi.org/10.7717/peerj.10891 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Kua et al. title: Development and clinical validation of a 3-miRNA signature to predict prognosis of gastric cancer link: https://peerj.com/articles/10462 last-modified: 2021-02-03 description: AimsIdentification of miRNA signature to predict the prognosis of gastric cancer (GC) patients by integrating bioinformatics and experimental validation.MethodsThe miRNA expression profile and clinical data of GC were collected. The univariable and LASSO-Cox regression were used to construct the risk signature. The receiver operating characteristic (ROC) curve analysis confirmed the good performance of the prognostic model.ResultsA 3-miRNA prognostic signature was constructed, which included hsa-miR-126-3p, hsa-miR-143-5p, and hsa-miR-1275. A nomogram, including the prognostic signature to predict the overall survival, was established, and internal validation in the The Cancer Genome Atlas (TCGA) cohort was performed. We found that compared with the traditional pathological stage, the nomogram was the best at predicting the prognosis.ConclusionsThe predictive model and the nomogram will enable patients with GC to be more accurately managed in clinical practice. creator: Wenqian Qi creator: Qian Zhang uri: https://doi.org/10.7717/peerj.10462 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Qi and Zhang title: High expression of MCM10 is predictive of poor outcomes in lung adenocarcinoma link: https://peerj.com/articles/10560 last-modified: 2021-02-03 description: BackgroundsLung adenocarcinoma is a complex disease that results in over 1.8 million deaths a year. Recent advancements in treating and managing lung adenocarcinoma have led to modest decreases in associated mortality rates, owing in part to the multifactorial etiology of the disease. Novel prognostic biomarkers are needed to accurately stage the disease and act as the basis of adjuvant treatments.Material and MethodsThe microarray datasets GSE75037, GSE31210 and GSE32863 were downloaded from the Gene Expression Omnibus (GEO) database to identify prognostic biomarkers for lung adenocarcinoma and therapy. The differentially expressed genes (DEGs) were identified by GEO2R. Functional and pathway enrichment analysis were performed by Kyoto Encyclopedia of Genes and Genomes and Gene Ontology (GO). Validation was performed based on 72 pairs of lung adenocarcinoma and adjacent normal lung tissues.ResultsResults showed that the DEGs were mainly focused on cell cycle and DNA replication initiation. Forty-one hub genes were identified and further analyzed by CytoScape. Here, we provide evidence which suggests MCM10 is a potential target with prognostic, diagnostic and therapeutic value. We base this on an integrated approach of comprehensive bioinformatics analysis and in vitro validation using the A549 lung adenocarcinoma cell line. We show that MCM10 overexpression correlates with a poor prognosis, while silencing of this gene decreases aberrant growth by 2-fold. Finally, evaluation of 72 clinical biopsy samples suggests that overexpression of MCM10 in the lung adenocarcinoma highly correlates with larger tumor size. Together, this work suggests that MCM10 may be a clinically relevant gene with both predictive and therapeutic value in lung adenocarcinoma. creator: Mingrui Shao creator: Shize Yang creator: Siyuan Dong uri: https://doi.org/10.7717/peerj.10560 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Shao et al. title: Ginsenoside Rb1, salvianolic acid B and their combination modulate gut microbiota and improve glucolipid metabolism in high-fat diet induced obese mice link: https://peerj.com/articles/10598 last-modified: 2021-02-03 description: BackgroundTo observe the effect of ginsenoside Rb1, salvianolic acid B and their combination on glucolipid metabolism and structural changes of gut microbiota.MethodsEight-week-old C57BL/6J mice were fed 45% high-fat diet to induce obesity. The obese mice were randomly divided into four groups, Con group as model control, ginsenoside Rb1 (Rb1) group, salvianolic acid B (SalB) group and ginsenoside Rb1+ salvianolic acid B (Rb1SalB) group. Mice in Rb1, SalB and Rb1SalB group were treated by gavage with ginsenoside Rb1, salvianolic acid B and the combination of the two ingredients, respectively. While mice in Con group were given the same amount of sterile water. The intervention lasted 8 weeks. Body weight and fasting blood glucose were measured every 2 weeks. Oral glucose tolerance test was conducted on the 4th and 8th week of drug intervention. At the end of the experiment, total cholesterol, triglyceride, high density lipoprotein cholesterol, low density lipoprotein cholesterol and non-esterified fatty acid content as well as glycated hemoglobin were measured and feces were collected for 16S rDNA sequencing.ResultsBoth ginsenoside Rb1 and Rb1SalB combination decreased body weight significantly (P < 0.05). Ginsenoside Rb1, salvianolic acid B and their combination alleviated fasting blood glucose, glycated hemoglobin and blood lipid profiles effectively (P < 0.05, compared with the corresponding indicators in Con group). Oral glucose tolerance test results at the 8th week showed that glucose tolerance was significantly improved in all three treatment groups. Ginsenoside Rb1, salvianolic acid B and their combination reduced the overall diversity of gut microbiota in feces and changed the microbial composition of the obese mice. LDA effect size (LefSe) analysis revealed the key indicator taxa corresponding to the treatment.ConclusionGinsenoside Rb1, salvianolic acid B and their combination could lower blood glucose and lipid level, and improve glucose tolerance of obese mice. The above effect may be at least partially through modulation of gut microbial composition. creator: Ying Bai creator: Xueli Bao creator: Qianqian Mu creator: Xin Fang creator: Ruyuan Zhu creator: Chenyue Liu creator: Fangfang Mo creator: Dongwei Zhang creator: Guangjian Jiang creator: Ping Li creator: Sihua Gao creator: Dandan Zhao uri: https://doi.org/10.7717/peerj.10598 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Bai et al. title: Liver cancer mortality over six decades in an epidemic area: what we have learned link: https://peerj.com/articles/10600 last-modified: 2021-02-03 description: Background and aims:Liver cancer is one of the most dominant malignant tumors in the world. The trends of liver cancer mortality over the past six decades have been tracked in the epidemic region of Qidong, China. Using epidemiological tools, we explore the dynamic changes in age-standardized rates to characterize important aspects of liver cancer etiology and prevention.MethodsMortality data of liver cancer in Qidong from 1958 to 1971 (death retrospective survey) and from 1972 to 2017 (cancer registration) were tabulated for the crude rate (CR), and age-standardized rate and age-birth cohorts. The average annual percentage change was calculated by the Joinpoint Regression Program.ResultsThe natural death rate during 1958–2017 decreased from 9‰ to 5.4‰ and then increased to 8‰ as the population aged; cancer mortality rates rose continuously from 57/105 to 240/105. Liver cancer mortality increased from 20/105 to 80/105, and then dropped to less than 52/105 in 2017. Liver cancer deaths in 1972–2017 accounted for 30.53% of all cancers, with a CR of 60.48/105, age-standardized rate China (ASRC) of 34.78/105, and ASRW (world) of 45.71/105. Other key features were the CR for males and females of 91.86/105 and 29.92/105, respectively, with a sex ratio of 3.07:1. Period analysis showed that the ASRs for mortality of the age groups under 54 years old had a significant decreasing trend. Importantly, birth cohort analysis showed that the mortality rate of liver cancer in 40–44, 35–39, 30–34, 25–29, 20–24, 15–19 years cohort decreased considerably, but the rates in 70–74, and 75+ increased.ConclusionsThe crude mortality rate of liver cancer in Qidong has experienced trends from lower to higher levels, and from continued increase at a high plateau to most recently a gradual decline, and a change greatest in younger people. Many years of comprehensive prevention and intervention measures have influenced the decline of the liver cancer epidemic in this area. The reduction of intake levels of aflatoxin might be one of the most significant factors as evidenced by the dramatic decline of exposure biomarkers in this population during the past three decades. creator: Jian-Guo Chen creator: Jian Zhu creator: Yong-Hui Zhang creator: Yong-Sheng Chen creator: Jian-Hua Lu creator: Yuan-Rong Zhu creator: Hai-Zhen Chen creator: Ai-Guo Shen creator: Gao-Ren Wang creator: John D. Groopman creator: Thomas W. Kensler uri: https://doi.org/10.7717/peerj.10600 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Chen et al. title: Effects of oxygen exposure on relative nucleic acid content and membrane integrity in the human gut microbiota link: https://peerj.com/articles/10602 last-modified: 2021-02-03 description: While the diversity of the human gut microbiota is becoming increasingly well characterized, bacterial physiology is still a critical missing link in understanding how the gut microbiota may be implicated in disease. The current best practice for studying bacterial physiology involves the immediate storage of fecal samples in an anaerobic chamber. This reliance on immediate access to anaerobic chambers greatly limits the scope of sample populations that can be studied. Here, we assess the effects of short-term oxygen exposure on gut bacterial physiology and diversity. We use relative nucleic acid content and membrane integrity as markers of bacterial physiology, and 16S rRNA gene amplicon sequencing to measure bacterial diversity. Samples were stored for up to 6 h in either ambient conditions or in anoxic environments created with gas packs or in an anaerobic chamber. Our data indicate that AnaeroGen sachets preserve bacterial membrane integrity and nucleic acid content over the course of 6 h similar to storage in an anaerobic chamber. Short-term oxygen exposure increases bacterial membrane permeability, without exceeding inter-individual differences. As oxygen exposure remains an important experimental consideration for bacterial metabolism, our data suggest that AnaeroGen sachets are a valid alternative limiting loss of membrane integrity for short-term storage of samples from harder-to-access populations. creator: Mariia Taguer creator: Ophélie Quillier creator: Corinne F. Maurice uri: https://doi.org/10.7717/peerj.10602 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Taguer et al. title: COVID-19: exploring impacts of the pandemic and lockdown on mental health of Pakistani students link: https://peerj.com/articles/10612 last-modified: 2021-02-03 description: BackgroundAs of the present, the twenty-first century is experiencing what may be one of its most devastating events, in respect to infected and dead people by the virus. Now known to the world as COVID-19, the devastating disease of what has become a pandemic started its spread from Wuhan, China and swiftly engulfed the whole world with almost 11 million cases, in a span of around six months. It has not only increased the global burden of disease but has heavily dented many social institutions, including education.MethodsThis study investigates how the COVID-19 pandemic and subsequent measures of lockdown, quarantine, and social distancing have affected students. We look specifically into the effects on individuals’ mental health, that is, the stress and anxiety levels of college and university students using the Zung Self-rating Anxiety Scale (SAS).ResultsAmong 494 respondents, 61% were females, and the majority (77.3%) of the students were in the age group of 19–25 years. Among the respondents, 125 (25.3%), 45 (9.1%) and 34 (6.9%) experienced minimal to moderate, severe, and most extreme levels of anxiety, respectively. The variables of gender, age and year of study were significant at the 0.25 level by univariate analyses. Nevertheless, the ordinal regression indicates that only gender was significant. The odds of a female student being more anxious are higher compared to a male student (OR = 1.779, 95% CI [1.202–2.634], P = 0.004). The most prominent stressors attained from the qualitative feedback from the Pakistani students are associated with online teaching, concerns about their academic performance and completion of the current semester, uncertainty related to exam dates, and the status of the following semester.ConclusionsThis study will add to the existing body of literature on the impacts of the COVID-19 pandemic on the social and psychological health of students. The study outcomes will provide basic data for further applied and action research and a framework for universities and policy makers in Pakistan and the neighboring countries in the region with the same cultural contexts. Thus, relevant health interventions can be designed for better mental health and educational attainments of students from higher educational institutions. This pathological pandemic may well lead to another pandemic of mental and behavioral illness. All stakeholders should join force regardless of pre-existing differences and inequalities to ensure the well-being of future generations, specifically students from higher educational institutions. The long-lasting impacts and the aftermath of this pandemic will unquestionably need further and future investigations. Keywords: Anxiety, students, mental health, COVID-19, Pakistan creator: Gul Muhammad Baloch creator: Sheela Sundarasen creator: Karuthan Chinna creator: Mohammad Nurunnabi creator: Kamilah Kamaludin creator: Heba Bakr Khoshaim creator: Syed Far Abid Hossain creator: Areej AlSukayt uri: https://doi.org/10.7717/peerj.10612 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Baloch et al. title: Effects of self-isolation and quarantine on loot box spending and excessive gaming—results of a natural experiment link: https://peerj.com/articles/10705 last-modified: 2021-02-03 description: COVID-19 has prompted widespread self-isolation and citywide/countrywide lockdowns. The World Health Organisation (WHO) has encouraged increased digital social activities such as video game play to counteract social isolation during the pandemic. However, there is active debate about the potential for video game overuse, and some video games contain randomised purchases (loot boxes) that may psychologically approximate gambling. In this pre-registered study, we examined the effects of self-isolation and quarantine on excessive gaming and loot box spending. We recruited 1,144 (619 male, 499 female, 26 other) Australian, Aotearoa New Zealand, and US residents who self reported being quarantined or self-isolating (n = 447) or not (n = 619) during the COVID-19 pandemic to a cross-sectional natural experiment. We compared the associations between problem gambling symptomology, excessive gaming and loot box spending for isolated and non-isolated participants. Participants completed the Kessler-10 Psychological Distress Scale, Problem Gambling Severity Index, Internet Gaming Disorder Checklist, a measure of risky engagement with loot boxes, concern about contamination, and reported money spent on loot boxes in the past month, as well as whether they were quarantined or under self-isolation during the pandemic. Although, in our data, excessive gaming and loot box spending were not higher for isolated (self-isolated/ quarantined) compared to non-isolated gamers, the established association between problem gambling symptomology and loot box spending was stronger among isolated gamers than those not isolated. Concerns about being contaminated by germs was also significantly associated with greater excessive gaming and, to a lesser extent, loot box spending irrespective of isolation status. Gamers might be managing concerns about the pandemic with greater video game use, and more problem gamblers may be purchasing loot boxes during the pandemic. It is unclear whether these relationships may represent temporary coping mechanisms which abate when COVID-19 ends. Re-examination as the pandemic subsides may be required. More generally, the results suggest that social isolation during the pandemic may inflate the effect size of some media psychology and gaming effects. We urge caution not to generalise psychological findings from research conducted during the COVID-19 pandemic to be necessarily representative of the magnitude of relationships when not in a pandemic. creator: Lauren C. Hall creator: Aaron Drummond creator: James D. Sauer creator: Christopher J. Ferguson uri: https://doi.org/10.7717/peerj.10705 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Hall et al. title: A novel bicyclic 2,4-diaminopyrimidine inhibitor of Streptococcus suis dihydrofolate reductase link: https://peerj.com/articles/10743 last-modified: 2021-02-03 description: Streptococcus suis is a Gram-positive bacterial pathogen of pigs and an emerging zoonotic pathogen. It has become increasingly resistant to multiple classes of antibiotics. New drug candidates and knowledge of their targets are needed to combat antibiotic-resistant S. suis. In this study, the open-source Pathogen Box compound library was screened. Thirty hits that effectively inhibited S. suis growth at 10 µM were identified. Among the most potent hits, MMV675968 (a diaminoquinazoline analog) was shown to target S. suis dihydrofolate reductase (SsDHFR) via (1) growth inhibition of an E. coli surrogate whose growth is dependent on exogenously expressed SsDHFR and (2) inhibition of in vitro SsDHFR activity. Thymidine supplement is able to reverse growth inhibition by MMV675968 in both E. coli surrogate and S. suis, indicating that a thymidine-related pathway is a major target of MMV675968. Comparison of MMV675968 with seven DHFR inhibitors representing different core structures revealed that bicyclic 2,4-diaminopyrimidines with long and flexible side chains are highly effective in inhibiting SsDHFR and S. suis growth. MMV675968 and related compounds thus may serve as starting points for developing antibiotics against drug resistant S. suis. creator: Warangkhana Songsungthong creator: Sunisa Prasopporn creator: Louise Bohan creator: Potjanee Srimanote creator: Ubolsree Leartsakulpanich creator: Suganya Yongkiettrakul uri: https://doi.org/10.7717/peerj.10743 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Songsungthong et al. title: Offspring and adult chemosensory recognition by an amphisbaenian reptile may allow maintaining familiar links in the fossorial environment link: https://peerj.com/articles/10780 last-modified: 2021-02-03 description: Kin recognition is a phenomenon with an important function in maintaining cohesive social groups in animals. Several studies have examined parent–offspring recognition in species with direct parental care. Few studies have, however, explored parent–offspring recognition in animals that, at best, only show apparent indirect parental care, such as some reptiles. In this study, we investigated reciprocal parent–offspring recognition in the fossorial amphisbaenian Trogonophis wiegmanni, a viviparous species that shows potential stable ‘family groups’ in the form of parent-offspring long-term associations. We examined whether adult males and females could discriminate via chemical cues between familiar juveniles which associate with them within their family groups, and are potentially their offspring, to that of unfamiliar juveniles, and whether juveniles could discriminate between familiar adult males and females of their family group (probably their parents) and unfamiliar unrelated adults. We measured tongue flick behavior to study chemosensory responses to the scent of conspecifics. We found that adult female amphisbaenians, but not males, could discriminate between scents of familiar and unfamiliar juveniles. Juvenile amphisbaenians did not discriminate between familiar and unfamiliar adult females, but recognize familiar from unfamiliar males. We discuss our results of parent–offspring recognition according to its potential social function in an ecological fossorial context where visibility is limited and chemosensory kin recognition may contribute to the establishment of stable family groups. creator: José Martín creator: Ernesto Raya-García creator: Jesús Ortega creator: Pilar López uri: https://doi.org/10.7717/peerj.10780 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Martín et al. title: Laparoscopic sleeve gastrectomy under general anesthesia in severely obese patients: a single-centered retrospective study link: https://peerj.com/articles/10802 last-modified: 2021-02-03 description: BackgroundThis study aims to summarize and analyze the clinical characteristics and outcomes of severely obese patients who underwent laparoscopic sleeve gastrectomy (LSG) under general anesthesia with multidisciplinary collaboration.MethodsA retrospective analysis was performed for 100 severely obese patients who were hospitalized in Zhongshan Hospital, Fudan University from January 2017 to December 2019, and included preoperative general information, laboratory examinations, anesthesia and outcomes.ResultsA total of 100 patients (46 males, 54 females) were admitted to the department of endocrinology: 100 had hepatic steatosis (100%), 43 had sleep apnea hypopnea syndrome (43%), 25 had hypertension (25%), 11 had type 2 diabetes (11%) and 8 had polycystic ovary syndrome (14% of women). The mean age and BMI were 31.52 ± 10.53 years and 43.31 ± 6.80 kg/m2, respectively. Visual laryngoscope intubation was successfully performed with routine intravenous induction in the optimum sniffing position at one time. The surgeries were successfully performed under general anesthesia, without conversion, and the operation time was 140.92 ± 31.23 min. The follow-up data for 41 patients were obtained. The postoperative BMI showed a downward trend. The BMI at 1 month and 3 months after surgery were 38.40 ± 6.77 kg/m2and 35.52 ± 7.94 kg/m2, respectively.ConclusionsMultidisciplinary collaboration may contribute to better management and recovery during the perioperative period. Visual laryngoscope intubation with intravenous induction was performed successfully in the optimum sniffing position at one time. creator: Yuanyuan Ma creator: Yu Fan creator: Di Zhou creator: Junjun Chen creator: Shengjin Ge uri: https://doi.org/10.7717/peerj.10802 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Ma et al. title: Assessing the effects of time-dependent restrictions and control actions to flatten the curve of COVID-19 in Kazakhstan link: https://peerj.com/articles/10806 last-modified: 2021-02-03 description: This article presents the assessment of time-dependent national-level restrictions and control actions and their effects in fighting the COVID-19 pandemic. By analysing the transmission dynamics during the first wave of COVID-19 in the country, the effectiveness of the various levels of control actions taken to flatten the curve can be better quantified and understood. This in turn can help the relevant authorities to better plan for and control the subsequent waves of the pandemic. To achieve this, a deterministic population model for the pandemic is firstly developed to take into consideration the time-dependent characteristics of the model parameters, especially on the ever-evolving value of the reproduction number, which is one of the critical measures used to describe the transmission dynamics of this pandemic. The reproduction number alongside other key parameters of the model can then be estimated by fitting the model to real-world data using numerical optimisation techniques or by inducing ad-hoc control actions as recorded in the news platforms. In this article, the model is verified using a case study based on the data from the first wave of COVID-19 in the Republic of Kazakhstan. The model is fitted to provide estimates for two settings in simulations; time-invariant and time-varying (with bounded constraints) parameters. Finally, some forecasts are made using four scenarios with time-dependent control measures so as to determine which would reflect on the actual situations better. creator: Ton Duc Do creator: Meei Mei Gui creator: Kok Yew Ng uri: https://doi.org/10.7717/peerj.10806 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Do et al. title: 6mA-Pred: identifying DNA N6-methyladenine sites based on deep learning link: https://peerj.com/articles/10813 last-modified: 2021-02-03 description: With the accumulation of data on 6mA modification sites, an increasing number of scholars have begun to focus on the identification of 6mA sites. Despite the recognized importance of 6mA sites, methods for their identification remain lacking, with most existing methods being aimed at their identification in individual species. In the present study, we aimed to develop an identification method suitable for multiple species. Based on previous research, we propose a method for 6mA site recognition. Our experiments prove that the proposed 6mA-Pred method is effective for identifying 6mA sites in genes from taxa such as rice, Mus musculus, and human. A series of experimental results show that 6mA-Pred is an excellent method. We provide the source code used in the study, which can be obtained from http://39.100.246.211:5004/6mA_Pred/. creator: Qianfei Huang creator: Wenyang Zhou creator: Fei Guo creator: Lei Xu creator: Lichao Zhang uri: https://doi.org/10.7717/peerj.10813 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Huang et al. title: The expression characteristics and prognostic roles of autophagy-related genes in gastric cancer link: https://peerj.com/articles/10814 last-modified: 2021-02-03 description: BackgroundAutophagy is an evolutionally highly conserved process, accompanied by the dynamic changes of various molecules, which is necessary for the orderly degradation and recycling of cellular components. The aim of the study was to identify the role of autophagy-related (ATG) genes in the occurrence and development of gastric cancer (GC).MethodsData from Oncomine dataset was used for the differential expression analysis between cancer and normal tissues. The association of ATG genes expression with clinicopathologic indicators was evaluated by The Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database. Moreover, using the TCGA datasets, the prognostic role of ATG genes was assessed. A nomogram was further built to assess the independent prognostic factors.ResultsThe expression of autophagy-related genes AMBRA1, ATG4B, ATG7, ATG10, ATG12, ATG16L2, GABARAPL2, GABARAPL1, ULK4 and WIPI2 showed differences between cancer and normal tissues. After verification, ATG14 and ATG4D were significantly associated with TNM stage. ATG9A, ATG2A, and ATG4D were associated with T stage. VMP1 and ATG4A were low-expressed in patients without lymph node metastasis. No gene in autophagy pathway was associated with M stage. Further multivariate analysis suggested that ATG4D and MAP1LC3C were independent prognostic factors for GC. The C-index of nomogram was 0.676 and the 95% CI was 0.628 to 0.724.ConclusionOur study provided a comprehensive illustration of ATG genes expression characteristics in GC. Abnormal expressions of the ubiquitin-like conjugated system in ATG genes plays a key role in the occurrence of GC. ATG8/LC3 sub-system may play an important role in development and clinical outcome of GC. In the future, it is necessary to further elucidate the alterations of specific ATG8/LC3 forms in order to provide insights for the discovery, diagnosis, or targeting for GC. creator: Mengya Wang creator: Jingjing Jing creator: Hao Li creator: Jingwei Liu creator: Yuan Yuan creator: Liping Sun uri: https://doi.org/10.7717/peerj.10814 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: Evaluation of FGFR1 as a diagnostic biomarker for ovarian cancer using TCGA and GEO datasets link: https://peerj.com/articles/10817 last-modified: 2021-02-03 description: BackgroundMalignant ovarian cancer is associated with the highest mortality of all gynecological tumors. Designing therapeutic targets that are specific to OC tissue is important for optimizing OC therapies. This study aims to identify different expression patterns of genes related to FGFR1 and the usefulness of FGFR1 as diagnostic biomarker for OC.MethodsWe collected data from The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) databases. In the TCGA cohort we analyzed clinical information according to patient characteristics, including age, stage, grade, longest dimension of the tumor and the presence of a residual tumor. GEO data served as a validation set. We obtained data on differentially expressed genes (DEGs) from the two microarray datasets. We then used gene set enrichment analysis (GSEA) to analyze the DEG data in order to identify enriched pathways related to FGFR1.ResultsDifferential expression analysis revealed that FGFR1 was significantly downregulated in OC specimens. 303 patients were included in the TCGA cohort. The GEO dataset confirmed these findings using information on 75 Asian patients. The GSE105437 and GSE12470 database highlighted the significant diagnostic value of FGFR1 in identifying OC (AUC = 1, p = 0.0009 and AUC = 0.8256, p = 0.0015 respectively).ConclusionsOur study examined existing TCGA and GEO datasets for novel factors associated with OC and identified FGFR1 as a potential diagnostic factor. Further investigation is warranted to characterize the role played by FGFR1 in OC. creator: Huiting Xiao creator: Kun Wang creator: Dan Li creator: Ke Wang creator: Min Yu uri: https://doi.org/10.7717/peerj.10817 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Xiao et al. title: Cytochrome P450 monooxygenase genes in the wild silkworm, Bombyx mandarina link: https://peerj.com/articles/10818 last-modified: 2021-02-03 description: Wild (Bombyx mandarina) and domestic silkworms (B. mori) are good models for investigating insect domestication, as 5000 years of artificial breeding and selection have resulted in significant differences between B. mandarina and B. mori. In this study, we improved the genome assemblies to the chromosome level and updated the protein-coding gene annotations for B. mandarina. Based on this updated genome, we identified 68 cytochrome P450 genes in B. mandarina. The cytochrome P450 repository in B. mandarina is smaller than in B. mori. Certain currently unknown key genes, rather than gene number, are critical for insecticide resistance in B. mandarina, which shows greater resistance to insecticides than B. mori. Based on the physical maps of B. mandarina, we located 66 cytochrome P450s on 18 different chromosomes, and 27 of the cytochrome P450 genes were concentrated into seven clusters. KEGG enrichment analysis of the P450 genes revealed the involvement of cytochrome P450 genes in hormone biosynthesis. Analyses of the silk gland transcriptome identified candidate cytochrome P450 genes (CYP306A) involved in ecdysteroidogenesis and insecticide metabolism in B. mandarina. creator: Linrong Wan creator: Anlian Zhou creator: Wenfu Xiao creator: Bangxing Zou creator: Yaming Jiang creator: Jinshu Xiao creator: Cao Deng creator: Youhong Zhang uri: https://doi.org/10.7717/peerj.10818 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wan et al. title: Social behavior of musk deer during the mating season potentially influences the diversity of their gut microbiome link: https://peerj.com/articles/10860 last-modified: 2021-02-03 description: An increasing body of research has revealed that social behavior shapes the animal gut microbiome community and leads to the similarity among the same social group. However, some additional factors (e.g., diet and habitat within each social group) may also contribute to this similarity within the social group and dissimilarity between social groups. Here, we investigated the potential correlation between social behavior and the gut microbiome community in 179 musk deer from four breeding regions in the Maerkang Captive Center, Sichuan. The dominant gut microbiome phyla in the musk deer in this study were Firmicutes, Bacteroidetes, and Proteobacteria. We found significant effects on the alpha and beta diversity of the gut microbiome due to the breeding regions. The similarity within breeding regions was higher than that between the breeding regions. Due to their solitary lifestyle, captive musk deer are raised in single cages with no direct social contact most of the time. Deer in all of the breeding regions have the same diet and similar living conditions. However, during each mating season from November to January, in each region, one adult male and about six adult females will be put together into a large cage. Social behavior happens during cohabitation, including mating behavior, grooming within the same sex or between different sexes, and other social contact. Therefore, we speculated that high similarity within the breeding region might be associated with the social behavior during the mating season. This was a simple and straightforward example of the relationship between animal social behavior and the gut microbiome. creator: Jianmei Li creator: Wei Luo creator: Yudong Zhu creator: Qinlong Dai creator: Guoqi Liu creator: Chengli Zheng creator: Lei Zhou creator: Shengqiang Li creator: Zhu Chen creator: Jianming Wang creator: Dayong Feng creator: Kunlin Yang creator: Zhisong Yang creator: Lifeng Zhu uri: https://doi.org/10.7717/peerj.10860 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Li et al. title: A method to adjust a prior distribution in Bayesian second-level fMRI analysis link: https://peerj.com/articles/10861 last-modified: 2021-02-03 description: Previous research has shown the potential value of Bayesian methods in fMRI (functional magnetic resonance imaging) analysis. For instance, the results from Bayes factor-applied second-level fMRI analysis showed a higher hit rate compared with frequentist second-level fMRI analysis, suggesting greater sensitivity. Although the method reported more positives as a result of the higher sensitivity, it was able to maintain a reasonable level of selectivity in term of the false positive rate. Moreover, employment of the multiple comparison correction method to update the default prior distribution significantly improved the performance of Bayesian second-level fMRI analysis. However, previous studies have utilized the default prior distribution and did not consider the nature of each individual study. Thus, in the present study, a method to adjust the Cauchy prior distribution based on a priori information, which can be acquired from the results of relevant previous studies, was proposed and tested. A Cauchy prior distribution was adjusted based on the contrast, noise strength, and proportion of true positives that were estimated from a meta-analysis of relevant previous studies. In the present study, both the simulated images and real contrast images from two previous studies were used to evaluate the performance of the proposed method. The results showed that the employment of the prior adjustment method resulted in improved performance of Bayesian second-level fMRI analysis. creator: Hyemin Han uri: https://doi.org/10.7717/peerj.10861 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Han title: Mitochondrial genomes of eight Scelimeninae species (Orthoptera) and their phylogenetic implications within Tetrigoidea link: https://peerj.com/articles/10523 last-modified: 2021-02-02 description: Scelimeninae is a key member of the pygmy grasshopper community, and an important ecological indicator. No mitochondrial genomes of Scelimeninae have been reported to date, and the monophyly of Scelimeninae and its phylogenetic relationship within Tetrigidae is still unclear. We sequenced and analyzed eight nearly complete mitochondrial genomes representing eight genera of Scelimeninae. These mitogenomes ranged in size from 13,112 to 16,380 bp and the order of tRNA genes between COII and ATP8 was reversed compared with the ancestral order of insects. The protein-coding genes (PCGs) of tetrigid species mainly with the typical ATN codons and most terminated with complete (TAA or TAG) stop codons. Analyses of pairwise genetic distances showed that ATP8 was the least conserved gene within Tetrigidae, while COI was the most conserved. The longest intergenic spacer (IGS) region in the mitogenomes was always found between tRNASer(UCN) and ND1. Additionally, tandem repeat units were identified in the longest IGS of three mitogenomes. Maximum likelihood (ML) and Bayesian Inference (BI) analyses based on the two datasets supported the monophyly of Tetriginae. Scelimeninae was classified as a non-monophyletic subfamily. creator: Ran Li creator: Xiaoli Ying creator: Weian Deng creator: Wantao Rong creator: Xiaodong Li uri: https://doi.org/10.7717/peerj.10523 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Li et al. title: A molecular phylogeny of Geotrochus and Trochomorpha species (Gastropoda: Trochomorphidae) in Sabah, Malaysia reveals convergent evolution of shell morphology driven by environmental influences link: https://peerj.com/articles/10526 last-modified: 2021-02-02 description: There are currently eleven Geotrochus and four Trochomorpha species in Sabah. The primary diagnostic character that separates the two genera is the intensity of sculpture on the shell upper surface. All Trochomorpha species have a coarse nodular sculpture while Geotrochus species has a non-nodular sculpture or smooth shell. However, it is known that shell characters are often evolutionary labile with high plasticity in response to environmental factors. Hence, identifying the phylogenetic and ecological determinants for the shell characters will shed light on the shell-based taxonomy. This study aims to estimate the phylogenetic relationship between Geotrochus and Trochomorpha species in Sabah based in two mitochondrial genes (COI, 16S) and one nuclear gene (ITS) and also to examine the influence of temperature, elevation and annual precipitation on the coarseness of shell upper surface sculpture and shell sizes of the species of both genera. Additionally, we also investigated the phylogenetic signal of the shell characters. The phylogenetic analysis showed that Geotrochus and Trochomorpha species are not reciprocally monophyletic. The phylogenetic signal test suggested that shell size and upper surface sculpture are homoplastic, and these shell traits are strongly influenced by elevation and annual precipitation, particularly at the cloud zone of Mount Kinabalu. The highland species of both genera have a coarser shell surface than lowland species. The shell and aperture width decrease with increasing elevation and annual precipitation. In the view of finding above, the current taxonomy of Geotrochus and Trochmorpha in this region and elsewhere that based on shell characters need to be revised with sufficient specimens throughout the distribution range of the two genera. creator: Zi-Yuan Chang creator: Thor-Seng Liew uri: https://doi.org/10.7717/peerj.10526 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Chang and Liew title: Effects of mixing two legume species at seedling stage under different environmental conditions link: https://peerj.com/articles/10615 last-modified: 2021-02-02 description: While intercropping is known to have positive effects on crop productivity, it is unclear whether the effects of mixing species start at the early plant stage, that is, during germination. We tested whether the germination of two legume species, alsike clover and black medic, characterized by a contrasting response to water availability and temperature is affected by mixing. We set up four experiments in each of which we compared a 1:1 mixture against the two monocultures, and combined this with various other experimental factors. These additional factors were (i) varied seed densities (50%, 100% and 150% of a reference density) in two field trials in 2016 and 2017, (ii) varied seed densities (high and low) and water availability (six levels, between 25% and 100% of water holding capacity (WHC)) in a greenhouse pot trial, (iii) varied seed spacing in a climate chamber, and (iv) varied temperatures (12 °C, 20 °C and 28 °C) and water availability (four levels between 25% and 100% of WHC) in a climate chamber. Across all experiments, the absolute mixture effects (AME) on germination ranged between −9% and +11%, with a median of +1.3%. Within experiments, significant mixture effects were observed, but the direction of these effects was inconsistent. In the field, AME on germination was significantly negative at some of the tested seed densities. A positive AME was observed in the climate chamber at 12 °C, and the mean AME decreased with increasing temperature. Higher density was associated with decreased germination in the field, indicating negative interaction through competition or allelopathy, among seedlings. Our findings indicate that interaction among seeds in species mixtures may be ongoing during germination, but that the direction of the mixture effect is affected by complex interactions with abiotic and biotic factors. creator: Heba Elsalahy creator: Sonoko Bellingrath-Kimura creator: Timo Kautz creator: Thomas Döring uri: https://doi.org/10.7717/peerj.10615 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Elsalahy et al. title: Tolerance of two anhydrobiotic tardigrades Echiniscus testudo and Milnesium inceptum to hypomagnetic conditions link: https://peerj.com/articles/10630 last-modified: 2021-02-02 description: The open space is a hostile environment for all lifeforms not only due to vacuum, high radiation, low atmospheric pressure, and extremely low temperature, but also the absence of the geomagnetic field. The geomagnetic field protects Earth mainly from corpuscular radiation, that is, solar wind and cosmic radiation, but above all it influences organisms, including their cells, tissues and organs. Moreover, numerous studies conducted on plants and animals confirmed that hypomagnetic conditions (the term referring to all situations when the magnetic field is weaker than the typical geomagnetic field) have significant influence on the metabolism of living organisms. Although many studies dealt with a variety of aspects related mainly to the influence of hypomagnetic conditions on human health. Very few studies have considered the influence of hypomagnetic conditions on extremophiles. Astrobiologists have long been testing different extremofiles to find out if any multicellular organisms are able to survive in extreme conditions of open space. Among all multicellular extremophiles fit for such research, water bears (Tardigrada) are the most interesting. Not only are they one of the most resistant organisms on Earth, but results obtained from studies on these invertebrates can be extrapolated or applied to vertebrates (including humans). Despite this, studies on the influence of hypomagnetic conditions on tardigrades are rare, so far. In the present study, to test the influence of hypomagnetic conditions on the process of anhydrobiosis while entering and returning from anhydrobiosis, we used two terrestrial anhydrobiotic species that are Echiniscus testudo and Milnesium inceptum. To exclude the ambient magnetic field, experiments were carried out in a special magnetic field shielding chamber. In total, three experiments were conducted: (a) tardigrades in anhydrobiosis, (b) tardigrades entering anhydrobiosis and (c) tardigrades returning to active life. The obtained results clearly showed that even partial isolation from the geomagnetic field, that is, hypomagnetic conditions, has negative influence on anhydrobiotic abilities of both tested tardigrade species. In both species we observed lower survivability rate while entering anhydrobiosis, in anhydrobiotic state and returning to the active state. What is more, we observed a higher mortality rate in Ech. testudo than Mil. inceptum which suggest that different species response to the hypomagnetic conditions in different way. In conclusion, while current knowledge on the influence of hypomagnetic conditions on mortality of invertebrates is very limited, our results suggest that the presence of the magnetic field is a very important factor which should be considered in further research focused on potential survival of Earth organisms in outer space, spacecrafts or different planets and moons. creator: Weronika Erdmann creator: Bogdan Idzikowski creator: Wojciech Kowalski creator: Jakub Z. Kosicki creator: Łukasz Kaczmarek uri: https://doi.org/10.7717/peerj.10630 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Erdmann et al. title: Inhibitory effect and mechanism of action (MOA) of hirsutine on the proliferation of T-cell leukemia Jurkat clone E6-1 cells link: https://peerj.com/articles/10692 last-modified: 2021-02-02 description: BackgroundThe bark of Uncaria rhynchophylla has been traditionally used to treat convulsion, bleeding, hypertension, auto-immune conditions, cancer, and other diseases. The main focus of this research is done for the purpose of exploring the antitumor activity and mechanism of action (MOA) for hirsutine isolated from U. rhynchophylla.MethodsJurkat clone E6-1 cells were treated using 10, 25 and 50 μM for 48 h. Inhibition of cell proliferation due to hirsutine treatment was evaluated by CCK8 assay. Flow cytometry was applied to ascertain Jurkat cell cycle progression and apoptosis after treatment with 10, 25 and 50 μM hirsutine for 48 h. The expression and level of the apoptosis-related genes and proteins was analyzed by Real-time Quantitative polymerase chain reaction (qPCR) and Western blotting method, respectively.ResultsCCK8 analyses revealed that hirsutine could significantly inhibit the proliferation of Jurkat clone E6-1 cells, in a concentration and time-dependent fashion. Flow cytometry assays revealed that hirsutine could drive apoptotic death and G0/G1 phase arrest in Jurkat cells. Apoptotic cells frequencies were 4.99 ± 0.51%, 13.69 ± 2.00% and 40.21 ± 15.19%, and respective cell cycle arrest in G0/G1 accounted for 34.85 ± 1.81%, 42.83 ± 0.70% and 49.12 ± 4.07%. Simultaneously, compared with the control group, Western blot assays indicated that the up-regulation of pro-apoptotic Bax, cleaved-caspase3, cleaved-caspase9 and Cyto c proteins, as well as the down-regulation of Bcl-2 protein which guards against cell death, might be correlated with cell death induction and inhibition of cell proliferation. QPCR analyses indicated that hirsutine could diminish BCL2 expression and, at the same time, improve Bax, caspase-3 and caspase-9 mRNA levels, thus reiterating a putative correlation of hirsutine treatment in vitro with apoptosis induction and inhibition of cell proliferation (p-value < 0.05). Excessive hirsutine damages the ultrastructure in mitochondria, leading to the release of Cyt c from the mitochondria to cytoplasm in Jurkat clone E6-1 cells, thereby inducing the activated caspase cascade apoptosis process through a mitochondria-mediated pathway.ConclusionAn important bioactive constituent—hirsutine—appears to have antitumor effects in human T-cell leukemia, thus enlightening the use of phytomedicines as a novel source for tumor therapy. It is speculated that hirsutine may induce apoptosis of Jurkat Clone E6-1 cells through the mitochondrial apoptotic pathway. creator: Jie Meng creator: Rui Su creator: Luping Wang creator: Bo Yuan creator: Ling Li uri: https://doi.org/10.7717/peerj.10692 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Meng et al. title: Local dynamics of a white syndrome outbreak and changes in the microbial community associated with colonies of the scleractinian brain coral Pseudodiploria strigosa link: https://peerj.com/articles/10695 last-modified: 2021-02-02 description: Reef corals in the Mexican Reef System have been severely affected by the emergence of a white syndrome that resembles both White Plague II and SCTLD descriptions. Meandroid scleractinian coral species are among the most severely affected. To gain insight into this affliction we conducted a broad study in the brain coral Pseudodiploria strigosa at a rear reef site in the NE Mexican Caribbean. We describe macro and microscopical signals of the disease, characterize the outbreak dynamics, the tissue histopathology, explore immunological responses in the individuals, and compare microbial assemblages associated with the surface mucus layer of healthy and unhealthy colonies. At the study site, the white syndrome outbreak on P. strigosa showed a high incidence rate in summer-fall and a low one in winter, as well as low survival expectation of diseased colonies at the end of the study. After 306 days of observation, out of 96 tracked colonies, eight remained apparently healthy and seven were diseased. No effective resistance to colony disease progression was observed once white syndrome signs developed. Tissue loss rate during the study varied among colonies (mean = 10.8 cm2, s.d. = 7.8 cm2) suggesting a complex relation between causal agents and colony resistance. The deterioration of tissues was evidenced from the basal to the surface body wall of polyps (up to 66% hypertrophy and liquefactive necrosis in unhealthy colonies), implying that microscopic alterations begin before macroscopic signals develop, suggesting this may be a systemic disease. We measured high levels of phenoloxidase (two orders of magnitude higher PO activity than P. strigosa affected by BBD) and antibacterial activity without significant reduction in unhealthy samples from the mucus layer, indicative of an enhanced immunological response. Results showed that opportunistic bacteria dominated damaged colonies, where six genera of the Bacteroidia class were found with significant changes in unhealthy colonies after DeSeq2 analysis. Nevertheless, histological observations did not support infection of the tissues. The opportunistic overload seems to be contained within the mucus layer but may be associated with the mortality of tissues in a yet unclear way. Future research should focus on experimental infections, the tracking of natural infections, and the immunocompetence of corals in the face of environmental pressures due to local, regional, and global impacts. If environmental deterioration is the primary cause of the continuing emergence and re-emergence of lethal coral diseases, as has been proposed by many authors, the only true option to effectively help preserve the coral reef biodiversity and services, is to restore the environmental quality of reef waters at the local scale and reduce greenhouse gases at the global scale. creator: Patricia E. Thome creator: Jacqueline Rivera-Ortega creator: Jenny C. Rodríguez-Villalobos creator: Daniel Cerqueda-García creator: Edgar O. Guzmán-Urieta creator: José Q. García-Maldonado creator: Natalia Carabantes creator: Eric Jordán-Dahlgren uri: https://doi.org/10.7717/peerj.10695 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Thome et al. title: Taxonomic, functional and expression analysis of viral communities associated with marine sponges link: https://peerj.com/articles/10715 last-modified: 2021-02-02 description: Viruses play an essential role in shaping the structure and function of ecological communities. Marine sponges have the capacity to filter large volumes of ‘virus-laden’ seawater through their bodies and host dense communities of microbial symbionts, which are likely accessible to viral infection. However, despite the potential of sponges and their symbionts to act as viral reservoirs, little is known about the sponge-associated virome. Here we address this knowledge gap by analysing metagenomic and (meta-) transcriptomic datasets from several sponge species to determine what viruses are present and elucidate their predicted and expressed functionality. Sponges were found to carry diverse, abundant and active bacteriophages as well as eukaryotic viruses belonging to the Megavirales and Phycodnaviridae. These viruses contain and express auxiliary metabolic genes (AMGs) for photosynthesis and vitamin synthesis as well as for the production of antimicrobials and the defence against toxins. These viral AMGs can therefore contribute to the metabolic capacities of their hosts and also potentially enhance the survival of infected cells. This suggest that viruses may play a key role in regulating the abundance and activities of members of the sponge holobiont. creator: Mary Nguyen creator: Bernd Wemheuer creator: Patrick W. Laffy creator: Nicole S. Webster creator: Torsten Thomas uri: https://doi.org/10.7717/peerj.10715 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Nguyen et al. title: Increased precipitation enhances soil respiration in a semi-arid grassland on the Loess Plateau, China link: https://peerj.com/articles/10729 last-modified: 2021-02-02 description: BackgroundPrecipitation influences the vulnerability of grassland ecosystems, especially upland grasslands, and soil respiration is critical for carbon cycling in arid grassland ecosystems which typically experience more droughty conditions.MethodsWe used three precipitation treatments to understand the effect of precipitation on soil respiration of a typical arid steppe in the Loess Plateau in north-western China. Precipitation was captured and relocated to simulate precipitation rates of 50%, 100%, and 150% of ambient precipitation.Results and DiscussionSoil moisture was influenced by all precipitation treatments. Shoot biomass was greater, though non-significantly, as precipitation increased. However, both increase and decrease of precipitation significantly reduced root biomass. There was a positive linear relationship between soil moisture and soil respiration in the study area during the summer (July and August), when most precipitation fell. Soil moisture, soil root biomass, pH, and fungal diversity were predictors of soil respiration based on partial least squares regression, and soil moisture was the best of these.ConclusionOur study highlights the importance of increased precipitation on soil respiration in drylands. Precipitation changes can cause significant alterations in soil properties, microbial fungi, and root biomass, and any surplus or transpired moisture is fed back into the climate, thereby affecting the rate of soil respiration in the future. creator: Yutao Wang creator: Yingzhong Xie creator: Gillian Rapson creator: Hongbin Ma creator: Le Jing creator: Yi Zhang creator: Juan Zhang creator: Jianping Li uri: https://doi.org/10.7717/peerj.10729 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: Identification of candidate genes influencing anthocyanin biosynthesis during the development and ripening of red and white strawberry fruits via comparative transcriptome analysis link: https://peerj.com/articles/10739 last-modified: 2021-02-02 description: Strawberries are one of the most economically important berry fruits worldwide and exhibit colours ranging from white to dark red, providing a rich genetic resource for strawberry quality improvement. In the present study, we conducted transcriptome analyses of three strawberry cultivars, namely, ‘Benihoppe’, ‘Xiaobai’, and ‘Snow White’, and compared their gene expression profiles. Among the high-quality sequences, 5,049 and 53,200 differentially expressed genes (DEGs) were obtained when comparing the diploid and octoploid strawberry genomes and analysed to identify anthocyanin-related candidate genes. Sixty-five DEGs in the diploid genome (transcriptome data compared to the diploid strawberry genome) and 317 DEGs in the octoploid genome (transcriptome data compared to the octoploid strawberry genome) were identified among the three cultivars. Among these DEGs, 19 and 70 anthocyanin pathway genes, six and 42 sugar pathway genes, 23 and 101 hormone pathway genes, and 17 and 104 transcription factors in the diploid and octoploid genomes, respectively, correlated positively or negatively with the anthocyanin accumulation observed among the three cultivars. Real-time qPCR analysis of nine candidate genes showed a good correlation with the transcriptome data. For example, the expression of PAL was higher in ‘Benihoppe’ and ‘Xiaobai’ than in ‘Snow White’, consistent with the RNA-seq data. Thus, the RNA-seq data and candidate DEGs identified in the present study provide a sound basis for further studies of strawberry fruit colour formation. creator: Fengli Zhao creator: Pan Song creator: Xiangfen Zhang creator: Gang Li creator: Panpan Hu creator: Ali Aslam creator: Xia Zhao creator: Houcheng Zhou uri: https://doi.org/10.7717/peerj.10739 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Zhao et al. title: Intestinal fatty acid-binding protein, a biomarker of intestinal barrier dysfunction, increases with the progression of type 2 diabetes link: https://peerj.com/articles/10800 last-modified: 2021-02-02 description: ObjectiveTo investigate serum intestinal fatty acid-binding protein (I-FABP) in two groups of patients with different duration of hyperglycemia in a cross-sectional study.Materials and MethodsIn the present study, a total of 280 individuals (158 outpatients and 122 inpatients) suffering from hyperglycemia were recruited between May and September 2019. The clinical information of all participants was collected from the hospital information system, including the duration of hyperglycemia, age, gender, hemoglobin A1c (HbA1c), 75-g oral glucose tolerance test including fasting plasma glucose (FPG), 2-hour plasma glucose (2hPG), fasting C-peptide (FC-pep), 2-hour C-peptide (2hC-pep), fasting insulin (FIns), and 2-hour insulin (2hIns). In addition, the morbidity of diabetic complications (retinopathy, neuropathy, and nephropathy) in the inpatient group was determined. Furthermore, the difference between 2hPG and FPG (ΔPG), the difference between 2hC-pep and FC-pep (ΔC-pep), and the difference between 2hIns and FIns (ΔIns) were calculated. The level of serum I-FABP, a biomarker of intestinal barrier (IB) dysfunction, was estimated by an enzyme-linked immunosorbent assay.ResultsFor the outpatient group, the median duration of hyperglycemia was less than a year; the serum I-FABP level was positively correlated with age (R = 0.299, P < 0.001). For the inpatient group, the median duration of hyperglycemia was ten years; correlation analysis showed that the serum I-FABP level was positively associated with age and ΔPG (R = 0.286, P = 0.001; R = 0.250, P = 0.006, respectively) while negatively associated with FC-pep and 2hC-pep (R =  − 0.304, P = 0.001; R =  − 0.241, P = 0.008, respectively); multiple linear regression analysis showed that the serum I-FABP level was positively associated with the duration of hyperglycemia (β = 0.362, P < 0.001); moreover, patients with retinopathy had a significantly higher I-FABP level than those without retinopathy (P = 0.001).ConclusionsIn the outpatients whose duration of hyperglycemia was less than a year, the serum I-FABP level was positively associated with age. In the inpatients with different courses of diabetes, the serum I-FABP level was positively associated with the duration of hyperglycemia and glycemic variability but negatively associated with islet beta-cell function; moreover, the serum I-FABP level was higher in patients with retinopathy than in those without retinopathy, suggesting that the IB dysfunction got worse with the progression of diabetes. creator: Yifeng Wang creator: Licheng Ding creator: Jiayue Yang creator: Lijun Liu creator: Liang Dong uri: https://doi.org/10.7717/peerj.10800 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Wang et al. title: Epigenetic identification of mitogen-activated protein kinase 10 as a functional tumor suppressor and clinical significance for hepatocellular carcinoma link: https://peerj.com/articles/10810 last-modified: 2021-02-02 description: BackgroundMitogen-activated protein kinase 10 (Mapk10) is a member of the c-jun N-terminal kinases (jnk) subgroup in the MAPK superfamily, and was proposed as a tumor suppressor inactivated epigenetically. Its role in hepatocellular carcinoma (HCC) has not yet been illustrated. We aimed to investigate the expression and epigenetic regulation of mapk10 as well as its clinical significance in HCC.ResultsMapk10 was expressed in almost all the normal tissues including liver, while we found that the protein expression of MAPK10 was significantly downregulated in clinical samples of HCC patients compared with these levels in adjacent normal tissues (29/46, P < 0.0001). Clinical significance of MAPK10 expression was then assessed in a cohort of 59 HCC cases, which indicated its negative expression was significantly correlated with advanced tumor stage (P = 0.001), more microsatellite nodules (P = 0.025), higher serum AFP (P = 0.001) and shorter overall survival time of HCC patients. Methylation was further detected in 58% of the HCC cell lines we tested and in 66% of primary HCC tissues by methylation-specific PCR (MSP), which was proved to be correlated with the silenced or downregulated expression of mapk10. To get the mechanisms more clear, the transcriptional silencing of mapk10 was reversed by pharmacological demethylation, and ectopic expression of mapk10 in silenced HCC cell lines significantly inhibited the colony formation ability, induced apoptosis, or enhanced the chemosensitivity of HCC cells to 5-fluorouracil.ConclusionMapk10 appears to be a functional tumor suppressor gene frequently methylated in HCC, which could be a valuable biomarker or a new diagnosis and therapy target in a clinical setting. creator: Liping Tang creator: Shasha Zhu creator: Weiyan Peng creator: Xuedong Yin creator: Cui Tan creator: Yaying Yang uri: https://doi.org/10.7717/peerj.10810 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Tang et al. title: Comparison of physical fitness and mental health status among school-age children with different sport-specific training frequencies link: https://peerj.com/articles/10842 last-modified: 2021-02-02 description: This cross-sectional study compared the physical fitness and mental health status of 140 school-age children who participated in sport-specific training with 180 age-matched peers. All the participants were grouped by sport-specific training frequencies in extracurricular time into the following: (i) high sports training frequency group (HFG): training three to five times per week (n = 77, mean [SD] age: 9.60 [0.12] years); (ii) low sports training frequency group (LFG): training once per week (n = 63, mean [SD] age: 9.88 [0.14] years); and (iii) control group (CG): maintaining routine life (n = 180, mean (SD) age: 9.77(0.09) years). Physical fitness status, including body composition (body mass index), endurance (vital capacity; 50 × 8 round trip), speed and agility (50 m sprint), flexibility (sit-and-reach), coordination (1-min rope skipping), and core strength (1-min sit-ups) as well as mental health status was measured. Overall, the results showed that Grade 3 to 4 HFG students showed better total physical fitness scores than the LFG and CG students. Grade 2 and 5 participants in the three groups showed no significant difference in the total physical fitness score. Children in HFG performed better in several PF indicators (i.e., cardiopulmonary function, flexibility, core strength, and coordination) than those in LFG and CG, and children in LFG got a higher score than those in CG on a testing item of 1-min rope skipping. The mental health test results showed that HFG performed better than LFG and CG. The results indicated that participating in sport-specific training 3–5 times per week was beneficial for children’s physical and mental health. Additionally, there was a weak and negative correlation between physical fitness and mental health in LFG and CG, while no correlation was found between physical fitness and mental health in HFG. creator: Ruichen Jiang creator: Chun Xie creator: Jilong Shi creator: Xuechen Mao creator: Qin Huang creator: Fanying Meng creator: Zhiguang Ji creator: Anmin Li creator: Chunhua Zhang uri: https://doi.org/10.7717/peerj.10842 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Jiang et al. title: Undiagnosed cardiovascular risk factors in overweight and obese individuals: a low income country experience link: https://peerj.com/articles/10870 last-modified: 2021-02-02 description: BackgroundOverweight and obesity are associated with diabetes, hypertension and chronic kidney disease (CKD). However, there is scarce information from lower income countries about undiagnosed obesity-associated conditions. This information is necessary for healthcare planning and for assessment of Global Burden of Disease.MethodsWe assessed the prevalence of obesity-associated conditions in 656 overweight (n = 360) and obese (n = 296) adults from inner-city Portoviejo (Ecuador), in descriptive field research, based on an opportunistic and selective sampling strategy.ResultsOf 316 men and 340 women, 73% met criteria for prehypertension (27%) or hypertension (46%), 50% met criteria for prediabetes (30%) or diabetes (20%), 11% had an estimated glomerular filtration rate (eGFR) <60 ml/min/1.73 m2 consistent with chronic kidney disease (CKD) and 5.5% had pathological albuminuria for a total CKD prevalence of 16%. Age-related prevalence data were generated. In all participants, serum total cholesterol and triglycerides were >200 and >150 mg/dl, respectively. Hyperuricemia and microhematuria (<2%) were uncommon. Women were more likely to have low eGFR (18 vs 5%, p 0.000). Diabetes and pathological albuminuria prevalence were higher in obese than in overweight participants (15 vs 12%, p 0.018; and 8 vs 4%, p 0.0199, respectively).DiscussionIn conclusion, undiagnosed hypertension, diabetes and CKD were more common than expected in overweight and obese persons from Ecuador. Detection rates exceeded official estimates of prevalene of these conditions. Screening the overweight/obese for these conditions, especially at the age ranges at higher risk, may be cost-effective to identify a high number of persons who may benefit from early inexpensive intervention. creator: Patricio Alfredo Vallejo-Valdivieso creator: Graciela Zambrano-Pincay creator: Alberto Ortiz uri: https://doi.org/10.7717/peerj.10870 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Vallejo-Valdivieso et al. title: The long non-coding RNA GHSROS reprograms prostate cancer cell lines toward a more aggressive phenotype link: https://peerj.com/articles/10280 last-modified: 2021-02-01 description: It is now appreciated that long non-coding RNAs (lncRNAs) are important players in orchestrating cancer progression. In this study we characterized GHSROS, a human lncRNA gene on the opposite DNA strand (antisense) to the ghrelin receptor gene, in prostate cancer. The lncRNA was upregulated by prostate tumors from different clinical datasets. Transcriptome data revealed that GHSROS alters the expression of cancer-associated genes. Functional analyses in vitro showed that GHSROS mediates tumor growth, migration and survival, and resistance to the cytotoxic drug docetaxel. Increased cellular proliferation of GHSROS-overexpressing PC3, DU145, and LNCaP prostate cancer cell lines in vitro was recapitulated in a subcutaneous xenograft model. Conversely, in vitro antisense oligonucleotide inhibition of the lncRNA reciprocally regulated cell growth and migration, and gene expression. Notably, GHSROS modulates the expression of PPP2R2C, the loss of which may drive androgen receptor pathway-independent prostate tumor progression in a subset of prostate cancers. Collectively, our findings suggest that GHSROS can reprogram prostate cancer cells toward a more aggressive phenotype and that this lncRNA may represent a potential therapeutic target. creator: Patrick B. Thomas creator: Penny Jeffery creator: Manuel D. Gahete creator: Eliza Whiteside creator: Carina Walpole creator: Michelle Maugham creator: Lidija Jovanovic creator: Jennifer Gunter creator: Elizabeth Williams creator: Colleen Nelson creator: Adrian Herington creator: Raul M. Luque creator: Rakesh Veedu creator: Lisa K. Chopin creator: Inge Seim uri: https://doi.org/10.7717/peerj.10280 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Thomas et al. title: Seasonal patterns in nest survival of a subtropical wading bird, the Hawaiian Stilt (Himantopus mexicanus knudseni) link: https://peerj.com/articles/10399 last-modified: 2021-02-01 description: Nest survival is influenced by where and when birds decide to breed. For ground-nesting species, nest-site characteristics, such as vegetation height and proximity to water, may impact the likelihood of nest flooding or depredation. Further, habitat characteristics, and thus nest survival, may fluctuate across the breeding season. The Hawaiian Stilt (‘Ae‘o; Himantopus mexicanus knudseni) is an endangered Hawaiian waterbird that nests in wetlands across the Hawaiian Islands. In this study, we used observational surveys and nest cameras to examine the impact of nest-site characteristics and day of nesting season on nest survival of the Hawaiian Stilt. Early nests had a higher chance of survival than late nests. For most of the nesting season, taller vegetation was correlated with increased nest survival, while shorter vegetation was correlated with increased nest survival late in the nesting season. Seasonal patterns in nest survival may be due to changes in parental behavior or predator activity. Nest depredation was responsible for 55% of confirmed nest failures and introduced mammals were the primary nest predators. Our study is the first to examine seasonality in nest survival of Hawaiian Stilts and suggests that, despite longer nesting seasons and year-round occupation of wetlands, late nesters in subtropical regions may have lower nest survival than early nesters, similar to trends observed in temperate regions. creator: Kristen C. Harmon creator: Nathaniel H. Wehr creator: Melissa R. Price uri: https://doi.org/10.7717/peerj.10399 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Harmon et al. title: Effect of ultra-processed foods consumption on glycemic control and gestational weight gain in pregnant with pregestational diabetes mellitus using carbohydrate counting link: https://peerj.com/articles/10514 last-modified: 2021-02-01 description: AimsThe aims were to evaluate the consumption of ultra-processed foods by pregnant women with pre-existing diabetes mellitus (DM) using the carbohydrate counting method, in addition to investigating the association with total gestational weight gain and glycemic control.MethodsA cohort study of adult Brazilian pregnant women with pre-existing DM. Dietary intake was evaluated adopting the NOVA classification to identify the reported consumption of ultra-processed foods. Weight was measured at all consultations and laboratory tests were evaluated at each gestational trimester. Multivariate linear regression was used in the analysis.ResultsPregnant women (n = 42) presented mean total gestational weight gain of 12.02 ± 4.8 kg, 65.8% of them with inadequate weight gain. Daily consumption of ultra-processed foods was 272.37 ± 170.55 kcal. The increase of every 1 kcal in the calorie intake from ultra-processed foods in the third trimester increased glycated hemoglobin by 0.007% (β = 0.007, p = 0.025), raised 1-h postprandial glucose by 0.14 mg/dL (β = 0.143, p = 0.011), and added 0.11 kg to total gestational weight gain (β = 0.11, p = 0.006).ConclusionUltra-processed food consumption influenced glycemic control and total gestational weight gain in pregnant women with DM. creator: Carolina F. M. Silva creator: Claudia Saunders creator: Wilza Peres creator: Bárbara Folino creator: Taiana Kamel creator: Mayara Silva dos Santos creator: Patrícia Padilha uri: https://doi.org/10.7717/peerj.10514 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2021 Silva et al. title: Night of the hunter: using cameras to quantify nocturnal activity in desert spiders link: https://peerj.com/articles/10684 last-modified: 2021-02-01 description: Invertebrates dominate the animal world in terms of abundance, diversity and biomass, and play critical roles in maintaining ecosystem function. Despite their obvious importance, disproportionate research attention remains focused on vertebrates, with knowledge and understanding of invertebrate ecology still lacking. Due to their inherent advantages, usage of camera traps in ecology has risen dramatically over the last three decades, especially for research on mammals. However, few studies have used cameras to reliably detect fauna such as invertebrates or used cameras to examine specific aspects of invertebrate ecology. Previous research investigating the interaction between wolf spiders (Lycosidae: Lycosa spp.) and the lesser hairy-footed dunnart (Sminthopsis youngsoni) found that camera traps provide a viable method for examining temporal activity patterns and interactions between these species. Here, we re-examine lycosid activity to determine whether these patterns vary with different environmental conditions, specifically between burned and unburned habitats and the crests and bases of sand dunes, and whether cameras are able to detect other invertebrate fauna. Twenty-four cameras were deployed over a 3-month period in an arid region in central Australia, capturing 2,356 confirmed images of seven invertebrate taxa, including 155 time-lapse images of lycosids. Overall, there was no clear difference in temporal activity with respect to dune position or fire history, but twice as many lycosids were detected in unburned compared to burned areas. Despite some limitations, camera traps appear to have considerable utility as a tool for determining the diel activity patterns and habitat use of larger arthropods such as wolf spiders, and we recommend greater uptake in their usage in future. creator: Tamara I. Potter creator: Aaron C. Greenville creator: Christopher R. Dickman uri: https://doi.org/10.7717/peerj.10684 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Potter et al. title: TLR4 promoter rs1927914 variant contributes to the susceptibility of esophageal squamous cell carcinoma in the Chinese population link: https://peerj.com/articles/10754 last-modified: 2021-02-01 description: BackgroundToll-like receptor 4 (TLR4), as a key regulator of both innate and acquired immunity, has been linked with the development of various cancers, including esophageal cancer. This study aims to analyze the association of potential functional genetic polymorphisms in TLR4 with the risk of esophageal cancer.MethodsThis case-control study involved in 480 ESCC patients and 480 health controls. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to genotype TLR4 rs1927914 polymorphism. Taqman probe method was used to determine the genotypes of TLR4 rs11536891 and rs7873784 variants. The relationship between TLR4 genetic variation and ESCC risk was analyzed by Logistic regression model by calculating the odds ratio (OR) and 95% confidence interval (95% CI).ResultsCompared with TLR4 rs1927914 AA genotype carriers, GG carriers had a lower ESCC risk (OR = 0.59, 95% CI [0.38–0.93], P = 0.023). Stratification analysis by age showed that TLR4 rs1927914 GG could affect the risk of ESCC in elderly people (OR = 0.59, 95% CI [0.36–0.97]). Smoking stratification analysis indicated that rs1927914 GG carriers were related to ESCC susceptibility among non-smokers (OR = 0.36, 95% CI [0.18–0.73]). Dual luciferase reporter assay suggested that rs1927914 G-containing TLR4 promoter displayed a 1.76-fold higher luciferase activity than rs1927914 A-containing counterpart in KYSE30 cells. Electrophoretic mobility shift assay (EMSA) showed the KYSE30 cell nuclear extract was able to bind the probe with rs1927914 G allele and this DNA-protein interaction could be eliminated by competition assays with unlabeled rs1927914 G probe, which indicating that the binding is sequence-specific. Our results also showed that TLR4 rs7873784 (G>C) and rs11536891 (T>C) conformed to complete genetic linkage. The genotype distributions of TLR4 rs11536891 variant among ESCC patients and normal controls have no statistical significance.ConclusionThe TLR4 rs1927914 variant contributes to the ESCC risk by effecting the promoter activity. creator: Jiaying Li creator: Hongjiao Wu creator: Hui Gao creator: Ruihuan Kou creator: Yuning Xie creator: Zhi Zhang creator: Xuemei Zhang uri: https://doi.org/10.7717/peerj.10754 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Li et al. title: Unexpected mitochondrial lineage diversity within the genus Alonella Sars, 1862 (Crustacea: Cladocera) across the Northern Hemisphere link: https://peerj.com/articles/10804 last-modified: 2021-02-01 description: Representatives of the genus Alonella Sars (Crustacea: Cladocera: Chydorinae) belong to the smallest known water fleas. Although species of Alonella are widely distributed and often abundant in acidic and mountain water bodies, their diversity is poorly studied. Morphological and genetic approaches have been complicated by the minute size of these microcrustaceans. As a result, taxonomists have avoided revising these species. Here, we present genetic data on Alonella species diversity across the Northern Hemisphere with particular attention to the A. excisa species complex. We analyzed 82 16S rRNA sequences (all newly obtained), and 78 COI sequences (39 were newly obtained). The results revealed at least twelve divergent phylogenetic lineages, possible cryptic species, of Alonella, with different distribution patterns. As expected, the potential species diversity of this genus is significantly higher than traditionally accepted. The A. excisa complex is represented by nine divergent clades in the Northern Hemisphere, some of them have relatively broad distribution ranges and others are more locally distributed. Our results provide a genetic background for subsequent morphological analyses, formal descriptions of Alonella species and detailed phylogeographical studies. creator: Anna N. Neretina creator: Dmitry P. Karabanov creator: Veronika Sacherova creator: Alexey A. Kotov uri: https://doi.org/10.7717/peerj.10804 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2021 Neretina et al.