title: PeerJ description: Articles published in PeerJ link: https://peerj.com/articles/index.rss3?journal=peerj&month=2020-05 creator: info@peerj.com PeerJ errorsTo: info@peerj.com PeerJ language: en title: Evaluation of leaf rust resistance in the Chinese wheat cultivar ‘Een1’ link: https://peerj.com/articles/8993 last-modified: 2020-05-29 description: Wheat cultivar Een1, 34 near isogenic lines (NILs), and two cultivars were used as plant materials to evaluate the resistance of Een1 to leaf rust disease. Infection type identification and gene postulation were carried out by inoculation of 12 Chinese Puccinia triticina (Pt) pathotypes. Based on the unique phenotype of Een1, we speculated that Een1 might carry Lr gene(s) different from the tested ones. The chromosomal locations for resistance gene to leaf rust disease was employed using SSR primers mapping the populations derived from the cross between Een1 and susceptible Thatcher. A total of 285 plants in the F2 population were tested by inoculating Pt pathotype FHNQ during the seedling stage. Results from the segregation analysis fits a ratio of 3:1 (${\chi }_{3:1}^{2}=2.37$χ3:12=2.37, P = 0.12), indicating the presence of a single dominant gene in Een1 conferring resistance to FHNQ. A total of 1,255 simple sequence repeat (SSR) primers were first used to identify the likely linked markers based on bulk segregation analysis (BSA), and then those likely linked markers were further genotyped in the F2 population for linkage analysis. Our linkage analysis found that the resistance gene (LrE1) was distal to seven SSR loci on the long arm of chromosome 7B, with distances from 2.6 cM (Xgwm344) to 27.1 cM (Xgwm131). The closest marker Xgwm344 was further verified with F3 lines. creator: Na Zhang creator: Lina Zhao creator: Kahsay Tadesse Mawcha creator: Chenguang Zhao creator: Wenxiang Yang creator: Daqun Liu uri: https://doi.org/10.7717/peerj.8993 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: Ancient genetic divergence in bumblebee catfish of the genus Pseudopimelodus (Pseudopimelodidae: Siluriformes) from northwestern South America link: https://peerj.com/articles/9028 last-modified: 2020-05-29 description: Pseudopimelodus is a Neotropical genus of bumblebee catfish, composed of four valid species occurring in both trans- and cis-Andean rivers of South America. The orogeny of the Andes has led to diversification in the genus Pseudopimelodus in Colombia. This study analyzed partial sequences of mitochondrial cox1 and nuclear rag2 genes to test the hypothesis that the species, nominally recognized as P. schultzi and P. bufonius in Colombia, correspond to more than two different evolutionary lineages. Results indicate high levels of genetic divergence among individuals of nominal P. schultzi and P. bufonius, from trans- and cis-Andean basins in Colombia. In addition, five divergent lineages of Pseudopimelodus were confidently delimited by using a single-locus species-discovery approach and confirmed by species tree analyses. Additionally, molecular-clock dating showed that most diversification processes in Pseudopimelodus took place during the Miocene, when Andean tectonic evolution was occurring in northwestern South America. The present study provides, for the first time, phylogeographic insight into this Neotropical genus. creator: José D. Rangel-Medrano creator: Armando Ortega-Lara creator: Edna J. Márquez uri: https://doi.org/10.7717/peerj.9028 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Rangel-Medrano et al. title: Meiofaunal communities and nematode diversity characterizing the Secca delle Fumose shallow vent area (Gulf of Naples, Italy) link: https://peerj.com/articles/9058 last-modified: 2020-05-29 description: Hydrothermal venting is rather prevalent in many marine areas around the world, and marine shallow vents are relatively abundant in the Mediterranean Sea, especially around Italy. However, investigations focusing on the characterization of meiofaunal organisms inhabiting shallow vent sediments are still scant compared to that on macrofauna. In the present study, we investigated the meiobenthic assemblages and nematode diversity inhabiting the Secca delle Fumose (SdF), a shallow water vent area located in the Gulf of Naples (Italy). In this area, characterized by a rapid change in the environmental conditions on a relative small spatial scale (i.e., 100 m), we selected four sampling sites: one diffusive emission site (H); one geyser site (G) and two inactive sites (CN, CS). Total meiofauna abundance did not vary significantly between active and inactive sites and between surface and deeper sediment layers due to a high inter-replicate variability, suggesting a pronounced spatial-scale patchiness in distribution of meiofauna. Nematofauna at site H presented the typical features of deep-sea vents with low structural and functional diversity, high biomass and dominance of few genera (i.e., Oncholaimus; Daptonema) while from site G we reported diversity values comparable to that of the inactive sites. We hypothesized that site G presented a condition of “intermediate disturbance” that could maintain a high nematode diversity. Environmental features such as sediment temperature, pH, total organic carbon and interstitial waters ions were found to be key factors influencing patterns of meiofauna and nematofauna assemblages at SdF. Even though the general theory is that nematodes inhabiting shallow vent areas include a subset of species that live in background sediments, this was not the case for SdF vent area. Due to a marked change in nematode composition between all sites and to the presence of many exclusive species, every single investigated site was characterized by a distinct nematofauna reflecting the high spatial heterogeneity of SdF. creator: Elisa Baldrighi creator: Daniela Zeppilli creator: Luca Appolloni creator: Luigia Donnarumma creator: Elena Chianese creator: Giovanni Fulvio Russo creator: Roberto Sandulli uri: https://doi.org/10.7717/peerj.9058 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Baldrighi et al. title: Mountain goat survival and mortality during a period of increased puma abundance in the Black Hills, South Dakota link: https://peerj.com/articles/9143 last-modified: 2020-05-29 description: We investigated survival and cause-specific mortality for a mountain goat (Oreamnos americanus) population during a period when the puma (Puma concolor) population was growing in the Black Hills, South Dakota, 2006–2018. We obtained survival data from 47 adult goats (n = 33 females, n = 14 males). Annual survival varied from 0.538 (95% CI [0.285–0.773]) to 1.00 (95% CI [1.00–1.00]) and puma predation was the primary cause-specific mortality factor over a 12-year period. Cumulative hectares of mountain pine beetle (Dendroctonus ponderosae) disturbance was a covariate of importance (wi = 0.972; β = 0.580, 95% CI [0.302–0.859]) influencing survival. To our knowledge, this is the first account of puma being the primary mortality factor of mountain goats over a long-term study. The Black Hills system is unique because we could examine the expanded realized niche of puma in the absence of other large carnivores and their influence on mountain goats. We hypothesize that puma were being sustained at higher densities due to alternate prey sources (e.g., white-tailed deer; Odocoileous virginianus) and this small population of mountain goats was susceptible to predation by one or several specialized puma in the Black Hills. However, we also hypothesize a changing landscape with increased tree mortality due to insect infestation provided conditions for better predator detection by goats and increased survival. Alternatively, open canopy conditions may have increased understory forage production potentially increasing mountain goat survival but we did not evaluate this relationship. Survival and mortality rates of mountain goats should continue to be monitored as this small population may be highly susceptible to population declines due to slow growth rates. creator: Chadwick P. Lehman creator: Eric M. Rominger creator: Brady Y. Neiles uri: https://doi.org/10.7717/peerj.9143 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Lehman et al. title: Proliferating activity in a bryozoan lophophore link: https://peerj.com/articles/9179 last-modified: 2020-05-29 description: Bryozoans are small benthic colonial animals; their colonies consist of zooids which are composed of a cystid and polypide. According to morphological and molecular data, three classes of bryozoans are recognized: Phylactolaemata, Gymnolaemata and Stenolaemata. Bryozoans are active suspension feeders and their feeding apparatus, the lophophore, is fringed with a single row of ciliated tentacles. In gymnolaemates, the lophophore is bell-shaped and its tentacles may be equal in length (equitentacled lophophores) or some tentacles may be longer than others (obliquely truncated lophophores). In encrusting colonies, polypides with obliquely truncated lophophores usually border specific sites of excurrent water outlets (colony periphery and chimneys) where depleted water has to be removed. It is known that during colony astogeny, colony-wide water currents rearrange: new chimneys are formed and/or location of the chimneys within a given colony changes with time. Such rearrangement requires remodeling of the lophophore shape and lengthening of some tentacles in polypides surrounding water outlets. However, proliferating activity has not been described for bryozoans. Here, we compared the distribution of S-phase and mitotic cells in young and adult polypides in three species of Gymnolaemata. We tested the hypothesis that tentacle growth/elongation is intercalary and cell proliferation takes place somewhere at the lophophore base because such pattern does not interfere with the feeding process. We also present a detailed description of ultrastructure of two parts of the lophophore base: the oral region and ciliated pits, and uncover the possible function of the latter. The presence of stem cells within the ciliated pits and the oral region of polypides provide evidence that both sites participate in tentacle elongation. This confirms the suggested hypothesis about intercalary tentacle growth which provides a potential to alter a lophophore shape in adult polypides according to rearrangement of colony wide water currents during colony astogeny. For the first time deuterosome-like structures were revealed during kinetosome biogenesis in the prospective multiciliated epithelial cells in invertebrates. Tentacle regeneration experiments in Electra pilosa demonstrated that among all epidermal cell types, only non-ciliated cells at the abfrontal tentacle surface are responsible for wound healing. Ciliated cells on the frontal and lateral tentacle surfaces are specialized and unable to proliferate, not even under wound healing. Tentacle regeneration in E. pilosa is very slow and similar to the morphallaxis type. We suggest that damaged tentacles recover their length by a mechanism similar to normal growth, powered by proliferation of cells both within ciliated pits and the oral region. creator: Natalia Shunatova creator: Ilya Borisenko uri: https://doi.org/10.7717/peerj.9179 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Shunatova and Borisenko title: Future sea-level rise drives rocky intertidal habitat loss and benthic community change link: https://peerj.com/articles/9186 last-modified: 2020-05-29 description: The impacts of sea-level rise (SLR) are likely to be the greatest for ecosystems that exist at the land-sea interface, where small changes in sea-level could result in drastic changes in habitat availability. Rocky intertidal ecosystems possess a number of characteristics which make them highly vulnerable to changes in sea-level, yet our understanding of potential community-scale responses to future SLR scenarios is limited. Combining remote-sensing with in-situ large-area imaging, we quantified habitat extent and characterized the biological community at two rocky intertidal study locations in California, USA. We then used a model-based approach to estimate how a range of SLR scenarios would affect total habitat area, areal extent of dominant benthic space occupiers, and numerical abundance of invertebrates. Our results suggest that SLR will reduce total available rocky intertidal habitat area at our study locations, leading to an overall decrease in areal extent of dominant benthic space occupiers, and a reduction in invertebrate abundances. As large-scale environmental changes, such as SLR, accelerate in the next century, more extensive spatially explicit monitoring at ecologically relevant scales will be needed to visualize and quantify their impacts to biological systems. creator: Nikolas J. Kaplanis creator: Clinton B. Edwards creator: Yoan Eynaud creator: Jennifer E. Smith uri: https://doi.org/10.7717/peerj.9186 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kaplanis et al. title: Recent advances in the contribution of noncoding RNAs to cisplatin resistance in cervical cancer link: https://peerj.com/articles/9234 last-modified: 2020-05-29 description: Cervical cancer (CC) remains a major disease burden on the female population worldwide. Chemotherapy with cisplatin (cis-diamminedichloroplatinum (II); CDDP) and related drugs are the main treatment option for CC; however, their efficacy is limited by the development of drug resistance. Noncoding RNAs (ncRNAs) have been found to play critical roles in numerous physiological and pathological cellular processes, including drug resistance of cancer cells. In this review, we describe some of the ncRNAs, including miRNAs, lncRNAs and circRNAs, that are involved in the sensitivity/resistance of CC to CDDP-based chemotherapy and discuss their mechanisms of action. We also describe some ncRNAs that could be therapeutic targets to improve the sensitivity of CC to CDDP-based chemotherapy. creator: Xin Wen creator: Shui Liu creator: Jiyao Sheng creator: Manhua Cui uri: https://doi.org/10.7717/peerj.9234 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wen et al. title: Decreased inhibitory control after partial sleep deprivation in individuals reporting binge eating: preliminary findings link: https://peerj.com/articles/9252 last-modified: 2020-05-29 description: BackgroundPoor executive functions are associated with dysregulated eating and greater caloric intake in healthy samples. In parallel, findings suggested that sleep deprivation impairs executive functions.MethodsWe investigated whether partial sleep deprivation impairs executive functions in individuals reporting binge eating (BE, N = 14) and healthy controls (C, N = 13). Switch cost and backward inhibition were measured using the Task Switching Paradigm after a habitual night of sleep and after a night of partial sleep deprivation.ResultsResults showed a Night by Group interaction on the backward inhibition. The two groups differed in the habitual night, evidencing higher inhibitory control in BE compared to C. Additionally, after partial sleep deprivation, compared to the habitual night, backward inhibition decreased in BE group. This preliminary study was the first to explore the impact of sleep deprivation on executive functions in participants reporting binge eating and healthy controls, thus highlighting their potential role in influencing eating behavior. creator: Silvia Cerolini creator: Andrea Ballesio creator: Fabio Ferlazzo creator: Fabio Lucidi creator: Caterina Lombardo uri: https://doi.org/10.7717/peerj.9252 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Cerolini et al. title: Uncovering the species diversity of subterranean rodents at the end of the World: three new species of Patagonian tuco-tucos (Rodentia, Hystricomorpha, Ctenomys) link: https://peerj.com/articles/9259 last-modified: 2020-05-29 description: Ctenomys Blainville 1826 is one of the most diverse genera of South American caviomorph rodents. Currently, six species of this genus are reported from Patagonia, south of 42°S. In this contribution, we assessed the taxonomic status of several populations from eastern and central Chubut province, northern Patagonia. Based on phylogenetic analyses of DNA sequences, morphology assessment (qualitative and quantitative), and previously published karyological data, we describe three new species of this genus, one formed by two subspecies, endemic to northern Patagonia. In addition, we include C. coyhaiquensis Kelt and Gallardo 1994 into the synonymy of C. sericeus J.A. Allen 1903. Finally, we discussed the need for additional integrative approaches, including field collection of specimens, to better understand the diversity of this highly speciose rodent genus. creator: Pablo Teta creator: Guillermo D’Elía uri: https://doi.org/10.7717/peerj.9259 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Teta et al. title: Isotopic biomonitoring of anthropic carbon emissions in a megalopolis link: https://peerj.com/articles/9283 last-modified: 2020-05-29 description: Atmospheric pollution has become a serious threat for human health and the environment. However, the deployment, operation and maintenance of monitoring networks can represent a high cost for local governments. In certain locations, the use of naturally occurring plants for monitoring pollution can be a useful supplement of existing monitoring networks, and even provide information when other types of monitoring are lacking. In this work, we (i) determined the tissue carbon content and the δ13C values for the epiphytic CAM bromeliad Tillandsia recurvata and the relationship of both parameters with the existing CO concentrations in the Valley of Mexico basin and (ii) mapped the spatial distribution of such elemental and isotopic composition for this plant within the basin, in order to assess its potential as an atmospheric biomonitor of carbon monoxide, a pollutant with important repercussions on public health. The CO concentrations in the basin ranged from 0.41 ppm at rural locations to 0.81 ppm at urban sites. The carbon content of T. recurvata, which averaged 42.9 ± 0.34% (dry weight), was not influenced by the surrounding CO concentration. In contrast, the δ13C depended on the sites where the plants were collected. For example, the values were −13.2‰ in rural areas and as low as –17.5‰ in an urban site. Indeed, the isotopic values had a positive linear relationship with the atmospheric CO concentrations. Given the close relationship observed between the isotopic composition of T. recurvata with the CO concentrations in the Valley of Mexico, the δ13C values can be useful for the detection of atmospheric carbonaceous emissions. creator: Edison A. Díaz-Álvarez creator: Erick de la Barrera uri: https://doi.org/10.7717/peerj.9283 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Díaz-Álvarez and de la Barrera title: Farming by soil in Europe: status and outlook of cropping systems under different pedoclimatic conditions link: https://peerj.com/articles/8984 last-modified: 2020-05-28 description: BackgroundDespite of the importance of soils in agronomy, to date no comprehensive assessment of cropping in Europe has been performed from the viewpoint of the soil variability and its relationship to cropping patterns. In order to fill this knowledge gap, we studied the cropping patterns in different soils of European climate zones with regards to the shares of their crop types in a comparative manner. The study highlights the main features of farming by soil in Europe. Farming by soil in this context means the consideration of soil characteristics when selecting crop types and cropping patterns.MethodsWe first assessed the dissimilarity between the cropping compositions of different pedoclimatic zones in Europe. Next, we assessed the differences of crop distribution in the climate zones by soil types and main crop types by analyzing the degree of association of crops to soil types. A detailed country scale assessment was performed using crops-specific soil productivity maps and land use survey data from Hungary.ResultsResults suggest that, in general, farmers consciously take pedoclimatic condition of farming into account when selecting their cropping patterns. In other words, farming by soil is a common practice in the different climatic regions of Europe. However, we have strong reasons to believe that soil suitability-based cropping is not practiced to its full potential over the continent. For example, the findings of our European assessment suggest that production areas of legumes are not always optimized for the local pedoclimatic conditions in some zones. These findings also underline that economic drivers are decisive, when farmers adopt their cropping (eg. oil crops on Albeluvsiols in Europe). Win-win situations of economic considerations and soil suitability based management are observed in all pedoclimatic zones of Europe. The country analysis shows that cropping is progressively practiced on more suitable areas, depending also the crop tolerance to variable pedoclimatic conditions In conclusion, we can assume that pedoclimatic conditions of cropping are respected in most of Europe and farmers crops according to edaphic conditions whenever economic considerations do not override the ecological concerns of farming. creator: Gergely Tóth creator: Tamás Kismányoky creator: Piroska Kassai creator: Tamás Hermann creator: Oihane Fernandez-Ugalde creator: Brigitta Szabó uri: https://doi.org/10.7717/peerj.8984 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Tóth et al. title: Characterization of biofilm production in different strains of Acinetobacter baumannii and the effects of chemical compounds on biofilm formation link: https://peerj.com/articles/9020 last-modified: 2020-05-28 description: Acinetobacter baumannii, an important emerging pathogen of nosocomial infections, is known for its ability to form biofilms. Biofilm formation increases the survival rate of A. baumannii on dry surfaces and may contribute to its persistence in the hospital environment, which increases the probability of nosocomial infections and outbreaks. This study was undertaken to characterize the biofilm production of different strains of A. baumannii and the effects of chemical compounds, especially antibiotics, on biofilm formation. In this study, no statistically significant relationship was observed between the ability to form a biofilm and the antimicrobial susceptibility of the A. baumannii clinical isolates. Biofilm formation caused by A. baumannii ATCC 17978 after gene knockout of two-component regulatory system gene baeR, efflux pump genes emrA/emrB and outer membrane coding gene ompA revealed that all mutant strains had less biofilm formation than the wild-type strain, which was further supported by the images from scanning electron microscopy and confocal laser scanning microscopy. The addition of amikacin, colistin, LL-37 or tannic acid decreased the biofilm formation ability of A. baumannii. In contrast, the addition of lower subinhibitory concentration tigecycline increased the biofilm formation ability of A. baumannii. Minimum biofilm eradication concentrations of amikacin, imipenem, colistin, and tigecycline were increased obviously for both wild type and multidrug resistant clinical strain A. baumannii VGH2. In conclusion, the biofilm formation ability of A. baumannii varied in different strains, involved many genes and could be influenced by many chemical compounds. creator: Ming-Feng Lin creator: Yun-You Lin creator: Chung-Yu Lan uri: https://doi.org/10.7717/peerj.9020 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Lin et al. title: The tetrapod fauna of the upper Permian Naobaogou Formation of China: 5. Caodeyao liuyufengi gen. et sp. nov., a new peculiar therocephalian link: https://peerj.com/articles/9160 last-modified: 2020-05-28 description: The upper Permian Naobaogou Formation has been the goal of recent contributions that notably increased the knowledge of its terrestrial vertebrate fauna and unravelled a hidden late Permian therocephalian diversity in China. Two very different species of therocephalians have been documented in the Naobaogou Formation and they were recovered as basal akidnognathids in cladistic analyses. In this contribution we describe Caodeyao liuyufengi gen. et sp. nov., represented by a partial skull and mandible, and a humerus. The new taxon features a short, high snout and a wide temporal opening with the coronoid process of the mandible separated by a wide space medially to the zygomatic arch. The latter feature is only recognized in the Russian therocephalian Purlovia maxima and it is also characteristic of non-mammaliaform cynodonts. Phylogenetic analysis indicates a close relationship of the new Chinese taxon with Purlovia maxima, producing a monophyletic Laurasian group in therocephalian phylogeny. With the representation of three different species, the Naobaogou Formation is now the most prolific unit documenting therocephalian late Permian diversity in China. creator: Jun Liu creator: Fernando Abdala uri: https://doi.org/10.7717/peerj.9160 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liu and Abdala title: The impact of smog on the concentration of particulate matter in the antelope house in the Silesian zoological garden link: https://peerj.com/articles/9191 last-modified: 2020-05-28 description: Persistent negligence in the field of environmental protection in Poland as well as strong dependance of the energy sector on the fossil fuels have led to serious pollution of the air with particulate matter, which at high concentrations is capable of penetrating into the buildings. The aim of this study is to assess the impact of particulate airborne pollution on the concentration of particulate matter inside the antelope house in the Silesian Zoological Garden located within the Upper Silesia in Poland. The research was conducted from February to May in 2018. The records taken in the research period show that the concentration of PM10 exceeded the level of 50 µg/m3 outside the building during 26 days and 11 days when it comes to the concentration of particulate matter inside the antelope house. The quantity of particulate matter in the antelope house is strongly correlated with the concentration of the particles in the air. Despite fitting existing ventilation system with a filter that reduces the dust level by 60% during the highest level of smog, particulate matter concentration in the antelope house exceeded acceptable limit for PM10 more than twofold. Particle size-fraction analysis revealed that as much as 85% of the particles detected in the studied compartment constitute PM2.5. creator: Krzysztof Pawlak creator: Zenon Nieckarz uri: https://doi.org/10.7717/peerj.9191 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Pawlak and Nieckarz title: Development and initial validation of the Psychological Need Frustration Scale for Physical Activity link: https://peerj.com/articles/9210 last-modified: 2020-05-28 description: BackgroundThe frustration of basic psychological needs can be detrimental to people’s health. To date, a scale developed specifically for measuring such perceived negative experiences, derived from a need thwarting environment in the physical activity context, is lacking. The present research attempted to develop and validate the Psychological Need Frustration Scale for Physical Activity (PNFS-PA) grounded in self-determination theory via multiple studies.MethodIn Study 1, an item pool was created, and its face and content validity were established. In Study 2, the factor structure of the scale was demonstrated using exploratory structural equation modelling (ESEM). In Study 3, its factor structure was cross-validated. Also, the nomological validity, reliability and measurement invariance of the scale were established.ResultTaken together, the research suggests the newly developed PNFS-PA is valid and reliable and can be applied to assess psychological needs frustration experiences in the physical activity context. creator: Pak-Kwong Chung creator: Tao Zhong creator: Jing-Dong Liu creator: Chun-Qing Zhang creator: Ming Yu Claudia Wong uri: https://doi.org/10.7717/peerj.9210 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chung et al. title: Consequences of removal of exotic species (eucalyptus) on carbon and nitrogen cycles in the soil-plant system in a secondary tropical Atlantic forest in Brazil with a dual-isotope approach link: https://peerj.com/articles/9222 last-modified: 2020-05-28 description: The impact of exotic species on heterogeneous native tropical forest requires the understanding on which temporal and spatial scales these processes take place. Functional tracers such as carbon (δ13C) and nitrogen (δ15N) isotopic composition in the soil-plant system might help track the alterations induced by the exotic species. Thus, we assess the effects from the removal of the exotic species eucalyptus (Corymbia cytriodora) in an Atlantic forest Reserve, and eucalyptus removal on the alteration of the nutrient dynamics (carbon and nitrogen). The hypotheses were: (1) the eucalyptus permanence time altered δ13C and δ15N in leaves, soils and litter fractions (leaves, wood, flowers + fruits, and rest); and (2) eucalyptus removal furthered decomposition process of the soil organic matter. Hence, we determined the soil granulometry, the δ13C and δ15N in leaves, in the superficial soil layer, and litter in three sites: a secondary forest in the Atlantic forest, and other two sites where eucalyptus had been removed in different times: 12 and 3 months ago (M12 and M3, respectively). Litter samples presented intermediate δ13C and δ15N values in comparison with leaves and soil. In the M3, the greater δ13C values in both litter rest fraction and soil indicate the presence, cycling and soil incorporation of C, coming from the C4 photosynthesis of grassy species (Poaceae). In the secondary forest, the soil δ15N values were twice higher, compared with the eucalyptus removal sites, revealing the negative influence from these exotic species upon the ecosystem N dynamics. In the M12, the leaves presented higher δ13C mean value and lower δ15N values, compared with those from the other sites. The difference of δ13C values in the litter fractions regarding the soil led to a greater fractioning of 13C in all sites, except the flower + fruit fractions in the secondary forest, and the rest fraction in the M3 site. We conclude that the permanence of this exotic species and the eucalyptus removal have altered the C and N isotopic and elemental compositions in the soil-plant system. Our results suggest there was organic matter decomposition in all litter fractions and in all sites. However, a greater organic matter decomposition process was observed in the M3 soil, possibly because of a more intense recent input of vegetal material, as well as the presence of grassy, easily-decomposing herbaceous species, only in this site. Therefore, the dual-isotope approach generated a more integrated picture of the impact on the ecosystem after removing eucalyptus in this secondary Atlantic forest, and could be regarded as an option for future eucalyptus removal studies. creator: Milena Carvalho Teixeira creator: Angela Pierre Vitória creator: Carlos Eduardo de Rezende creator: Marcelo Gomes de Almeida creator: Gabriela B. Nardoto uri: https://doi.org/10.7717/peerj.9222 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Teixeira et al. title: Here are the polyps: in situ observations of jellyfish polyps and podocysts on bivalve shells link: https://peerj.com/articles/9260 last-modified: 2020-05-28 description: Most Scyphozoan jellyfish species have a metagenic life cycle involving a benthic, asexually reproducing polyp stage and a sexually reproducing medusa stage. Medusae can be large and conspicuous and most can be identified using morphological characteristics. Polyps on the other hand are small, live a cryptic life attached to hard substrates and often are difficult or impossible to distinguish based on morphology alone. Consequently, for many species the polyp stage has not been identified in the natural environment. We inspected hard substrates in various habitats for the presence of Scyphozoan polyps. Three polyps were found on Dogger Bank, Central North Sea, attached to the inside of the umbo of empty valves of the bivalves Mactra stultorum and Spisula subtruncata. One polyp was accompanied by four podocysts. With this knowledge, the inside of bivalve shells washed ashore in Oostende (Belgium) was inspected and supposed podocysts on the inside of empty valves of Cerastoderma edule and Spisula elliptica were found. Polyps and podocysts were identified to species level by 18S rDNA and mitochondrial COI sequencing. The three polyps found on Dogger Bank all belonged to the compass jellyfish Chrysaora hysoscella. One podocyst from the Oostende beach also belonged to this species but another podocyst belonged to Cyanea lamarkii. These are the first in situ observations of C. hysoscella and C. lamarckii polyps and podocysts in the natural environment. Mactra, Cerastoderma and Spisula species are abundant in many North Sea regions and empty bivalve shells could provide an abundant settling substrate for jellyfish polyps in the North Sea and other areas. Several new strategies to increase the detection of polyps on bivalve shells are presented. creator: Lodewijk van Walraven creator: Judith van Bleijswijk creator: Henk W. van der Veer uri: https://doi.org/10.7717/peerj.9260 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 van Walraven et al. title: Dynamics of Influenza A (H5N1) virus protein sequence diversity link: https://peerj.com/articles/7954 last-modified: 2020-05-27 description: BackgroundInfluenza A (H5N1) virus is a global concern with potential as a pandemic threat. High sequence variability of influenza A viruses is a major challenge for effective vaccine design. A continuing goal towards this is a greater understanding of influenza A (H5N1) proteome sequence diversity in the context of the immune system (antigenic diversity), the dynamics of mutation, and effective strategies to overcome the diversity for vaccine design.MethodsHerein, we report a comprehensive study of the dynamics of H5N1 mutations by analysis of the aligned overlapping nonamer positions (1–9, 2–10, etc.) of more than 13,000 protein sequences of avian and human influenza A (H5N1) viruses, reported over at least 50 years. Entropy calculations were performed on 9,408 overlapping nonamer position of the proteome to study the diversity in the context of immune system. The nonamers represent the predominant length of the binding cores for peptides recognized by the cellular immune system. To further dissect the sequence diversity, each overlapping nonamer position was quantitatively analyzed for four patterns of sequence diversity motifs: index, major, minor and unique.ResultsAlmost all of the aligned overlapping nonamer positions of each viral proteome exhibited variants (major, minor, and unique) to the predominant index sequence. Each variant motif displayed a characteristic pattern of incidence change in relation to increased total variants. The major variant exhibited a restrictive pyramidal incidence pattern, with peak incidence at 50% total variants. Post this peak incidence, the minor variants became the predominant motif for majority of the positions. Unique variants, each sequence observed only once, were present at nearly all of the nonamer positions. The diversity motifs (index and variants) demonstrated complex inter-relationships, with motif switching being a common phenomenon. Additionally, 25 highly conserved sequences were identified to be shared across viruses of both hosts, with half conserved to several other influenza A subtypes.DiscussionThe presence of distinct sequences (nonatypes) at nearly all nonamer positions represents a large repertoire of reported viral variants in the proteome, which influence the variability dynamics of the viral population. This work elucidated and provided important insights on the components that make up the viral diversity, delineating inherent patterns in the organization of sequence changes that function in the viral fitness-selection. Additionally, it provides a catalogue of all the mutational changes involved in the dynamics of H5N1 viral diversity for both avian and human host populations. This work provides data relevant for the design of prophylactics and therapeutics that overcome the diversity of the virus, and can aid in the surveillance of existing and future strains of influenza viruses. creator: Hadia Syahirah Abd Raman creator: Swan Tan creator: Joseph Thomas August creator: Asif M. Khan uri: https://doi.org/10.7717/peerj.7954 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Abd Raman et al. title: Changes of liver transcriptome profiles following oxidative stress in streptozotocin-induced diabetes in mice link: https://peerj.com/articles/8983 last-modified: 2020-05-27 description: BackgroundOxidative-stress (OS) was causal in the development of cell dysfunction and insulin resistance. Streptozotocin (STZ) was an alkylation agent that increased reactive oxygen species (ROS) levels. Here we aimed to explore the oxidative-stress and related RNAs in the liver of STZ-induced diabetic mice.MethodsRNA-sequencing was performed using liver tissues from STZ induced diabetic mice and controls. Pathway and Gene Ontology (GO) analyses were utilized to annotate the target genes. The differentially expressed RNAs involved in the peroxisome pathway were validated by qRT-PCR. The glucose metabolite and OS markers were measured in the normal control (NC) and STZ-induced diabetic mellitus (DM) group.ResultsThe levels of serum Fasting insulin, HbA1c, Malondialdehyde (MDA) and 8-iso-prostaglandin F2α (8-iso-PGF2α) were significant higher in DM groups than NC group, while SOD activity decreased significantly in DM groups. We found 416 lncRNAs and 910 mRNAs were differentially expressed in the STZ-induced diabetic mice compared to the control group. OS associated RNAs were differentially expressed in the liver of STZ-induced diabetic mice.ConclusionThis study confirmed that the OS was increased in the STZ-induced DM mice as evidenced by the increase of lipid peroxidation product MDA and 8-iso-PGF2α, identified aberrantly expressed lncRNAs and mRNAs in STZ-induced diabetic mice. creator: Shuren Guo creator: Xiaohuan Mao creator: Yunmeng Yan creator: Yan Zhang creator: Liang Ming uri: https://doi.org/10.7717/peerj.8983 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Guo et al. title: Purpureocillium lilacinum and Metarhizium marquandii as plant growth-promoting fungi link: https://peerj.com/articles/9005 last-modified: 2020-05-27 description: BackgroundEspecially on commodities crops like soybean, maize, cotton, coffee and others, high yields are reached mainly by the intensive use of pesticides and fertilizers. The biological management of crops is a relatively recent concept, and its application has increased expectations about a more sustainable agriculture. The use of fungi as plant bioinoculants has proven to be a useful alternative in this process, and research is deepening on genera and species with some already known potential. In this context, the present study focused on the analysis of the plant growth promotion potential of Purpureocillium lilacinum, Purpureocillium lavendulum and Metarhizium marquandii aiming its use as bioinoculants in maize, bean and soybean.MethodsPurpureocillium spp. and M. marquandii strains were isolated from soil samples. They were screened for their ability to solubilize phosphorus (P) and produce indoleacetic acid (IAA) and the most promising strains were tested at greenhouse in maize, bean and soybean plants. Growth promotion parameters including plant height, dry mass and contents of P and nitrogen (N) in the plants and in the rhizospheric soil were assessed.ResultsThirty strains were recovered and characterized as Purpureocillium lilacinum (25), Purpureocillium lavendulum (4) and Metarhizium marquandii (1). From the trial for P solubilization and IAA production, seven strains were selected and inoculated in maize, bean and soybean plants. These strains were able to modify in a different way the evaluated parameters involving plant growth in each crop, and some strains distinctly increased the availability of P and N, for the last, an uncommon occurrence involving these fungi. Moreover, the expected changes identified at the in vitro analysis were not necessarily found in planta. In addition, this study is the first to evaluate the effect of the isolated inoculation of these fungi on the growth promotion of maize, bean and soybean plants. creator: Noemi Carla Baron creator: Andressa de Souza Pollo creator: Everlon Cid Rigobelo uri: https://doi.org/10.7717/peerj.9005 license: https://creativecommons.org/licenses/by-nc/4.0 rights: © 2020 Baron et al. title: Prevalence and molecular characterization of some circulating strains of the peste-des-petits-ruminants virus in Saudi Arabia between 2014–2016 link: https://peerj.com/articles/9035 last-modified: 2020-05-27 description: BackgroundThe peste-des-petits-ruminants virus (PPRV) is a highly devastating virus of small ruminants in many parts of the world, including the Kingdome of Saudi Arabia. Therefore, our objectives were (1) to conduct a molecular prevalence study of PPRV in sheep and goat across the KSA and (2) to isolate and identify currently circulating PPRV lineages. Swabs and tissue specimens were collected from 97 herds suspected to be infected with PPRV across the Kingdome of Saudi Arabia (KSA). Testing for the presence of the virus was done by the real-time PCR. Confirmation of the identity of the reactions was done by the gel-based-PCR then by sequencing of the partial PPRV genome.ResultsOur results revealed that 24.1% of the tested specimens were PPRV-positive. Isolation of PPRV was successful from samples using the Vero cell line. Sequence analysis of some partial PPRV genes (N, F, M, L, P, and H) revealed that these strains were belonging to lineage IV of the PPRV.ConclusionsThis is the first study to conduct both the nationwide prevalence, isolation, and molecular characterizations of the PPRV in the KSA. Continuous surveillance and monitoring of the circulating strains of PPRV among sheep and goats will contribute substantially to the global eradication campaign of such a virus. creator: Maged Gomaa Hemida creator: Hussain Mohammed Alghadeer creator: Mohammed Alhammadi creator: Sayed Ali uri: https://doi.org/10.7717/peerj.9035 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Hemida et al. title: A new sponge-associated starfish, Astrolirus patricki sp. nov. (Asteroidea: Brisingida: Brisingidae), from the northwestern Pacific seamounts link: https://peerj.com/articles/9071 last-modified: 2020-05-27 description: Seamounts are important deep ocean entities that serve as reservoirs for varied types of habitats and fauna. During the Chinese cruises in the northwestern Pacific seamount areas, a new starfish species of order Brisingida, Astrolirus patricki, was found at 1,458–2,125 m depth. All specimens of the new species were observed in situ attaching on hexactinellid sponges, suggesting a possible close relationship between the two taxa. A. patrickisp. nov. is the second known species of the genus, characterized by the abutting plates in the intercostal integument, separated first pair of adambulacral plates and densely distributed proximal spines. Phylogenetic analyses were conducted for order Brisingida to incorporate the new species as well as Hymenodiscus cf. fragilis (Fisher, 1906), Freyella cf. attenuata Sladen, 1889 and two Brisinga spp., for which we present the molecular data for the first time. Phylogenetic trees suggest a close relationship between A. patrickisp. nov. with Brisinga species rather than with Hymenodiscus species, which is inconsistent with morphological taxonomy. This study highlights the distinct morphological and ecological characters of the new species and provides new data for future investigation on Brisingida phylogeny. creator: Ruiyan Zhang creator: Yadong Zhou creator: Ning Xiao creator: Chunsheng Wang uri: https://doi.org/10.7717/peerj.9071 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: Evaluating Eucalyptus leaf colonization by Brasilonema octagenarum (Cyanobacteria, Scytonemataceae) using in planta experiments and genomics link: https://peerj.com/articles/9158 last-modified: 2020-05-27 description: BackgroundBrasilonema is a cyanobacterial genus found on the surface of mineral substrates and plants such as bromeliads, orchids and eucalyptus. B. octagenarum stands out among cyanobacteria due to causing damage to the leaves of its host in an interaction not yet observed in other cyanobacteria. Previous studies revealed that B. octagenaum UFV-E1 is capable of leading eucalyptus leaves to suffer internal tissue damage and necrosis by unknown mechanisms. This work aimed to investigate the effects of B. octagenarum UFV-E1 inoculation on Eucalyptus urograndis and to uncover molecular mechanisms potentially involved in leaf damage by these cyanobacteria using a comparative genomics approach.ResultsLeaves from E. urograndis saplings were exposed for 30 days to B. octagenarum UFV-E1, which was followed by the characterization of its genome and its comparison with the genomes of four other Brasilonema strains isolated from phyllosphere and the surface of mineral substrates. While UFV-E1 inoculation caused an increase in root and stem dry mass of the host plants, the sites colonized by cyanobacteria on leaves presented a significant decrease in pigmentation, showing that the cyanobacterial mats have an effect on leaf cell structure. Genomic analyses revealed that all evaluated Brasilonema genomes harbored genes encoding molecules possibly involved in plant-pathogen interactions, such as hydrolases targeting plant cell walls and proteins similar to known virulence factors from plant pathogens. However, sequences related to the type III secretory system and effectors were not detected, suggesting that, even if any virulence factors could be expressed in contact with their hosts, they would not have the structural means to actively reach plant cytoplasm.ConclusionsLeaf damage by this species is likely related to the blockage of access to sunlight by the efficient growth of cyanobacterial mats on the phyllosphere, which may hinder the photosynthetic machinery and prevent access to some essential molecules. These results reveal that the presence of cyanobacteria on leaf surfaces is not as universally beneficial as previously thought, since they may not merely provide the products of nitrogen fixation to their hosts in exchange for physical support, but in some cases also hinder regular leaf physiology leading to tissue damage. creator: Danillo O. Alvarenga creator: Maione W. Franco creator: Kaarina Sivonen creator: Marli F. Fiore creator: Alessandro M. Varani uri: https://doi.org/10.7717/peerj.9158 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Alvarenga et al. title: Acute effects of air pollution on all-cause mortality: a natural experiment from haze control measures in Chiang Mai Province, Thailand link: https://peerj.com/articles/9207 last-modified: 2020-05-27 description: BackgroundSerious haze episodes have been a seasonal event in Chiang Mai province for more than a decade. In 2008, local government agencies introduced comprehensive measures to control haze and limit its impacts on public health. This study assessed the acute effects of ambient air pollutants on all-cause mortality before and after the introduction of those haze control measures.MethodsWe obtained daily mortality counts and data on mass concentrations of particulate matter <10 micron in aerodynamic diameter (PM10), gaseous pollutants (SO2, NO2, O3, and CO), and meteorology in Chiang Mai Province between January 2002 and December 2016. We analyzed the data using a case-crossover approach adjusting for temperature, relative humidity, seasonality, and day-of-week. We assessed change in the excess risks of all-cause mortality associated with an increase in interquartile range (IQR) of pollutant concentration before and after control measures came into force.ResultsWe found decreased PM10 levels and markedly reduced excess risks of daily mortality associated with an IQR increase in PM10 concentrations in the years after haze-control measures were implemented (2009–2016). We found mixed results for gaseous pollutants: SO2 showed no significant change in excess risk of daily mortality throughout the study period, while NO2 and CO showed significant excess risks only in the period 2012–2016, and 8-h maximum O3 showed a decrease in excess risk despite an increase in its atmospheric levels after the introduction of haze control measures in 2008.ConclusionsThe findings indicate that the government haze control measures first introduced in Chiang Mai province in 2008 have successfully reduced episodic PM10 concentrations, which has led to a decrease in short-term all-cause mortality. creator: Nitaya Vajanapoom creator: Patcharee Kooncumchoo creator: Thuan-Quoc Thach uri: https://doi.org/10.7717/peerj.9207 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Vajanapoom et al. title: Rapid identification of melioidosis agent by an insulated isothermal PCR on a field–deployable device link: https://peerj.com/articles/9238 last-modified: 2020-05-27 description: Background Burkholderia pseudomallei causes melioidosis, a serious illness that can be fatal if untreated or misdiagnosed. Culture from clinical specimens remains the gold standard but has low diagnostic sensitivity.MethodIn this study, we developed a rapid, sensitive and specific insulated isothermal Polymerase Chain Reaction (iiPCR) targeting bimA gene (Burkholderia Intracellular Motility A; BPSS1492) for the identification of B. pseudomallei. A pair of novel primers: BimA(F) and BimA(R) together with a probe were designed and 121 clinical B. pseudomallei strains obtained from numerous clinical sources and 10 ATCC non-targeted strains were tested with iiPCR and qPCR in parallel.ResultsAll 121 B. pseudomallei isolates were positive for qPCR while 118 isolates were positive for iiPCR, demonstrating satisfactory agreement (97.71%; 95% CI [93.45–99.53%]; k = 0.87). Sensitivity of the bimA iiPCR/POCKIT assay was 97.52% with the lower detection limit of 14 ng/µL of B. pseudomallei DNA. The developed iiPCR assay did not cross-react with 10 types of non-targeted strains, indicating good specificity.ConclusionThis bimA iiPCR/POCKIT assay will undoubtedly complement other methodologies used in the clinical laboratory for the rapid identification of this pathogen. creator: Kek Heng Chua creator: E. Wei Tan creator: Hwa Chia Chai creator: SD Puthucheary creator: Ping Chin Lee creator: Suat Moi Puah uri: https://doi.org/10.7717/peerj.9238 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chua et al. title: Effects of temperature on metabolic scaling in black carp link: https://peerj.com/articles/9242 last-modified: 2020-05-27 description: The surface area (SA) of organs and cells may vary with temperature, which changes the SA exchange limitation on metabolic flows as well as the influence of temperature on metabolic scaling. The effect of SA change can intensify (when the effect is the same as that of temperature) or compensate for (when the effect is the opposite of that of temperature) the negative effects of temperature on metabolic scaling, which can result in multiple patterns of metabolic scaling with temperature among species. The present study aimed to examine whether metabolic scaling in black carp changes with temperature and to identify the link between metabolic scaling and SA at the organ and cellular levels at different temperatures. The resting metabolic rate (RMR), gill surface area (GSA) and red blood cell (RBC) size of black carp with different body masses were measured at 10 °C and 25 °C, and the scaling exponents of these parameters were compared. The results showed that both body mass and temperature independently affected the RMR, GSA and RBC size of black carp. A consistent scaling exponent of RMR (0.764, 95% CI [0.718–0.809]) was obtained for both temperatures. The RMR at 25 °C was 2.7 times higher than that at 10 °C. At both temperatures, the GSA scaled consistently with body mass by an exponent of 0.802 (95% CI [0.759–0.846]), while RBC size scaled consistently with body mass by an exponent of 0.042 (95% CI [0.010–0.075]). The constant GSA scaling can explain the constant metabolic scaling as temperature increases, as metabolism may be constrained by fluxes across surfaces. The GSA at 10 °C was 1.2 times higher than that at 25 °C, which suggests that the constraints of GSA on the metabolism of black carp is induced by the higher temperature. The RBC size at 10 °C was 1.1 times higher than that at 25 °C. The smaller RBC size (a larger surface-to-volume ratio) at higher temperature suggests an enhanced oxygen supply and a reduced surface boundary limit on bR, which offset the negative effect of temperature on bR. creator: Qian Li creator: Xiaoling Zhu creator: Wei Xiong creator: Yanqiu Zhu creator: Jianghui Zhang creator: Pathe Karim Djiba creator: Xiao Lv creator: Yiping Luo uri: https://doi.org/10.7717/peerj.9242 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Li et al. title: Role of echocardiography in screening for portopulmonary hypertension in liver transplant candidates: a meta-analysis link: https://peerj.com/articles/9243 last-modified: 2020-05-27 description: ObjectivesTo demonstrate the screening value of echocardiography for portopulmonary hypertension (POPH) in liver transplant candidates.DesignSystematic review and meta-analysis.BackgroundPOPH is a complication of end-stage liver disease that adversely affects the outcome of orthotopic liver transplant. There are no specific symptoms in the early stage of POPH. POPH reduce the survival rate of patients with end-stage liver disease specially if they are not diagnosed. Therefore, early detection may improve prognosis. The objective of this study is to explore the screening value of echocardiography on liver transplant candidates for screening of POPH compared to right heart catheterization (RHC).MethodPubMed, EMBASE and the Cochrane Library were searched by two independent reviewers for potentially eligible studies published up to 30 June 2019 to retrieve data based on per-patient analysis. STATA, Meta-DiSc, and RevMan were applied to perform this meta-analysis.ResultsOur search yielded 1576 studies, of which 11 satisfied the inclusion criteria. The pooled sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR) and area under the summary receiver operating characteristic (SROC) curve (AUC) of echocardiography for POPH were 0.85 (95% CI [0.65–0.94]), 0.83 (95% CI [0.73–0.90]), 4.99 (95% CI [3.03–8.21]), 0.19 (95% CI [0.07–0.46]), and 0.91 (95% CI [0.88–0.93]), respectively. Deeks’ funnel plot did not indicate the existence of publication bias (P = 0.66).ConclusionsEchocardiography, a noninvasive modality, provides superior screening for POPH, but the diagnosis of POPH still requires RHC. PROSPERO registration number CRD42019144589. creator: Xin Yin creator: Yueming Shao creator: Yu Zhang creator: Hui Gao creator: Tingting Qin creator: Xiaoyu Wen creator: Chen Yang uri: https://doi.org/10.7717/peerj.9243 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yin et al. title: Exact integer linear programming solvers outperform simulated annealing for solving conservation planning problems link: https://peerj.com/articles/9258 last-modified: 2020-05-27 description: The resources available for conserving biodiversity are limited, and so protected areas need to be established in places that will achieve objectives for minimal cost. Two of the main algorithms for solving systematic conservation planning problems are Simulated Annealing (SA) and exact integer linear programing (EILP) solvers. Using a case study in BC, Canada, we compare the cost-effectiveness and processing times of SA used in Marxan versus EILP using both commercial and open-source algorithms. Plans for expanding protected area systems based on EILP algorithms were 12–30% cheaper than plans using SA, due to EILP’s ability to find optimal solutions as opposed to approximations. The best EILP solver we examined was on average 1,071 times faster than the SA algorithm tested. The performance advantages of EILP solvers were also observed when we aimed for spatially compact solutions by including a boundary penalty. One practical advantage of using EILP over SA is that the analysis does not require calibration, saving even more time. Given the performance of EILP solvers, they can be used to generate conservation plans in real-time during stakeholder meetings and can facilitate rapid sensitivity analysis, and contribute to a more transparent, inclusive, and defensible decision-making process. creator: Richard Schuster creator: Jeffrey O. Hanson creator: Matthew Strimas-Mackey creator: Joseph R. Bennett uri: https://doi.org/10.7717/peerj.9258 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Schuster et al. title: Bioinformatics profiling identifies seven immune-related risk signatures for hepatocellular carcinoma link: https://peerj.com/articles/8301 last-modified: 2020-05-26 description: BackgroundDensity of tumor infiltrating lymphocytes (TIL) and expressions of certain immune-related genes have prognostic and predictive values in hepatocellular carcinoma (HCC); however, factors determining the immunophenotype of HCC patients are still unclear. In the current study, the transcript sequencing data of liver cancer were systematically analyzed to determine an immune gene marker for the prediction of clinical outcome of HCC.MethodsRNASeq data and clinical follow-up information were downloaded from The Cancer Genome Atlas (TCGA), and the samples were assigned into high-stage and low-stage groups. Immune pathway-related genes were screened from the Molecular Signatures Database v4.0 (MsigDB) database. LASSO regression analysis was performed to identify robust immune-related biomarkers in predicting HCC clinical outcomes. Moreover, an immune gene-related prognostic model was established and validated by test sets and Gene Expression Omnibus (GEO) external validation sets.ResultsWe obtained 319 immune genes from MsigDB, and the genes have different expression profiles in high-stage and low-stage of HCC. Univariate survival analysis found that 17 genes had a significant effect on HCC prognosis, among them, 13 (76.5%) genes were prognostically protective factors. Further lasso regression analysis identified seven potential prognostic markers (IL27, CD1D, NCOA6, CTSE, FCGRT, CFHR1, and APOA2) of robustness, most of which are related to tumor development. Cox regression analysis was further performed to establish a seven immune gene signature, which could stratify the risk of samples in training set, test set and external verification set (p < 0.01), and the AUC in both training set and test set was greater than 0.85, which also greater compared with previous studies.ConclusionThis study constructed a 7-immunogenic marker as novel prognostic markers for predicting survival of HCC patients. creator: Feng Xue creator: Lixue Yang creator: Binghua Dai creator: Hui Xue creator: Lei Zhang creator: Ruiliang Ge creator: Yanfu Sun uri: https://doi.org/10.7717/peerj.8301 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Xue et al. title: Descriptions of five new species of the salamander genus Chiropterotriton (Caudata: Plethodontidae) from eastern Mexico and the status of three currently recognized taxa link: https://peerj.com/articles/8800 last-modified: 2020-05-26 description: The genus Chiropterotriton is endemic to Mexico with a geographical distribution along the Sierra Madre Oriental, the Trans Mexican Volcanic Belt and the Sierra de Juárez. The recent use of molecular tools has shown that Mexico’s amphibian diversity is highly underestimated, including a large number of cryptic, unnamed species. Chiropterotriton has 18 described species including terrestrial, arboreal and cave-dwelling species. In previous molecular studies, the presence of multiple undescribed species was evident. We present a phylogenetic hypothesis based on mitochondrial data, which includes all described species and six undescribed taxa. Based on the morphological analyses and, when available, combined with molecular data, we describe five new species of the genus; Chiropterotriton casasi sp. nov., C. ceronorum sp. nov., C. melipona sp. nov., C. perotensis sp. nov. and C. totonacus sp. nov. In addition, we redescribe two others: Chiropterotriton chiropterus and C. orculus, and provide a comparable account of one additional sympatric congener. This increases the number of species in the genus to 23, which represent a considerable component of Mexican plethodontid richness. creator: Gabriela Parra Olea creator: Mirna G. Garcia-Castillo creator: Sean M. Rovito creator: Jessica A. Maisano creator: James Hanken creator: David B. Wake uri: https://doi.org/10.7717/peerj.8800 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Parra Olea et al. title: Effective connectivity differences in motor network during passive movement of paretic and non-paretic ankles in subacute stroke patients link: https://peerj.com/articles/8942 last-modified: 2020-05-26 description: BackgroundA better understanding of the neural changes associated with paresis in stroke patients could have important implications for therapeutic approaches. Dynamic Causal Modeling (DCM) for functional magnetic resonance imaging (fMRI) is commonly used for analyzing effective connectivity patterns of brain networks due to its significant property of modeling neural states behind fMRI signals. We applied this technique to analyze the differences between motor networks (MNW) activated by continuous passive movement (CPM) of paretic and non-paretic ankles in subacute stroke patients. This study aimed to identify CPM induced connectivity characteristics of the primary sensory area (S1) and the differences in extrinsic directed connections of the MNW and to explain the hemodynamic differences of brain regions of MNW.MethodsFor the network analysis, we used ten stroke patients’ task fMRI data collected under CPMs of both ankles. Regions for the MNW, the primary motor cortex (M1), the premotor cortex (PM), the supplementary motor area (SMA) and the S1 were defined in a data-driven way, by independent component analysis. For the network analysis of both CPMs, we compared twelve models organized into two model-families, depending on the S1 connections and input stimulus modeling. Using DCM, we evaluated the extrinsic connectivity strengths and hemodynamic parameters of both stimulations of all patients.ResultsAfter a statistical comparison of the extrinsic connections and their modulations of the “best model”, we concluded that three contralateral self-inhibitions (cM1, cS1 and cSMA), one contralateral inter-regional connection (cSMA→cM1), and one interhemispheric connection (cM1→iM1) were significantly different. Our research shows that hemodynamic parameters can be estimated with the Balloon model using DCM but the parameters do not change with stroke.ConclusionsOur results confirm that the DCM-based connectivity analyses combined with Bayesian model selection may be a useful technique for quantifying the alteration or differences in the characteristics of the motor network in subacute stage stroke patients and in determining the degree of MNW changes. creator: Marianna Nagy creator: Csaba Aranyi creator: Gábor Opposits creator: Tamás Papp creator: Levente Lánczi creator: Ervin Berényi creator: Csilla Vér creator: László Csiba creator: Péter Katona creator: Tamás Spisák creator: Miklós Emri uri: https://doi.org/10.7717/peerj.8942 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Nagy et al. title: Microbiome analysis of healthy and diseased sponges Lubomirskia baicalensis by using cell cultures of primmorphs link: https://peerj.com/articles/9080 last-modified: 2020-05-26 description: Endemic sponges (Demosponges, Lubomirskiidae) dominate the fauna of the littoral zone of Lake Baikal. These freshwater sponges live in symbiosis with diverse eukaryotes and prokaryotes, including chlorophyll-containing microalgae. Within the last 5 years, the incidence of sponge disease and mortality events in Lake Baikal has increased. The etiology and ecology of these events remain unknown, in part because of the lack of models to study sponge-microbe interactions. In this work, we tested the use of primmorph cell cultures of Lubomirskia baicalensis as a tool for investigating the microbiomes of sponges. We infected primmorphs, cultured in vitro, with samples from diseased sponges and observed, by microscopy, disease symptoms, including loss of green symbionts, associated with mass die-off events. Subsequent sequencing of 16S rRNA gene fragments revealed that the microbiome community of healthy sponge and primmorphs formed a group separate from the community of diseased sponges and infected primmorphs. This confirms the suitability of the primmorph cell culture as a model sponge system. We also discovered mass mortality of green symbionts (Chlorophyta) was associated with a shift in the microbial communities of sponges/primmorphs. Microbes in diseased sponges, and infected primmorphs, belonged mainly to the phyla Bacteroidetes and Proteobacteria and these families Flavobacteriaceae, Burkholderiaceae, and Moraxellaceae. Primmorphs cell culture may provide a model to study interactions between these bacteria and their host and elucidate the cause of mass mortality events. creator: Lubov Chernogor creator: Elizaveta Klimenko creator: Igor Khanaev creator: Sergei Belikov uri: https://doi.org/10.7717/peerj.9080 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chernogor et al. title: Cca-miR398 increases copper sulfate stress sensitivity via the regulation of CSD mRNA transcription levels in transgenic Arabidopsis thaliana link: https://peerj.com/articles/9105 last-modified: 2020-05-26 description: MicroRNAs play crucial roles during the process of plant development under stress conditions. Copper is an essential micronutrient for most organisms and serves as an important redox-active cofactor for various functional proteins. In the present study, we investigated the effects of copper sulfate stress on hickory (Carya cathayensis) root development. We identified that hickory cca-miR398 was related to copper sulfate stress response, targeting Copper/Zinc superoxide dismutases (cytosolic (CSD1) and chloroplastic (CSD2)) and a 5b subunit of mitochondrial cytochrome C oxidase (COX5b.1) that are linked directly to stress regulatory networks. The sequence of hickory cca-miR398 is highly similar to that of Arabidopsis miR398b and miR398c, regardless of one nucleotide variation. Therefore, target genes of cca-miR398 were investigated by using 5′-Rapid-amplification of cDNA ends. An overexpression of cca-miR398 in Arabidopsis caused a reduction not only in root length and cotyledon greening, but also in the CSD1, CSD2, and CSD3 transcription levels. These reductions had greater significance in transgenic Arabidopsis than in wild-type Arabidopsis under copper sulfate stress. The level of physiological indicators also changed in transgenic Arabidopsis. In addition, the expressions of copper-responsive microRNAs, such as miR397 and miR408, were affected by the copper sulfate stress. These results showed that CSD possesses the ability to enhance copper sulfate stress response in both transgenic Arabidopsis and hickory roots by increasing the production of superoxide dismutase. Our results also demonstrated that cca-miR398 weakens hickory tolerance to copper sulfate by regulating CSD targets. creator: Zhichao Sun creator: Lilu Shu creator: Wei Zhang creator: Zhengjia Wang uri: https://doi.org/10.7717/peerj.9105 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Sun et al. title: The combined effects of light intensity, temperature, and water potential on wall deposition in regulating hypocotyl elongation of Brassica rapa link: https://peerj.com/articles/9106 last-modified: 2020-05-26 description: Hypocotyl elongation is a critical sign of seed germination and seedling growth, and it is regulated by multi-environmental factors. Light, temperature, and water potential are the major environmental stimuli, and their regulatory mechanism on hypocotyl growth has been extensively studied at molecular level. However, the converged point in signaling process of light, temperature, and water potential on modulating hypocotyl elongation is still unclear. In the present study, we found cell wall was the co-target of the three environmental factors in regulating hypocotyl elongation by analyzing the extension kinetics of hypocotyl and the changes in hypocotyl cell wall of Brassica rapa under the combined effects of light intensity, temperature, and water potential. The three environmental factors regulated hypocotyl cell elongation both in isolation and in combination. Cell walls thickened, maintained, or thinned depending on growth conditions and developmental stages during hypocotyl elongation. Further analysis revealed that the imbalance in wall deposition and hypocotyl elongation led to dynamic changes in wall thickness. Low light repressed wall deposition by influencing the accumulation of cellulose, hemicellulose, and pectin; high temperature and high water potential had significant effects on pectin accumulation overall. It was concluded that wall deposition was tightly controlled during hypocotyl elongation, and low light, high temperature, and high water potential promoted hypocotyl elongation by repressing wall deposition, especially the deposition of pectin. creator: Hongfei Wang creator: Qingmao Shang uri: https://doi.org/10.7717/peerj.9106 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang and Shang title: At the crossroads of botanical collections and molecular genetics laboratory: a preliminary study of obtaining amplifiable DNA from moss herbarium material link: https://peerj.com/articles/9109 last-modified: 2020-05-26 description: BackgroundResearch focused on extreme environments is often associated with difficulties in obtaining fresh plant material. Herbaria may provide great support as they house large collections of specimens from different parts of the world. Accordingly, there is also a growing interest in methods using herbarium specimens in molecular studies. Much of the literature on herbarium DNA is aimed to improve extraction and PCR amplification and is focused mostly on vascular plants. Here, I provide a brief study of DNA extraction efficiency from moss herbarium specimens, emphasizing the importance of herbaria as an invaluable source of material from hard-to-access geographical areas, such as the Antarctic region.MethodsThe presented study is based on herbarium collections of 25 moss species collected in the austral polar regions between 1979 and 2013. The majority of samples were obtained using the DNeasy Plant Mini Kit (Qiagen, Hilden, Germany). The remaining, smaller part was extracted using an adapted CTAB-based approach. The performance of DNA extraction methods in terms of PCR amplification success was measured by testing several DNA fragments of various size. Furthermore, in order to estimate of DNA fragmentation level, an automated on-chip electrophoresis system was used.ResultsResults reveal that DNA purity and the length of the target genetic region are the fundamental agents which drive the successful PCR reaction. Conversely, the DNA yield and specimen age seem to be less relevant. With this study, I present also an optimized CTAB-based approach which may effectively suppress inhibitors in the herbarium DNA. This method can be considered a cheaper alternative to column-based technology, particularly useful for dealing with a large number of samples. Results of this study confirmed previous reports and contribute to filling the existing gap in molecular analyses which involve the use of herbarium collections of mosses. creator: Marta Saługa uri: https://doi.org/10.7717/peerj.9109 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Saługa title: Characterization of the bacterial microbiota composition and evolution at different intestinal tract in wild pigs (Sus scrofa ussuricus) link: https://peerj.com/articles/9124 last-modified: 2020-05-26 description: Commensal microorganisms are essential to the normal development and function of many aspects of animal biology, including digestion, nutrient absorption, immunological development, behaviors, and evolution. The specific microbial composition and evolution of the intestinal tracts of wild pigs remain poorly characterized. This study therefore sought to assess the composition, distribution, and evolution of the intestinal microbiome of wild pigs. For these analyses, 16S rRNA V3-V4 regions from five gut sections prepared from each of three wild sows were sequenced to detect the microbiome composition. These analyses revealed the presence of 6,513 operational taxonomic units (OTUs) mostly distributed across 17 phyla and 163 genera in these samples, with Firmicutes and Actinobacteria being the most prevalent phyla of microbes present in cecum and jejunum samples, respectively. Moreover, the abundance of Actinobacteria in wild pigs was higher than that in domestic pigs. At the genus level the Bifidobacterium and Allobaculum species of microbes were most abundant in all tested gut sections, with higher relative abundance in wild pigs relative to domestic pigs, indicating that in the process of pig evolution, the intestinal microbes also evolved, and changes in the intestinal microbial diversity could have been one of the evolutionary forces of pigs. Intestinal microbial functional analyses also revealed the microbes present in the small intestine (duodenum, jejunum, and ileum) and large intestine (cecum and colon) of wild pigs to engage distinct metabolic spatial structures and pathways relative to one another. Overall, these results offer unique insights that would help to advance the current understanding of how the intestinal microbes interact with the host and affect the evolution of pigs. creator: Guangli Yang creator: Chuanxin Shi creator: Shuhong Zhang creator: Yan Liu creator: Zhiqiang Li creator: Fengyi Gao creator: Yanyan Cui creator: Yongfeng Yan creator: Ming Li uri: https://doi.org/10.7717/peerj.9124 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yang et al. title: An improved similarity-based approach to predicting and mapping soil organic carbon and soil total nitrogen in a coastal region of northeastern China link: https://peerj.com/articles/9126 last-modified: 2020-05-26 description: Soil organic carbon (SOC) and soil total nitrogen (STN) are major soil indicators for soil quality and fertility. Accurate mapping SOC and STN in soils would help both managed and natural soils and ecosystem management. This study developed an improved similarity-based approach (ISA) to predicting and mapping topsoil (0–20 cm soil depth) SOC and STN in a coastal region of northeastern China. Six environmental variables including elevation, slope gradient, topographic wetness index, the mean annual temperature, the mean annual temperature, and normalized difference vegetation index were used as predictors. Soil survey data in 2012 was designed based on the clustering of the study area into six climatic vegetation landscape units. In each landscape unit, 20–25 sampling points were determined at different landform positions considering local climate, soil type, elevation and other environmental factors, and finally 126 sampling points were obtained. Soil sampling from the depth of 0–20 cm were used for model prediction and validation. The ISA model performance was compared with the geographically weighted regression (GWR), regression kriging (RK), boosted regression trees (BRT) considering mean absolute prediction error (MAE), root mean square error (RMSE), coefficient of determination (R2), and maximum relative difference (RD) indices. We found that the ISA method performed best with the highest R2 and lowest MAE, RMSE compared to GWR, RK, and BRT methods. The ISA method could explain 76% and 83% of the total SOC and STN variability, respectively, 12–40% higher than other models in the study area. Elevation had the largest influence on SOC and STN distribution. We conclude that the developed ISA model is robust and effective in mapping SOC and STN, particularly in the areas with complex vegetation-landscape when limited samples are available. The method needs to be tested for other regions in our future research. creator: Shuai Wang creator: Kabindra Adhikari creator: Qianlai Zhuang creator: Zijiao Yang creator: Xinxin Jin creator: Qiubing Wang creator: Zhenxing Bian uri: https://doi.org/10.7717/peerj.9126 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: The combined impacts of wheat spatial position and phenology on cereal aphid abundance link: https://peerj.com/articles/9142 last-modified: 2020-05-26 description: BackgroundWheat is a staple crop that suffers from massive yield losses caused by cereal aphids. Many factors can determine the abundance of cereal aphids and the damage they cause to plants; among them are the plant’s genetic background, as well as environmental conditions such as spatial position within the plot, the composition and the distance from neighboring vegetation. Although the effects of these factors have been under scrutiny for many years, the combined effect of both factors on aphid populations is not fully understood. The goal of this study was to examine the collective impact of genotype and environment on wheat phenology (developmental stages), chemical diversity (metabolites), and insect susceptibility, as manifested by cereal aphid abundance.MethodsTo determine the influence of plant genotype on the metrics mentioned above, we measured the phenology, chemical profile, and aphid abundance of four wheat genotypes, including the tetraploid wild emmer (Triticum turgidum ssp. dicoccoides cv. Zavitan), tetraploid durum (Triticum turgidum ssp. durum cv. Svevo), and two hexaploid spring bread (Triticum aestivum), ‘Rotem’ and ‘Chinese Spring’. These genotypes are referred to as “focal” plants. To evaluate the impact of the environment, we scored the distance of each focal plant (spatial position) from two neighboring vegetation types: (i) natural resource and (ii) monoculture wheat resource.ResultsThe results demonstrated that the wild emmer wheat was the most aphid-resistant, while the bread wheat Rotem was most aphid-susceptible. Aphids were more abundant in plants that matured early. The spatial position analysis demonstrated that aphids were more abundant in focal plants located closer to the margin monoculture wheat resource rather than to the natural resource, suggesting a resource concentration effect. The analysis of metabolic diversity showed that the levels of three specialized metabolites from the flavonoid class, differed between the wheat genotypes and some minor changes in central metabolites were shown as well. Altogether, these results demonstrate a combined effect of genetic background and spatial position on wheat phenology and aphid abundance on plants. This exposes the potential role of the marginal vegetation environment in shaping the insect population of desirable crops. These findings highlight the importance of maintaining plant intra-specific variation in the agriculture system because of its potential applications in reducing pest density. creator: Zhaniya S. Batyrshina creator: Alon Cna’ani creator: Tamir Rozenberg creator: Merav Seifan creator: Vered Tzin uri: https://doi.org/10.7717/peerj.9142 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Batyrshina et al. title: Isolation and functional analyses of PvFAD2 and PvFAD3 involved in the biosynthesis of polyunsaturated fatty acids from Sacha Inchi (Plukenetia volubilis) link: https://peerj.com/articles/9169 last-modified: 2020-05-26 description: The development of ω-3 fatty acid-rich vegetable oils is essential to enrich the production of functional foods. Sacha Inchi (Plukenetia volubilis L.) is a unique oilseed crop with much potential. Its seeds contain rich polyunsaturated fatty acids (PUFAs), especially linoleic acid (LA, C18:2) and α-linolenic acid (ALA, C18:3). Endoplasmic reticulum -located ω-6 and ω-3 fatty acid desaturases (FAD) are responsible for the biosynthesis of LA and ALA, respectively, in plant seeds. Here, we isolated two full-length FAD genes from Sacha Inchi, named PvFAD2 and PvFAD3, which encoded predicted amino acid residues of 384 and 379 in protein, respectively. Protein sequence and subcellular localization analysis revealed that they were located in the endoplasmic reticulum (ER). Heterologous expression in Saccharomyces cerevisiae confirmed that PvFAD2 and PvFAD3 could catalyze LA and ALA synthesis, respectively. The stability and catalytic efficiency of the PvFAD3 protein may be closely related to temperature. In transgenic tobacco, using seed-specific expression promoters, PvFAD2 and PvFAD3 significantly promotes the production of LA (from 68% to 70.5%) and ALA (from 0.7% to 3.1%) in seed oil. These results show that PvFAD2 and PvFAD3 do, indeed, function as crucial enzymes for PUFAs biosynthesis, and provide a key gene source for the sustainable production of lipids with tailored fatty acid compositions via genetic engineering in other oil crops. creator: Tianquan Yang creator: Xiaojuan Wang creator: Tingnan Dong creator: Wei Xu creator: Aizhong Liu uri: https://doi.org/10.7717/peerj.9169 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yang et al. title: Effects of mining on the molybdenum absorption and translocation of plants in the Luanchuan molybdenum mine link: https://peerj.com/articles/9183 last-modified: 2020-05-26 description: BackgroundThere is a critical need to examine whether mining of molybdenum (Mo) ore will affect Mo absorption and translocation by plants at a community level.MethodsIndigenous plants and their rhizospheric soil (0–20 cm) growing in two different areas including the mining and the unexploited areas were collected from the Luanchuan Mo mine—one of the largest Mo mines in Asia. The concentrations of Mo and other heavy metals of plants or soil were measured by ICP-AES. Mo absorption and translocation in plants growing in two areas were investigated and compared. Heavy metal pollution in soil was also evaluated by the potential ecological hazard index method.ResultsMo concentration in mining soils was higher with the changes from 108.13 to 268.13 mg kg−1 compared to unexploited area. Mo concentrations in shoots and roots of plants growing in the mining area were also significant higher than those growing in the unexploited area with 2.59 and 2.99 times, respectively. The Mo translocation factor of plants growing in the unexploited area was 1.61, which reached 1.69 times that of plants growing in the mining area. Mo was the main heavy metal pollutant in the soil of both the mining and the unexploited areas.ConclusionMining of Mo had changed not only the Mo concentration in soil but also Mo absorption and translocation in plants. Plants growing in the mining area absorbed more Mo from the soil but translocated relatively less to shoots than plants of the unexploited area. However, the mechanisms of Mo absorption and translocation of plants in mining area should be further studied in the future. creator: Kejing Yin creator: Zhaoyong Shi creator: Mengge Zhang creator: Yajuan Li uri: https://doi.org/10.7717/peerj.9183 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yin et al. title: The high expression of MTH1 and NUDT5 predict a poor survival and are associated with malignancy of esophageal squamous cell carcinoma link: https://peerj.com/articles/9195 last-modified: 2020-05-26 description: BackgroundMTH1 and NUDT5 effectively degrade nucleotides containing 8-oxoguanine. MTH1 and NUDT5 have been linked to the malignancy of multiple cancers. However, their functions in tumor growth and metastasis in esophageal squamous carcinoma (ESCC) remain obscure. Our present study aims to explore their prognostic value in ESCC and investigate their function in MTH1 or NUDT5-knockout tumor cells.MethodsMTH1 and NUDT5 protein expression in ESCC adjacent normal tissues and tumor tissues was examined by immunohistochemistry staining. Kaplan–Meier curves were used to assess the association between their expression and overall survival (OS) in ESCC patients. Univariate and Multivariate Cox regression analyses were generated to determine the correlation between these protein expression and OS of ESCC patients. Protein expression in ESCC cell lines were measured by Western blotting. To explore the potential effects of the MTH1 and NUDT5 protein in ESCC, cell models with MTH1 or NUDT5 depletion were established. CCK-8, cell cycle, Western blotting, migration and invasion assays were performed.ResultsOur present study demonstrated that the levels of MTH1 and NUDT5 were upregulated in ESCC cell lines and ESCC tissues, the expression of MTH1 and NUDT5 in ESCC tissues was significantly higher than in adjacent non-tumorous, and higher levels of MTH1 and NUDT5 predicted a worse prognosis in patients with ESCC. MTH1 and NUDT5 are novel biomarkers of the progression of ESCC and a poor prognosis. We also found for the first time that the high expression of NUDT5 independently predicted lower OS in patients with ESCC (hazard ratio (HR) 1.751; 95% confidence interval (CI) [1.056–2.903]; p = 0.030). In addition, the depletion of MTH1 and NUDT5 strongly suppressed the proliferation of ESCC cells and significantly delayed the G1 phase of the cell cycle. Furthermore, we found that MTH1 and NUDT5 silencing inhibited epithelial–mesenchymal transition mainly by the MAPK/MEK/ERK dependent pathway, which in turn significantly decreased the cell migration and invasion of ESCC cells. Our results suggested that the overexpression of MTH1 and NUDT5 is probably involved in the tumor development and poor prognosis of ESCC. creator: Jing-Jing Wang creator: Teng-Hui Liu creator: Jin Li creator: Dan-Ni Li creator: Xin-Yuan Tian creator: Qiu-Geng Ouyang creator: Jian-Ping Cai uri: https://doi.org/10.7717/peerj.9195 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: Comprehensive analysis reveals a metabolic ten-gene signature in hepatocellular carcinoma link: https://peerj.com/articles/9201 last-modified: 2020-05-26 description: BackgroundDue to the complicated molecular and cellular heterogeneity in hepatocellular carcinoma (HCC), the morbidity and mortality still remains high level in the world. However, the number of novel metabolic biomarkers and prognostic models could be applied to predict the survival of HCC patients is still small. In this study, we constructed a metabolic gene signature by systematically analyzing the data from The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO) and International Cancer Genome Consortium (ICGC).MethodsDifferentially expressed genes (DEGs) between tumors and paired non-tumor samples of 50 patients from TCGA dataset were calculated for subsequent analysis. Univariate cox proportional hazard regression and LASSO analysis were performed to construct a gene signature. The Kaplan–Meier analysis, time-dependent receiver operating characteristic (ROC), Univariate and Multivariate Cox regression analysis, stratification analysis were used to assess the prognostic value of the gene signature. Furthermore, the reliability and validity were validated in four types of testing cohorts. Moreover, the diagnostic capability of the gene signature was investigated to further explore the clinical significance. Finally, Go enrichment analysis and Gene Set Enrichment Analysis (GSEA) have been performed to reveal the different biological processes and signaling pathways which were active in high risk or low risk group.ResultsTen prognostic genes were identified and a gene signature were constructed to predict overall survival (OS). The gene signature has demonstrated an excellent ability for predicting survival prognosis. Univariate and Multivariate analysis revealed the gene signature was an independent prognostic factor. Furthermore, stratification analysis indicated the model was a clinically and statistically significant for all subgroups. Moreover, the gene signature demonstrated a high diagnostic capability in differentiating normal tissue and HCC. Finally, several significant biological processes and pathways have been identified to provide new insights into the development of HCC.ConclusionThe study have identified ten metabolic prognostic genes and developed a prognostic gene signature to provide more powerful prognostic information and improve the survival prediction for HCC. creator: Zhipeng Zhu creator: Lulu Li creator: Jiuhua Xu creator: Weipeng Ye creator: Borong Chen creator: Junjie Zeng creator: Zhengjie Huang uri: https://doi.org/10.7717/peerj.9201 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zhu et al. title: Estimating food resource availability in arid environments with Sentinel 2 satellite imagery link: https://peerj.com/articles/9209 last-modified: 2020-05-26 description: BackgroundIn arid environments, plant primary productivity is generally low and highly variable both spatially and temporally. Resources are not evenly distributed in space and time (e.g., soil nutrients, water), and depend on global (El Niño/ Southern Oscillation) and local climate parameters. The launch of the Sentinel2-satellite, part of the European Copernicus program, has led to the provision of freely available data with a high spatial resolution (10 m per pixel). Here, we aimed to test whether Sentinel2-imagery can be used to quantify the spatial variability of a minor tussock grass (Enneapogon spp.) in an Australian arid area and whether we can identify different vegetation cover (e.g., grass from shrubs) along different temporal scenarios. Although short-lasting, the Enneapogon grassland has been identified as a key primary food source to animals in the arid environment. If we are able to identify and monitor the productivity of this species remotely, it will provide an important new tool for examining food resource dynamics and subsequent animal responses to them in arid habitat.MethodsWe combined field vegetation surveys and Sentinel2-imagery to test if satellite spectral data can predict the spatial variability of Enneapogon over time, through GLMMs. Additionally, a cluster analysis (‘gower’ distance, ‘complete’ method), based on Enneapogon seed-productivity, and total vegetation cover in October 2016, identified three clusters: bare ground, grass dominated and shrub dominated. We compared the vegetation indices between these different clusters from October 2016 to January 2017.ResultsWe found that MSAVI2 and NDVI correlated with the proportion of Enneapogon with seeds across the landscape and this relationship changed over time. Both vegetation indices (MSAVI2 and NDVI) were higher in patches with high seed-productivity of Enneapogon than in bare soil, but only in October, a climatically-favorable period during which this dominant grass reached peak seed-productivity.DiscussionMSAVI2 and NDVI provided reliable estimates of the heterogeneity of vegetation type across the landscape only when measured in the Austral spring. This means that grass cover is related to seed-productivity and it is possible to remotely and reliably predict food resource availability in arid habitat, but only in certain conditions. The lack of significant differences between clusters in the summer was likely driven by the short-lasting nature of the vegetation in the study and the sparseness of the grass-dominated vegetation, in contrast to the shrub vegetation cluster that was particularly well measured by the NDVI.ConclusionsOverall, our study highlights the potential for Sentinel2-imagery to estimate and monitor the change in grass seed availability remotely in arid environments. However, heterogeneity in grassland cover is not as reliably measured as other types of vegetation and may only be well detected during periods of peak productivity (e.g., October 2016). creator: Caterina Funghi creator: René H.J. Heim creator: Wiebke Schuett creator: Simon C. Griffith creator: Jens Oldeland uri: https://doi.org/10.7717/peerj.9209 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Funghi et al. title: Genome-wide identification and characterization of the fibrillin gene family in Triticum aestivum link: https://peerj.com/articles/9225 last-modified: 2020-05-26 description: BackgroundThe fibrillin (FBN) gene family is highly conserved and widely distributed in the photosynthetic organs of plants. Members of this gene family are involved in the growth and development of plants and their response to biotic and abiotic stresses. Wheat (Triticum aestivum), an important food crop, has a complex genetic background and little progress has occurred in the understanding of its molecular mechanisms.MethodsIn this study, we identified 26 FBN genes in the whole genome of T. aestivum through bioinformatic tools and biotechnological means. These genes were divided into 11 subgroups and were distributed on 11 chromosomes of T. aestivum. Interestingly, most of the TaFBN genes were located on the chromosomes 2A, 2B and 2D. The gene structure of each subgroup of gene family members and the position and number of motifs were highly similar.ResultsThe evolutionary analysis results indicated that the affinities of FBNs in monocots were closer together. The tissue-specific analysis revealed that TaFBN genes were expressed in different tissues and developmental stages. In addition, some TaFBNs were involved in one or more biotic and abiotic stress responses. These results provide a basis for further study of the biological function of FBNs. creator: Yaoyao Jiang creator: Haichao Hu creator: Yuhua Ma creator: Junliang Zhou uri: https://doi.org/10.7717/peerj.9225 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Jiang et al. title: A new nematode species, Chromadorina tangaroa sp. nov. (Chromadorida: Chromadoridae) from the hull of a research vessel, New Zealand link: https://peerj.com/articles/9233 last-modified: 2020-05-26 description: Chromadorina is a globally distributed, largely marine nematode genus frequently found on a variety of organisms, including macro- and microalgae and crustaceans, as well as artificial substrates such as settlement plates and ship hulls. Here, Chromadorina tangaroa sp. nov. is described from filamentous seaweed growing on the hull of RV Tangaroa anchored in Wellington, North Island of New Zealand. It is characterized by body length 763–1,086 μm, and pore of secretory-excretory system located at or near level of teeth. Males have spicules with rounded capitulum followed by a narrower shaft and blade tapered distally, a gubernaculum as long as the spicules, and three cup-shaped precloacal supplements, and females are characterized by a cuticularized prevulvar pad, vagina located at 46–48% of body length from anterior, and vagina anteriorly directed. Chromadorina tangaroa sp. nov. is the first species of the genus to be described from New Zealand, but it is unclear whether it is native to the region because it may have dispersed as part of ship hull biofouling communities. Long-distance transport of nematodes through ship hull biofouling may be a common occurrence, but too little is known about the occurrence of nematodes on ship hulls to gauge the potential effect of shipping on nematode species distributions. creator: Daniel Leduc uri: https://doi.org/10.7717/peerj.9233 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Leduc title: Hamstring extensibility differences among elite adolescent and young dancers of different dance styles and non-dancers link: https://peerj.com/articles/9237 last-modified: 2020-05-26 description: BackgroundHamstrings have been analyzed extensively due to their tendency to shorten and their effect in the lumbo-pelvic dynamics and the sagittal position of the spine in trunk flexion with extended knees positions. It has been demonstrated that practicing a certain sport results in long-term changes in hamstring extensibility. Despite this, adequate extensibility of the hamstring musculature is essential for the dancer’s performance. Several studies have found differences in the extensibility of the hamstrings depending on the dance style, but none have compared ballet, Spanish dance and modern dance. The purpose of the present research was to analyze the differences in hamstring extensibility among professional dance students based on dance style practiced and non-dancers.MethodsThe sample was comprised of 210 students from the Professional Dance Conservatory (70 for ballet, 70 for Spanish dance and 70 for modern dance) and 70 non-dancers. For the assessment of hamstring extensibility, the angle in the passive and active straight leg raise (PSLR and ASLR, respectively) test, and the scores of the pelvic tilt in sit-and-reach (SR) test and the toe-touch (TT) test were randomly conducted.ResultsThe results showed significant differences for all the tests according to group (p < 0.001). In the PSLR and ASLR test, for both legs, and in the pelvic tilt in the SR test, the ballet dancers showed greater ranges of hamstring extensibility than the modern dancers and Spanish dancers (p ≤ 0.001). In the distance in the SR test and in the pelvic tilt in the TT test, the ballet dancers obtained higher values than the Spanish dancers (p = 0.004 and p = 0.003, respectively). The modern dancers showed higher ranges of hip flexion than the Spanish dancers in the ASLR test for both legs and in the pelvic tilt in the SR test (p from 0.007 to <0.001). Dancers showed significantly higher hamstring extensibility than non-dancers in all the tests (p < 0.001).ConclusionsThe systematic practice of dance, regardless of the style, seems to lead to high ranges of hamstring extensibility. Ballet dancers have the greatest hamstring extensibility. creator: Raquel Vaquero-Cristóbal creator: Patricia Molina-Castillo creator: Pedro A. López-Miñarro creator: Mario Albaladejo-Saura creator: Francisco Esparza-Ros uri: https://doi.org/10.7717/peerj.9237 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Vaquero-Cristóbal et al. title: Transcultural adaptation and validation of the questionnaire “Urgency, Weak stream, Incomplete emptying and Nocturia (UWIN)” for the Brazilian Portuguese link: https://peerj.com/articles/9039 last-modified: 2020-05-25 description: Lower urinary tract symptoms (LUTS) are common complaints in the adult male population and have a negative affect in the quality of life and represent an economic burden for the healthcare system worldwide. The International Prostatism Symptom Score (IPSS) is a validated tool for assessing these symptoms, but patients with low literacy may have difficulties comprehending and completing it accurately. The Urgency, Weak Stream, Incomplete Emptying, and Nocturia (UWIN) questionnaire was developed as a shorter tool in the assessment of LUTS to improve accuracy and minimize error. This study aimed at performing cross-cultural adaptation to Brazilian Portuguese and validation of UWIN questionnaire for patients with LUTS. The cross-cultural adaptation followed the steps of conceptual, item, semantic, operational, and pre-test equivalence to obtain the UWIN-Br version. The IPSS (gold standard) and UWIN-Br were coadministered, and information regarding the uroflowmetry examination was also recorded. We evaluated 306 men, median age 59 [52-66] years. There was a positive correlation r = 0.804 (p < 0.001) between the total IPSS score and the total UWIN-Br score, as well as the quality of life question (r = 0.761) (p < 0.001). The Bland-Altman plot showed good agreement between. Also, we observed that the maximum flow values decreased with the severity of the LUTS. UWIN-Br demonstrated excellent diagnostic accuracy in detecting the most severe cases. The area under the ROC curve was 89% [81–97%] 95% CI. 280 (91.5%) subjects completed the questionnaires without help, while 26 (8.5%) did so through an interview. The response time was 1.2 [1.0–1.5] min for UWIN-Br and 2.8 [2.2–3.4] min for IPSS (p < 0.001). UWIN-Br presents satisfactory and similar psychometric properties to the IPSS in the evaluation of LUTS and quality of life and is suitable for use in both clinical practice and research in our country. creator: Caroline S. Silva creator: Katia S. Freitas creator: Anna Paloma R. Ribeiro creator: Cristiano M. Gomes creator: Jose Bessa Junior uri: https://doi.org/10.7717/peerj.9039 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Silva et al. title: Identifying multiple collagen gene family members as potential gastric cancer biomarkers using integrated bioinformatics analysis link: https://peerj.com/articles/9123 last-modified: 2020-05-25 description: BackgroundGastric cancer is one of the most common malignant cancers worldwide. Despite substantial developments in therapeutic strategies, the five-year survival rate remains low. Therefore, novel biomarkers and therapeutic targets involved in the progression of gastric tumors need to be identified.MethodsWe obtained the mRNA microarray datasets GSE65801, GSE54129 and GSE79973 from the Gene Expression Omnibus database to acquire differentially expressed genes (DEGs). We used the Database for Annotation, Visualization, and Integrated Discovery (DAVID) to analyze DEG pathways and functions, and the Search Tool for the Retrieval of Interacting Genes (STRING) and Cytoscape to obtain the protein–protein interaction (PPI) network. Next, we validated the hub gene expression levels using the Oncomine database and Gene Expression Profiling Interactive Analysis (GEPIA), and conducted stage expression and survival analysis.ResultsFrom the three microarray datasets, we identified nine major hub genes: COL1A1, COL1A2, COL3A1, COL5A2, COL4A1, FN1, COL5A1, COL4A2, and COL6A3.ConclusionOur study identified COL1A1 and COL1A2 as potential gastric cancer prognostic biomarkers. creator: Zhaoxing Li creator: Zhao Liu creator: Zhiting Shao creator: Chuang Li creator: Yong Li creator: Qingwei Liu creator: Yifei Zhang creator: Bibo Tan creator: Yu Liu uri: https://doi.org/10.7717/peerj.9123 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Li et al. title: Herbicide applications increase greenhouse gas emissions of alfalfa pasture in the inland arid region of northwest China link: https://peerj.com/articles/9231 last-modified: 2020-05-25 description: Herbicides are used to control weeds in agricultural crops such as alfalfa (Medicago sativa L.), which is a forage crop. It is unclear what, if any, effect herbicides have on greenhouse gas (GHG) emissions when used on alfalfa. Our study was conducted in 2017 and 2018 to investigate the effects of two herbicides (Quizalofop-p-ethyl, QE and Bentazone, BT) on methane (CH4), carbon dioxide (CO2) and nitrous oxide (N2O) emissions from soil planted with alfalfa. QE is used to control grasses and BT is used for broadleaf weed control. Soil CO2 emissions and soil uptake of CH4 increased significantly in both years following the QE and BT treatments, although CO2 emissions differed significantly between the trial years. N2O emissions decreased relative to the control and showed no significant differences between the trial years. The application of QE and BT on alfalfa resulted in a significant increase in CO2 emissions which contributed to a significant increase in GHG emissions. The application of QE influenced GHG emissions more than BT. We demonstrated the potential effect that herbicide applications have on GHG fluxes, which are important when considering the effect of agricultural practices on GHG emissions and the potential for global warming over the next 100 years. creator: Lina Shi creator: Yarong Guo creator: Jiao Ning creator: Shanning Lou creator: Fujiang Hou uri: https://doi.org/10.7717/peerj.9231 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Shi et al. title: Revealing the selective mechanisms of inhibitors to PARP-1 and PARP-2 via multiple computational methods link: https://peerj.com/articles/9241 last-modified: 2020-05-25 description: BackgroundResearch has shown that Poly-ADP-ribose polymerases 1 (PARP-1) is a potential therapeutic target in the clinical treatment of breast cancer. An increasing number of studies have focused on the development of highly selective inhibitors that target PARP-1 over PARP-2, its closest isoform, to mitigate potential side effects. However, due to the highly conserved and similar binding sites of PARP-1 and PARP-2, there is a huge challenge for the discovery and design of PARP-1 inhibitors. Recently, it was reported that a potent PARP-1 inhibitor named NMS-P118 exhibited greater selectivity to PARP-1 over PARP-2 compared with a previously reported drug (Niraparib). However, the mechanisms underlying the effect of this inhibitor remains unclear.MethodsIn the present study, classical molecular dynamics (MD) simulations and accelerated molecular dynamics (aMD) simulations combined with structural and energetic analysis were used to investigate the structural dynamics and selective mechanisms of PARP-1 and PARP-2 that are bound to NMS-P118 and Niraparib with distinct selectivity.ResultsThe results from classical MD simulations indicated that the selectivity of inhibitors may be controlled by electrostatic interactions, which were mainly due to the residues of Gln-322, Ser-328, Glu-335, and Tyr-455 in helix αF. The energetic differences were corroborated by the results from aMD simulations.ConclusionThis study provides new insights about how inhibitors specifically bind to PARP-1 over PARP-2, which may help facilitate the design of highly selective PARP-1 inhibitors in the future. creator: Hongye Hu creator: Buran Chen creator: Danni Zheng creator: Guanli Huang uri: https://doi.org/10.7717/peerj.9241 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Hu et al. title: Agreement between the GAITRite® System and the Wearable Sensor BTS G-Walk® for measurement of gait parameters in healthy adults and Parkinson’s disease patients link: https://peerj.com/articles/8835 last-modified: 2020-05-22 description: BackgroundNowadays, the most widely used types of wearable sensors in gait analysis are inertial sensors. The aim of the study was to assess the agreement between two different systems for measuring gait parameters (inertial sensor vs. electronic walkway) on healthy control subjects (HC) and patients with Parkinson’s disease (PD).MethodsForty healthy volunteers (26 men, 14 women, mean age 58.7 ± 7.7 years) participated in the study and 24 PD patients (19 men, five women, mean age 62.7 ± 9.8 years). Each participant walked across an electronic walkway, GAITRite, with embedded pressure sensors at their preferred walking speed. Concurrently a G-Walk sensor was attached with a semi-elastic belt to the L5 spinal segment of the subject. Walking speed, cadence, stride duration, stride length, stance, swing, single support and double support phase values were compared between both systems.ResultsThe Passing-Bablock regression slope line manifested the values closest to 1.00 for cadence and stride duration (0.99 ≤ 1.00) in both groups. The slope of other parameters varied between 0.26 (double support duration in PD) and 1.74 (duration of single support for HC). The mean square error confirmed the best fit of the regression line for speed, stride duration and stride length. The y-intercepts showed higher systematic error in PD than HC for speed, stance, swing, and single support phases.ConclusionsThe final results of this study indicate that the G-Walk system can be used for evaluating the gait characteristics of the healthy subjects as well as the PD patients. However, the duration of the gait cycle phases should be used with caution due to the presence of a systematic error. creator: Slávka Vítečková creator: Hana Horáková creator: Kamila Poláková creator: Radim Krupička creator: Evžen Růžička creator: Hana Brožová uri: https://doi.org/10.7717/peerj.8835 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Vítečková et al. title: Maternal trauma and fear history predict BDNF methylation and gene expression in newborns link: https://peerj.com/articles/8858 last-modified: 2020-05-22 description: Trauma and related fear exert significant influence on mental and physical health throughout the lifespan and are associated with intergenerational patterns of development, health, and behavior. DNA methylation and gene expression are involved in our developmental adaptations to our experiences and can be influenced by social interventions. Patterns of DNA methylation and expression of a gene involved in neurodevelopment and psychiatric risk (BDNF) have been linked with childhood trauma. Given the intergenerational patterns of health and behavior, and previous links between childhood trauma and BDNF methylation and expression, this study investigated the potential for maternal history of traumatic experiences to influence development in her newborn, via changes in her newborn’s BDNF methylation and expression. We found that mothers’ trauma history was associated with epigenetic regulation of BDNF in their newborns. Moreover, the association between maternal trauma and BDNF methylation and expression patterns were moderated by newborn sex. Male newborns showed increased BDNF expression with maternal exposure to child abuse (p = .001), and increased BDNF methylation with greater maternal fear (p = .001). Female newborns showed reduced BDNF expression with greater maternal fear (p = .004). Practitioners strive to identify prevention and intervention avenues that will reduce the harmful effects of trauma. Future research should consider the potential for maternal historical trauma experiences to influence offspring DNA methylation and gene expression in a manner that could alter development and inform novel prevention strategies. creator: Stefanie R. Pilkay creator: Terri Combs-Orme creator: Frances Tylavsky creator: Nicole Bush creator: Alicia K. Smith uri: https://doi.org/10.7717/peerj.8858 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Pilkay et al. title: A new insect trackway from the Upper Jurassic—Lower Cretaceous eolian sandstones of São Paulo State, Brazil: implications for reconstructing desert paleoecology link: https://peerj.com/articles/8880 last-modified: 2020-05-22 description: The new ichnospecies Paleohelcura araraquarensis isp. nov. is described from the Upper Jurassic-Lower Cretaceous Botucatu Formation of Brazil. This formation records a gigantic eolian sand sea (erg), formed under an arid climate in the south-central part of Gondwana. This trackway is composed of two track rows, whose internal width is less than one-quarter of the external width, with alternating to staggered series, consisting of three elliptical tracks that can vary from slightly elongated to tapered or circular. The trackways were found in yellowish/reddish sandstone in a quarry in the Araraquara municipality, São Paulo State. Comparisons with neoichnological studies and morphological inferences indicate that the producer of Paleohelcura araraquarensis isp. nov. was most likely a pterygote insect, and so could have fulfilled one of the ecological roles that different species of this group are capable of performing in dune deserts. The producer could have had a herbivorous or carnivorous diet or been part of the fauna of omnivores, being able to adopt herbivorous, carnivorous, and saprophagous diets when necessary. In modern dune deserts, some species of pterygote insects are detritivores (like Tenebrionidae), relying on organic matter that accumulated among the sand grains of the dunes during dry periods with no plant growth. The presence of additional burrows suggests that the Botucatu paleodesert would have had a detritivorous fauna like this. Based on the interpretation of the ichnofossil producers, it was possible to reconstruct the food web of this paleodesert. All the omnivorous and herbivorous invertebrates and the herbivorous ornithopod dinosaurs made up the primary consumers. These animals were, in turn, the food source for bigger carnivorous or omnivorous animals unable to feed on detritus, like arachnids, possible predatory insects, mammaliaforms, and theropod dinosaurs. The highest trophic level was occupied by larger theropod dinosaurs and mammaliaforms, which, because of their size, could prey upon a wide range of animals. The producer of Paleohelcura araraquarensis isp. nov. could have been a primary consumer if it were an omnivorous detritivore or a herbivore, or a secondary consumer if it were produced by a predatory insect or an omnivore relying on animal biomass. The description of this new trackway expands the knowledge on the faunal composition of the Botucatu paleodesert and provides insights into the ecological relationships in ancient deserts. The presence of these arthropod trackways in Mesozoic eolian deposits helps to trace a continuity between Paleozoic and post-Paleozoic desert ichnofaunas, further reinforcing a single Octopodichnus—Entradichnus Ichnofacies for eolian deposits. creator: Bernardo de C.P. e M. Peixoto creator: M. Gabriela Mángano creator: Nicholas J. Minter creator: Luciana Bueno dos Reis Fernandes creator: Marcelo Adorna Fernandes uri: https://doi.org/10.7717/peerj.8880 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Peixoto et al. title: Biochemical parameters, dynamic tensiometry and circulating nucleic acids for cattle blood analysis: a review link: https://peerj.com/articles/8997 last-modified: 2020-05-22 description: The animal’s blood is the most complicated and important biological liquid for veterinary medicine. In addition to standard methods that are always in use, recent technologies such as dynamic tensiometry (DT) of blood serum and PCR analysis of particular markers are in progress. The standard and modern biochemical tests are commonly used for general screening and, finally, complete diagnosis of animal health. Interpretation of major biochemical parameters is similar across animal species, but there are a few peculiarities in each case, especially well-known for cattle. The following directions are discussed here: hematological indicators; “total protein” and its fractions; some enzymes; major low-molecular metabolites (glucose, lipids, bilirubin, etc.); cations and anions. As example, the numerous correlations between DT data and biochemical parameters of cattle serum have been obtained and discussed. Changes in the cell-free nucleic acids (cfDNA) circulating in the blood have been studied and analyzed in a variety of conditions; for example, pregnancy, infectious and chronic diseases, and cancer. CfDNA can easily be detected using standard molecular biological techniques like DNA amplification and next-generation sequencing. The application of digital PCR even allows exact quantification of copy number variations which are for example important in prenatal diagnosis of chromosomal aberrations. creator: Sergei Yu. Zaitsev creator: Nadezhda V. Bogolyubova creator: Xuying Zhang creator: Bertram Brenig uri: https://doi.org/10.7717/peerj.8997 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zaitsev et al. title: Plant-derived angiogenin fusion protein’s cytoprotective effect on trabecular meshwork damage induced by Benzalkonium chloride in mice link: https://peerj.com/articles/9084 last-modified: 2020-05-22 description: BackgroundBenzalkonium chloride (BAK), commonly used in glaucoma treatment, is an eye drop preservative with dose-dependent toxicity. Previous studies have observed the multi-functional benefits of angiogenin (ANG) against glaucoma. In our study, we evaluated ANG’s cytoprotective effect on the trabecular meshwork (TM) damage induced by BAK. Additionally, we developed a plant-derived ANG fusion protein and evaluated its effect on TM structure and function.MethodsWe synthesized plant-derived ANG (ANG-FcK) by fuzing immunoglobulin G’s Fc region and KDEL to conventional recombinant human ANG (Rh-ANG) purified from transgenic tobacco plants. We established a mouse model using BAK to look for degenerative changes in the TM, and to evaluate the protective effects of ANG-FcK and Rh-ANG. Intraocular pressure (IOP) was measured for 4 weeks and ultrastructural changes, deposition of fluorescent microbeads, type I and IV collagen, fibronectin, laminin and α-SMA expression were analyzed after the mice were euthanized.ResultsTM structural and functional degeneration were induced by 0.1% BAK instillation in mice. ANG co-treatment preserved TM outflow function, which we measured using IOP and a microbead tracer. ANG prevented phenotypic and ultrastructure changes, and that protective effect might be related to the anti-fibrosis mechanism. We observed a similar cytoprotective effect in the BAK-induced degenerative TM mouse model, suggesting that plant-derived ANG-FcK could be a promising glaucoma treatment. creator: Jae Hoon Jeong creator: Soo Jin Lee creator: Kisung Ko creator: Jeong Hwan Lee creator: Jungmook Lyu creator: Moon Hyang Park creator: Jaeku Kang creator: Jae Chan Kim uri: https://doi.org/10.7717/peerj.9084 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Jeong et al. title: Oldest co-occurrence of Varanus and Python from Africa—first record of squamates from the early Miocene of Moghra Formation, Western Desert, Egypt link: https://peerj.com/articles/9092 last-modified: 2020-05-22 description: Lizard and snake remains from the early Miocene (Burdigalian) of the Moghra Formation, Egypt, are described herein. This material comprises the first fossil remains of squamates recovered from the otherwise rich and well known vertebrate assemblage of Moghra. The material pertains to two different genera, the varanid lizard Varanus and the pythonid snake Python and adds to the so far rather poorly known squamate fossil record from Africa. On the basis of the new remains, Moghra marks the oldest so far described co-occurrence of Varanus and Python in the African continent. The close sympatry of these two genera in the African fossil record is thoroughly analyzed and discussed, a co-existence, which is still widespread in the extant herpetofauna of the continent. Situated rather close to the so called “Levantine Corridor” and dated at the Burdigalian, practically when Afro-Arabia collided with Eurasia, the Moghra squamate assemblage offers the potential of important insights in the biogeography and dispersal events of vertebrate groups during the early Miocene. creator: Georgios L. Georgalis creator: Mohamed K. Abdel Gawad creator: Safiya M. Hassan creator: Ahmed N. El-Barkooky creator: Mohamed A. Hamdan uri: https://doi.org/10.7717/peerj.9092 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Georgalis et al. title: Comparative analysis of four Zantedeschia chloroplast genomes: expansion and contraction of the IR region, phylogenetic analyses and SSR genetic diversity assessment link: https://peerj.com/articles/9132 last-modified: 2020-05-22 description: The horticulturally important genus Zantedeschia (Araceae) comprises eight species of herbaceous perennials. We sequenced, assembled and analyzed the chloroplast (cp) genomes of four species of Zantedeschia (Z. aethiopica, Z. odorata, Z. elliottiana, and Z. rehmannii) to investigate the structure of the cp genome in the genus. According to our results, the cp genome of Zantedeschia ranges in size from 169,065 bp (Z. aethiopica) to 175,906 bp (Z. elliottiana). We identified a total of 112 unique genes, including 78 protein-coding genes, 30 transfer RNA (tRNA) genes and four ribosomal RNA (rRNA) genes. Comparison of our results with cp genomes from other species in the Araceae suggests that the relatively large sizes of the Zantedeschia cp genomes may result from inverted repeats (IR) region expansion. The sampled Zantedeschia species formed a monophylogenetic clade in our phylogenetic analysis. Furthermore, the long single copy (LSC) and short single copy (SSC) regions in Zantedeschia are more divergent than the IR regions in the same genus, and non-coding regions showed generally higher divergence than coding regions. We identified a total of 410 cpSSR sites from the four Zantedeschia species studied. Genetic diversity analyses based on four polymorphic SSR markers from 134 cultivars of Zantedeschia suggested that high genetic diversity (I = 0.934; Ne = 2.371) is present in the Zantedeschia cultivars. High genetic polymorphism from the cpSSR region suggests that cpSSR could be an effective tool for genetic diversity assessment and identification of Zantedeschia varieties. creator: Shuilian He creator: Yang Yang creator: Ziwei Li creator: Xuejiao Wang creator: Yanbing Guo creator: Hongzhi Wu uri: https://doi.org/10.7717/peerj.9132 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 He et al. title: Creatinine is a biochemical marker for assessing how untrained people adapt to fitness training loads link: https://peerj.com/articles/9137 last-modified: 2020-05-22 description: BackgroundTo study the peculiarities of changes in creatinine concentration in blood serum of untrained men during the prolonged usage of training loads different in volume and intensity, and to determine the value of this biochemical marker for the assessment of adaptive body changes during fitness training.MethodsWe examined 50 untrained men aged 18–20 years who had no contraindications for practicing fitness. Taking into account the aim of the research, we divided these people into two groups: group A and group B. The research participants used training load regimes different in volume and intensity: representatives of group Aused low intensity training load regime (Ra = 0.53) and representatives of group B used high intensity training load regime (Ra = 0.72). To assess the adaptive body changes in the examined contingent, we used the anthropometry method (circumference body size) and bioimpedansometry (body composition indicators). We also determined the features of adaptation-compensatory body reactions to different training loads by means of biochemical control of creatinine concentration in blood serum.ResultsThe obtained results showed a significant increase in basal level of creatinine concentration in blood serum (by 17.6%) of group B representatives fixed after 3 months of practicing fitness in high intensity training load regime (Ra = 0.72). This group representative also revealed the most pronounced manifestation of adaptive body changes confirmed by the results of the maximal muscle strength growth (1RM) and circumference body size, which was almost 2.5 times higher than the results of group A representatives for the same period of time. The parameters indicating the load volume in a set (Wn) were almost 62.0 % higher in group A representatives in comparison with group B during all stages of control. Nonetheless, the highest creatinine concentration in blood serum (by 11.1 % (p < 0.05) was fixed in group A representatives in response to training load after 3 months of practicing. This fact testifies to the important role of the creatine phosphokinase mechanism of energy supply of muscular activity in the conditions of high volume and low intensity training load regime (Ra = 0.53).ConclusionThe analysis of the results obtained during a series of experimental studies indicates the need and feasibility of using the indicator of basal level of creatinine concentration in blood during fitness training, especially in the conditions of high intensity and low volume training load regime (Ra = 0.72), as an informative marker for assessing the process of long-term adaptation. creator: Andrii Chernozub creator: Vladimir Potop creator: Georgiy Korobeynikov creator: Olivia Carmen Timnea creator: Oleg Dubachinskiy creator: Oksana Ikkert creator: Yuriy Briskin creator: Yuriy Boretsky creator: Lesia Korobeynikova uri: https://doi.org/10.7717/peerj.9137 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chernozub et al. title: Endosymbiotic adaptations in three new bacterial species associated with Dictyostelium discoideum: Paraburkholderia agricolaris sp. nov., Paraburkholderia hayleyella sp. nov., and Paraburkholderia bonniea sp. nov link: https://peerj.com/articles/9151 last-modified: 2020-05-22 description: Here we give names to three new species of Paraburkholderia that can remain in symbiosis indefinitely in the spores of a soil dwelling eukaryote, Dictyostelium discoideum. The new species P. agricolaris sp. nov., P. hayleyella sp. nov., and P. bonniea sp. nov. are widespread across the eastern USA and were isolated as internal symbionts of wild-collected D. discoideum. We describe these sp. nov. using several approaches. Evidence that they are each a distinct new species comes from their phylogenetic position, average nucleotide identity, genome-genome distance, carbon usage, reduced length, cooler optimal growth temperature, metabolic tests, and their previously described ability to invade D. discoideum amoebae and form a symbiotic relationship. All three of these new species facilitate the prolonged carriage of food bacteria by D. discoideum, though they themselves are not food. Further studies of the interactions of these three new species with D. discoideum should be fruitful for understanding the ecology and evolution of symbioses. creator: Debra A. Brock creator: Suegene Noh creator: Alicia N.M. Hubert creator: Tamara S. Haselkorn creator: Susanne DiSalvo creator: Melanie K. Suess creator: Alexander S. Bradley creator: Mahboubeh Tavakoli-Nezhad creator: Katherine S. Geist creator: David C. Queller creator: Joan E. Strassmann uri: https://doi.org/10.7717/peerj.9151 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Brock et al. title: In silico analysis on the functional and structural impact of Rad50 mutations involved in DNA strand break repair link: https://peerj.com/articles/9197 last-modified: 2020-05-22 description: BackgroundDNA double strand break repair is important to preserve the fidelity of our genetic makeup after DNA damage. Rad50 is one of the components in MRN complex important for DNA repair mechanism. Rad50 mutations can lead to microcephaly, mental retardation and growth retardation in human. However, Rad50 mutations in human and other organisms have never been gathered and heuristically compared for their deleterious effects. It is important to assess the conserved region in Rad50 and its homolog to identify vital mutations that can affect functions of the protein.MethodIn this study, Rad50 mutations were retrieved from SNPeffect 4.0 database and literature. Each of the mutations was analyzed using various bioinformatic analyses such as PredictSNP, MutPred, SNPeffect 4.0, I-Mutant and MuPro to identify its impact on molecular mechanism, biological function and protein stability, respectively.ResultsWe identified 103 mostly occurred mutations in the Rad50 protein domains and motifs, which only 42 mutations were classified as most deleterious. These mutations are mainly situated at the specific motifs such as Walker A, Q-loop, Walker B, D-loop and signature motif of the Rad50 protein. Some of these mutations were predicted to negatively affect several important functional sites that play important roles in DNA repair mechanism and cell cycle signaling pathway, highlighting Rad50 crucial role in this process. Interestingly, mutations located at non-conserved regions were predicted to have neutral/non-damaging effects, in contrast with previous experimental studies that showed deleterious effects. This suggests that software used in this study may have limitations in predicting mutations in non-conserved regions, implying further improvement in their algorithm is needed. In conclusion, this study reveals the priority of acid substitution associated with the genetic disorders. This finding highlights the vital roles of certain residues such as K42E, C681A/S, CC684R/S, S1202R, E1232Q and D1238N/A located in Rad50 conserved regions, which can be considered for a more targeted future studies. creator: Juwairiah Remali creator: Wan Mohd Aizat creator: Chyan Leong Ng creator: Yi Chieh Lim creator: Zeti-Azura Mohamed-Hussein creator: Shazrul Fazry uri: https://doi.org/10.7717/peerj.9197 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Remali et al. title: Human milk microbiota associated with early colonization of the neonatal gut in Mexican newborns link: https://peerj.com/articles/9205 last-modified: 2020-05-22 description: BackgroundHuman milk microbiota plays a role in the bacterial colonization of the neonatal gut, which has important consequences in the health and development of the newborn. However, there are few studies about the vertical transfer of bacteria from mother to infant in Latin American populations.MethodsWe performed a cross-sectional study characterizing the bacterial diversity of 67 human milk-neonatal stool pairs by high-throughput sequencing of V3-16S rDNA libraries, to assess the effect of the human milk microbiota on the bacterial composition of the neonate’s gut at early days.ResultsHuman milk showed higher microbial diversity as compared to the neonatal stool. Members of the Staphylococcaceae and Sphingomonadaceae families were more prevalent in human milk, whereas the Pseudomonadaceae family, Clostridium and Bifidobacterium genera were in the neonatal stool. The delivery mode showed association with the neonatal gut microbiota diversity, but not with the human milk microbiota diversity; for instance, neonates born by C-section showed greater richness and diversity in stool microbiota than those born vaginally. We found 25 bacterial taxa shared by both ecosystems and 67.7% of bacteria found in neonate stool were predicted to originate from human milk. This study contributes to the knowledge of human milk and neonatal stool microbiota in healthy Mexican population and supports the idea of vertical mother-neonate transmission through exclusive breastfeeding. creator: Karina Corona-Cervantes creator: Igrid García-González creator: Loan Edel Villalobos-Flores creator: Fernando Hernández-Quiroz creator: Alberto Piña-Escobedo creator: Carlos Hoyo-Vadillo creator: Martín Noé Rangel-Calvillo creator: Jaime García-Mena uri: https://doi.org/10.7717/peerj.9205 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Corona-Cervantes et al. title: A within-subject comparison of different relaxation therapies in eliciting physiological and psychological changes in young women link: https://peerj.com/articles/9217 last-modified: 2020-05-22 description: BackgroundStress reactivity can be different in women compared to men, which might consequently influence disease risk.Stress in women may also generate adverse physiological effects on their offspring during pregnancy or lactation. The objective of this study was to compare the effects of different relaxation interventions on physiological outcomes and perceived relaxation in healthy young women, to assist in identifying the most appropriate intervention(s) for use in a subsequent trial for mothers who deliver prematurely.MethodsA within-subject study was conducted in 17 women of reproductive age comparing five different relaxation interventions (guided-imagery meditation audio (GIM), music listening (ML), relaxation lighting (RL), GIM+RL, ML+RL), with control (silence/sitting), assigned in random order over a 3–6 week period. Subjective feelings of relaxation (10-point scale), heart rate (HR), systolic and diastolic blood pressure (SBP, DBP), and fingertip temperature (FT) were measured before and after each techniqueResultsAll interventions significantly increased perceived relaxation and FT, while music also significantly reduced SBP (p < 0.05). Compared to control, HR significantly decreased following GIM (mean difference = 3.2 bpm, p < 0.05), and FT increased (mean difference = 2.2 °C, p < 0.05) and SBP decreased (mean difference = 3.3 mmHg, p < 0.01) following ML. GIM + RL followed by GIM were the most preferred interventions.ConclusionsBased on preference, simplicity, and the physiological and psychological effects, GIM and ML were identified as the most effective tools for reducing stress and improving relaxation. These techniques warrant further research in larger samples and other populations. creator: Sarah Dib creator: Jonathan C.K. Wells creator: Mary Fewtrell uri: https://doi.org/10.7717/peerj.9217 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Dib et al. title: Bioinformatics analyses of retinoblastoma reveal the retinoblastoma progression subtypes link: https://peerj.com/articles/8873 last-modified: 2020-05-21 description: IntroductionRetinoblastoma (RB) is one common pediatric malignant tumor with dismal outcomes. Heterogeneity of RB and subtypes of RB were identified but the association between the subtypes of RB and RB progression have not been fully investigated.MethodsFour public datasets were downloaded from Gene expression omnibus and normalization was performed to remove batch effect. Two public datasets were explored to obtain the RB progression gene signatures by differentially expression analysis while another two datasets were iterated for RB subtypes identification using consensus clustering. After the RB progressive subtype gene signatures were identified, we tested the diagnostic capacity of these gene signatures by receiver operation curve.ResultsThree hundreds and forty six genes that were enriched in cell cycle were identified as the progression signature in RB from two independent datasets. Four subtypes of RB were stratified by consensus clustering. A total of 21 genes from RB progression signature were differentially expressed between RB subtypes. One subtype with low expression cell division genes have less progression of all four subtypes. A panel of five RB subtype genes (CLUL1, CNGB1, ROM1, LRRC39 and RDH12) predict progression of RB.ConclusionRetinoblastoma is a highly heterogeneous tumor and the level of cell cycle related gene expression is associated with RB progression. A subpopulation of RB with high expression of visual perception has less progressive features. LRRC39 is potentially the RB progression subtype biomarker. creator: Manjing Cao creator: Sha Wang creator: Jing Zou creator: Wanpeng Wang uri: https://doi.org/10.7717/peerj.8873 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Cao et al. title: VIRdb: a comprehensive database for interactive analysis of genes/proteins involved in the pathogenesis of vitiligo link: https://peerj.com/articles/9119 last-modified: 2020-05-21 description: Vitiligo is a chronic asymptomatic disorder affecting melanocytes from the basal layer of the epidermis which leads to a patchy loss of skin color. Even though it is one of the neglected disease conditions, people suffering from vitiligo are more prone to psychological disorders. As of now, various studies have been done in order to project auto-immune implications as the root cause. To understand the complexity of vitiligo, we propose the Vitiligo Information Resource (VIRdb) that integrates both the drug-target and systems approach to produce a comprehensive repository entirely devoted to vitiligo, along with curated information at both protein level and gene level along with potential therapeutics leads. These 25,041 natural compounds are curated from Natural Product Activity and Species Source Database. VIRdb is an attempt to accelerate the drug discovery process and laboratory trials for vitiligo through the computationally derived potential drugs. It is an exhaustive resource consisting of 129 differentially expressed genes, which are validated through gene ontology and pathway enrichment analysis. We also report 22 genes through enrichment analysis which are involved in the regulation of epithelial cell differentiation. At the protein level, 40 curated protein target molecules along with their natural hits that are derived through virtual screening. We also demonstrate the utility of the VIRdb by exploring the Protein–Protein Interaction Network and Gene–Gene Interaction Network of the target proteins and differentially expressed genes. For maintaining the quality and standard of the data in the VIRdb, the gold standard in bioinformatics toolkits like Cytoscape, Schrödinger’s GLIDE, along with the server installation of MATLAB, are used for generating results. VIRdb can be accessed through “http://www.vitiligoinfores.com/”. creator: Priyansh Srivastava creator: Alakto Choudhury creator: Mehak Talwar creator: Sabyasachi Mohanty creator: Priyanka Narad creator: Abhishek Sengupta uri: https://doi.org/10.7717/peerj.9119 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Srivastava et al. title: SMRT sequencing of the full-length transcriptome of the Rhynchophorus ferrugineus (Coleoptera: Curculionidae) link: https://peerj.com/articles/9133 last-modified: 2020-05-21 description: BackgroundRed palm weevil Rhynchophorus ferrugineus (Coleoptera: Curculionidae) is one of the most destructive insects for palm trees in the world. However, its genome resources are still in the blank stage, which limits the study of molecular and growth development analysis.MethodsIn this study, we used PacBio Iso-Seq and Illumina RNA-seq to first generate transcriptome from three developmental stages of R. ferrugineus (pupa, 7th larva, female and male) to increase our understanding of the life cycle and molecular characteristics of R. ferrugineus.ResultsA total of 63,801 nonredundant full-length transcripts were generated with an average length of 2,964 bp from three developmental stages, including the 7th instar larva, pupa, female adult and male adult. These transcripts showed a high annotation rate in seven public databases, with 54,999 (86.20%) successfully annotated. Meanwhile, 2,184 alternative splicing (AS) events, 2,084 transcription factors (TFs), 66,230 simple sequence repeats (SSR) and 9,618 Long noncoding RNAs (lncRNAs) were identified. In summary, our results provide a new source of full-length transcriptional data and information for the further study of gene expression and genetics in R. ferrugineus. creator: Hongjun Yang creator: Danping Xu creator: Zhihang Zhuo creator: Jiameng Hu creator: Baoqian Lu uri: https://doi.org/10.7717/peerj.9133 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yang et al. title: Isolation, characterization, and comparative genomic analysis of a phage infecting high-level aminoglycoside-resistant (HLAR) Enterococcus faecalis link: https://peerj.com/articles/9171 last-modified: 2020-05-21 description: Enterococcus is a genus of Gram-positive bacteria that are commensal to the gastrointestinal tracts of humans but some species have been increasingly implicated as agents of nosocomial infections. The increase in infections and the spread of antibiotic-resistant strains have contributed to renewed interest in the discovery of Enterococcus phages. The aims of this study were (1) the isolation, characterization, and genome sequencing of a phage capable of infecting an antibiotic-resistant E. faecalis strain, and (2) the comparative genomic analysis of publicly-available Enterococcus phages. For this purpose, multiple phages were isolated from wastewater treatment plant (WWTP) influent using a high-level aminoglycoside-resistant (HLAR) E. faecalis strain as the host. One phage, phiNASRA1, demonstrated a high lytic efficiency (∼97.52%). Transmission electron microscopy (TEM) and whole-genome sequencing (WGS) showed that phiNASRA1 belongs to the Siphoviridae family of double-stranded DNA viruses. The phage was approximately 250 nm in length and its complete genome (40,139 bp, 34.7% GC) contained 62 open reading frames (ORFs). Phylogenetic comparisons of phiNASRA1 and 31 publicly-available Enterococcus phages, based on the large subunit terminase and portal proteins, grouped phage by provenance, size, and GC content. In particular, both phylogenies grouped phages larger than 100 kbp into distinct clades. A phylogeny based on a pangenome analysis of the same 32 phages also grouped phages by provenance, size, and GC content although agreement between the two single-locus phylogenies was higher. Per the pangenome phylogeny, phiNASRA1 was most closely related to phage LY0322 that was similar in size, GC content, and number of ORFs (40,139 and 40,934 bp, 34.77 and 34.80%, and 60 and 64 ORFs, respectively). The pangenome analysis did illustrate the high degree of sequence diversity and genome plasticity as no coding sequence was homologous across all 32 phages, and even ‘conserved’ structural proteins (e.g., the large subunit terminase and portal proteins) were homologous in no more than half of the 32 phage genomes. These findings contribute to a growing body of literature devoted to understanding phage biology and diversity. We propose that this high degree of diversity limited the value of the single-locus and pangenome phylogenies. By contrast, the high degree of homology between phages larger than 100 kbp suggests that pangenome analyses of more similar phages is a viable method for assessing subclade diversity. Future work is focused on validating phiNASRA1 as a potential therapeutic agent to eradicate antibiotic-resistant E. faecalis infections in an animal model. creator: Danial Nasr Azadani creator: Daiyuan Zhang creator: J. Robert Hatherill creator: David Silva creator: Jeffrey W. Turner uri: https://doi.org/10.7717/peerj.9171 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Nasr Azadani et al. title: Transcriptome profiling of avian pathogenic Escherichia coli and the mouse microvascular endothelial cell line bEnd.3 during interaction link: https://peerj.com/articles/9172 last-modified: 2020-05-21 description: BackgroundAvian pathogenic Escherichia coli (APEC), an important extraintestinal pathogenic E. coli, causes colibacillosis, an acute and mostly systemic disease involving multiple organ lesions such as meningitis. Meningitis-causing APEC can invade the host central nervous system by crossing the blood–brain barrier (BBB), which is a critical step in the development of meningitis. However, the bacteria-host interaction mechanism in this process remains unclear.MethodsIn this study, we examined E. coli and bEnd.3 cells transcriptomes during infection and mock infection to investigate the global transcriptional changes in both organisms using RNA sequencing approach.ResultsWhen APEC infected the bEnd.3 cells, several significant changes in the expression of genes related to cell junctional complexes, extracellular matrix degradation, actin cytoskeleton rearrangement, immune activation and the inflammatory response in bEnd.3 cells were observed as compared to the mock infection group. Thus, the immune activation of bEnd.3 cells indicated that APEC infection activated host defenses. Furthermore, APEC may exploit cell junction degradation to invade the BBB. In addition, amino acid metabolism and energy metabolism related genes were downregulated and the protein export pathway related genes were upregulated in APEC cultured with bEnd.3 cells, compared to that in control. Thus, APEC may encounter starvation and express virulence factors during incubation with bEnd.3 cells.ConclusionThis study provides a comprehensive overview of transcriptomic changes that occur during APEC infection of bEnd.3 cells, and offers insights into the bacterial invasion strategies and the subsequent host defense mechanism. creator: Peili Wang creator: Xia Meng creator: Jianji Li creator: Yanfei Chen creator: Dong Zhang creator: Haoran Zhong creator: Pengpeng Xia creator: Luying Cui creator: Guoqiang Zhu creator: Heng Wang uri: https://doi.org/10.7717/peerj.9172 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: Development and validation of a prognostic nomogram to predict overall survival and cancer-specific survival for patients with anaplastic thyroid carcinoma link: https://peerj.com/articles/9173 last-modified: 2020-05-21 description: BackgroundAnaplastic thyroid carcinoma (ATC) is a rare malignant tumor with a poor prognosis. However, there is no useful clinical prognostic predictive tool for ATC so far. Our study identified risk factors for survival of ATC and created a reliable nomogram to predict overall survival (OS) and cancer-specific survival (CSS) of patients with ATC.MethodsA total of 1,404 cases of ATC diagnosed between 1983 and 2013 were extracted from on the Surveillance, Epidemiology and End Results database based on our inclusion criteria. OS and CSS were compared among patients between each variable by Kaplan–Meier methods. The Cox proportional hazards model was used to evaluate multiple prognostic factors and obtain independent predictors. All independent risk factors were included to build nomograms, whose accuracy and practicability were tested by concordance index (C-index), calibration curves, ROC curves, DCA, net reclassification improvement (NRI) and integrated discrimination improvement (IDI).ResultsHistoric stage, tumor size, surgery and radiotherapy were independent risk factors associated with ATC according to multivariate Cox regression analysis of OS. However, gender was also an important prognostic predictor in CSS besides the factors mentioned above. These characteristics were included in the nomograms predicting OS and CSS of patients with ATC. The nomograms predicting OS and CSS performed well with a C-index of 0.765 and 0.773. ROC curves, DCA, NRI and IDI suggested that the nomogram was superior to TNM staging and age.ConclusionThe proposed nomogram is a reliable tool based on the prediction of OS and CSS for patients with ATC. Such a predictive tool can help to predict the survival of the patients. creator: Weiwei Gui creator: Weifen Zhu creator: Weina Lu creator: Chengxin Shang creator: Fenping Zheng creator: Xihua Lin creator: Hong Li uri: https://doi.org/10.7717/peerj.9173 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Gui et al. title: Extensive protein expression changes induced by pamidronate in RAW 264.7 cells as determined by IP-HPLC link: https://peerj.com/articles/9202 last-modified: 2020-05-21 description: BackgroundBisphosphonate therapy has become a popular treatment for osteoporosis, Paget’s disease, multiple myeloma, osteogenesis imperfecta, myocardial infarction, and cancer despite its serious side effects. Bisphosphonate-induced molecular signaling changes in cells are still not clearly elucidated.MethodsAs bisphosphonates are primarily engulfed by macrophages, we treated RAW 264.7 cells (a murine macrophage cell line) with pamidronate and investigated global protein expressional changes in cells by immunoprecipitation high performance liquid chromatography (IP-HPLC) using 218 antisera.ResultsPamidronate upregulated proliferation-activating proteins associated with p53/Rb/E2F and Wnt/β-catenin pathways, but downregulated the downstream of RAS signaling, pAKT1/2/3, ERK-1, and p-ERK-1, and subsequently suppressed cMyc/MAX/MAD network. However, in situ proliferation index of pamidronate-treated RAW264.7 cells was slightly increased by 3.2% vs. non-treated controls. Pamidronate-treated cells showed increase in the expressions of histone- and DNA methylation-related proteins but decrease of protein translation-related proteins. NFkB signaling was also suppressed as indicated by the down-regulations of p38 and p-p38 and the up-regulation of mTOR, while the protein expressions related to cellular protection, HSP-70, NRF2, JNK-1, and LC3 were upregulated. Consequently, pamidronate downregulated the protein expressions related to immediate inflammation,cellular differentiation, survival, angiogenesis, and osteoclastogenesis, but upregulated PARP-1 and FAS-mediated apoptosis proteins. These observations suggest pamidronate affects global protein expressions in RAW 264.7 cells by stimulating cellular proliferation, protection, and apoptosis but suppressing immediate inflammation, differentiation, osteoclastogenesis, and angiogenesis. Accordingly, pamidronate appears to affect macrophages in several ways eliciting not only its therapeutic effects but also atypical epigenetic modification, protein translation, RAS and NFkB signalings. Therefore, our observations suggest pamidronate-induced protein expressions are dynamic, and the affected proteins should be monitored by IP-HPLC to achieve the therapeutic goals during treatment. creator: Sang Shin Lee creator: Soung Min Kim creator: Yeon Sook Kim creator: Suk Keun Lee uri: https://doi.org/10.7717/peerj.9202 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Lee et al. title: Comparison between radiofrequency ablation and sublobar resections for the therapy of stage I non-small cell lung cancer: a meta-analysis link: https://peerj.com/articles/9228 last-modified: 2020-05-21 description: BackgroundSublobar resection (SLR) and radiofrequency ablation (RFA) are the two minimally invasive procedures performed for treating stage I non-small cell lung cancer (NSCLC). This study aimed to compare SLR and RFA for the treatment of stage I NSCLC using the meta-analytical method.MethodsWe searched PubMed and Embase for articles published till December 2019 to evaluate the comparative studies and assess the survival and progression-free survival rates and postoperative complications (PROSPERO registration number: CRD42018087587). A meta-analysis was performed by combining the outcomes of the reported incidences of short-term morbidity and long-term mortality. The fixed or random effects model was utilized to calculate the pooled odds ratios (OR) and the 95% confidence intervals.ResultsFour retrospective studies were considered in the course of this study. The studies included a total of 309 participants; 154 were assigned to the SLR group, and 155 were assigned to the RFA group. Moreover, there were statistically significant differences between the one- and three-year survival rates and one- and three-year progression-free survival rates for the two groups, which were in favor of the SLR group. Among the post-surgical complications, pneumothorax and pleural effusion were more common for the SLR group, while cardiac abnormalities were prevalent in the RFA group. There was no difference in prevalence of hemoptysis between SLR and RFA groups, which might be attributed to the limited study sample size.ConclusionConsidering the higher survival rates and disease control in the evaluated cases, surgical resection is the preferred treatment method for stage I NSCLC. RFA can be considered a valid alternative in patients not eligible for surgery and in high-risk patients as it is less invasive and requires shorter hospital stay. creator: Shuang Chen creator: Shize Yang creator: Shun Xu creator: Siyuan Dong uri: https://doi.org/10.7717/peerj.9228 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: Evaluation of the effective dose of amygdalin for the improvement of antioxidant gene expression and suppression of oxidative damage in mice link: https://peerj.com/articles/9232 last-modified: 2020-05-21 description: BackgroundLittle is known regarding the toxic and therapeutic doses of amygdalin. Treatment regimens and schedules can vary between humans and animal models, and there have been reports of cyanide toxicity due to amygdalin use.ObjectiveThe aim of this study was to evaluate the effect of different doses of amygdalin on antioxidant gene expression and suppression of oxidative damage in mice.MethodsForty adult male mice were divided randomly into four groups (n = 10) as follows and treated orally for two weeks: a control group treated with saline solution, a group treated with amygdalin at 200 mg/kg body weight, a group treated with amygdalin at 100 mg/kg body weight, and a group treated with amygdalin at 50 mg/kg body weight. Liver and testis samples were collected for gene expression, biochemical and histopathological analyses.ResultsThe mice treated with medium-dose amygdalin (100 mg/kg) showed upregulated mRNA expression of glutathione peroxidase (P < 0.01) and superoxide dismutase (P < 0.05) and significantly decreased lipid peroxidation (P < 0.05) in hepatic and testicular tissues compared to those in the untreated groups (controls), with mild histopathological effects. The mice treated with high-dose of amygdalin (200 mg/kg) showed downregulated mRNA expression of glutathione peroxidase and superoxide dismutase (P < 0.01) and significantly increased lipid peroxidation (P < 0.05) in both hepatic and testicular tissues compared to those in the untreated groups (controls), with an apparent effect at the histopathological level. No effects were observed in the mice treated with low-dose amygdalin (50 mg/kg) at the gene, protein and histopathological level.ConclusionLow-and medium-dose amygdalin did not induce toxicity in the hepatic and testicular tissues of male mice, unlike high-dose amygdalin, which had a negative effect on oxidative balance in mice. Therefore, amygdalin at a moderate dose may improve oxidative balance in mice. creator: Sarah Albogami creator: Aziza Hassan creator: Nibal Ahmed creator: Alaa Alnefaie creator: Afnan Alattas creator: Lama Alquthami creator: Afaf Alharbi uri: https://doi.org/10.7717/peerj.9232 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Albogami et al. title: Insights on early mutational events in SARS-CoV-2 virus reveal founder effects across geographical regions link: https://peerj.com/articles/9255 last-modified: 2020-05-21 description: Here we aim to describe early mutational events across samples from publicly available SARS-CoV-2 sequences from the sequence read archive and GenBank repositories. Up until 27 March 2020, we downloaded 50 illumina datasets, mostly from China, USA (WA State) and Australia (VIC). A total of 30 datasets (60%) contain at least a single founder mutation and most of the variants are missense (over 63%). Five-point mutations with clonal (founder) effect were found in USA next-generation sequencing samples. Sequencing samples from North America in GenBank (22 April 2020) present this signature with up to 39% allele frequencies among samples (n = 1,359). Australian variant signatures were more diverse than USA samples, but still, clonal events were found in these samples. Mutations in the helicase, encoded by the ORF1ab gene in SARS-CoV-2 were predominant, among others, suggesting that these regions are actively evolving. Finally, we firmly urge that primer sets for diagnosis be carefully designed, since rapidly occurring variants would affect the performance of the reverse transcribed quantitative PCR (RT-qPCR) based viral testing. creator: Carlos Farkas creator: Francisco Fuentes-Villalobos creator: Jose Luis Garrido creator: Jody Haigh creator: María Inés Barría uri: https://doi.org/10.7717/peerj.9255 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Farkas et al. title: High expression of stromal signatures correlated with macrophage infiltration, angiogenesis and poor prognosis in glioma microenvironment link: https://peerj.com/articles/9038 last-modified: 2020-05-20 description: Glioma is one of the most fatal tumors in central nervous system. Previous studies gradually revealed the association between tumor microenvironment and the prognosis of gliomas patients. However, the correlation between tumor-infiltrating immune cell and stromal signatures are unknown. In our study, we obtained gliomas samples from the Chinese Glioma Genome Atlas (CGGA) and The Cancer Genome Atlas (TCGA). The landscape of tumor infiltrating immune cell subtypes in gliomas was calculated by CIBERSORT. As a result, we found high infiltration of macrophages was correlated with poor outcome (P < 0.05). Then functional enrichment analysis of high/low macrophage-infiltrating groups was performed by GSEA. The results showed three gene sets includes 102 core genes about angiogenesis were detected in high macrophage-infiltrating group. Next, we constructed PPI network and analyzed prognostic value of 102 core genes. We found that five stromal signatures indicated poor prognosis which including HSPG2, FOXF1, KDR, COL3A1, SRPX2 (P < 0.05). Five stromal signatures were adopted to construct a classifier. The classifier showed powerful predictive ability (AUC = 0.748). Patients with a high risk score showed poor survival. Finally, we validated this classifier in TCGA and the result was consistent with CGGA. Our investigation of tumor microenvironment in gliomas may stimulate the new strategy in immunotherapy. Five stromal signature correlated with poor prognosis also provide a strong predator of gliomas patient outcome. creator: Yixin Tian creator: Yiquan Ke creator: Yanxia Ma uri: https://doi.org/10.7717/peerj.9038 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Tian et al. title: Distribution of the acoustic occurrence of dolphins during the summers 2011 to 2015 in the Upper Gulf of California, Mexico link: https://peerj.com/articles/9121 last-modified: 2020-05-20 description: Baseline knowledge of spatial and temporal distribution patterns is essential for cetacean management and conservation. Such knowledge is particularly important in areas where gillnet fishing occurs, as the Upper Gulf of California, which increases the probability of bycatch of cetaceans. In this area, the vaquita porpoise (Phocoena sinus) has been widely studied, but the knowledge of other cetaceans is scarce and based on traditional visual survey methods. We used data collected by an array of acoustic click detectors (C-PODs) during the summers 2011 to 2015 to analyze the distribution of dolphins in the Vaquita Refuge in the Upper Gulf of California. We recorded 120,038 echolocation click trains of dolphins during 12,371 days of recording effort at 46 sampling sites. Based on simultaneous visual and acoustic data, we estimated a false positive acoustic detection rate of 19.4%. Dolphin acoustic activity varied among sites, with higher activity in the east of the Vaquita Refuge. Acoustic activity was higher at night than during the day. We used negative binomial generalized linear models to study the count of clicks of dolphins in relation to spatial, temporal, physical, biological and anthropogenic explanatory variables. The best model selected for the response variable included sampling site, day-night condition, and vertical component of tide speed. Patterns in the spatial distribution of predicted acoustic activity of dolphins were similar to the acoustic activity observed per sampling season. Higher acoustic activity was predicted at night, but the tide speed variable was not relevant under this condition. Acoustic activity patterns could be related to the availability of prey resources since echolocation click trains are associated with foraging activities of dolphins. This is the first study of the distribution of dolphins in Mexico using medium-term systematic passive acoustic monitoring, and the results can contribute to better management to the natural protected area located in the Upper Gulf of California. creator: Gustavo Cárdenas Hinojosa creator: Horacio de la Cueva creator: Tim Gerrodette creator: Armando M. Jaramillo-Legorreta uri: https://doi.org/10.7717/peerj.9121 license: https://creativecommons.org/publicdomain/zero/1.0/ rights: title: Impact of cost distance and habitat fragmentation on the daily path length of Rhinopithecus bieti link: https://peerj.com/articles/9165 last-modified: 2020-05-20 description: An understanding of primate movement patterns in response to natural and anthropogenically induced changes in habitat heterogeneity, food availability, and plant species distribution is essential for developing effective management and conservation programs. Therefore, from July 2013 to June 2014, we examined the effects of landscape configuration on the ranging behavior (daily path length, DPL) of the Endangered Yunnan snub-nosed monkey (Rhinopithecus bieti) in the Baimaxueshan National Nature Reserve (27°34′N, 99°17′E) in Gehuaqing, China. Given the extreme difficulties in following the study group across high altitude mountainous terrain across an elevation of 2,500–4,000 m, we were only able to collect DPL using 3-4 GPS points per day on 21 individual days. We found that R. bieti traveled the shortest DPL in winter (1,141.31 m), followed by spring (2,034.06 m) and autumn (2,131.19 m). The cost distance, a statistical tool designed to estimate the difficulty of a species moving across its distributional range, was lowest in autumn (205.47), followed by spring (225.93) and winter (432.59) (one-way ANOVA: F = 3.852, P = 0.026, df = 2). The habitat fragmentation index (HFI), which measures the density of forest patches, indicated areas visited in the winter were more fragmented (HFI = 2.16) compared to spring (HFI = 1.83) or autumn (HFI = 1.3). Although our results should be considered preliminary, they suggest that both the availability of suitable travel routes and habitat fragmentation, driven by high-intensity human disturbance, constrain the movement of R. bieti. We found that undisturbed areas of the bands’ range contained a high density of lichens, which represent a nutritious and abundant and year-round food source for Yunnan snub-nosed monkeys. In order to protect this Endangered species, we recommend that researchers construct detailed maps of landscape heterogeneity, particularly habitat connectivity, forest fragmentation, and seasonal variation in the location of major food patches in order to better understand and mitigate the effects of seasonal habitat change on patterns of R. bieti habitat utilization and population viability. creator: Cong Li creator: Xumao Zhao creator: Dayong Li creator: Paul Alan Garber creator: Zuofu Xiang creator: Ming Li creator: Huijuan Pan uri: https://doi.org/10.7717/peerj.9165 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Li et al. title: The effect of landscape on functional connectivity and shell shape in the land snail Humboldtiana durangoensis link: https://peerj.com/articles/9177 last-modified: 2020-05-20 description: The populations of Humboldtiana durangoensis have experienced a drastic reduction in the effective population size; in addition, the species is threatened by anthropogenic activities. For the aforementioned, landscape genetics will serve as a tool to define the potential evolutionarily significant units (ESU) for this species. To complete our objective, we evaluated the effect of cover vegetation and climate on the functional connectivity of the species from the last glacial maximum (LGM) to the present as well as the effect of climate on shell shape. Partial Mantel tests, distance-based redundance analysis and a Bayesian framework were used to evaluate connectivity. On the other hand, geometric morphometrics, phylogenetic principal component analysis and redundancy analysis were used for the analysis of shell shape. Our results suggest that the suitable areas have been decreasing since the LGM; also, vegetation cover rather than climate has influenced the genetic connectivity among land snail populations, although temperature had a high influence on shell shape in this species. In conclusion, vegetation cover was the main factor that determined the functional connectivity for the land snail; however, local selective pressures led to different phenotypes in shell shape that allowed us to postulate that each one of the previously defined genetic groups must be considered as a different ESU. creator: Benjamín López creator: Omar Mejía creator: Gerardo Zúñiga uri: https://doi.org/10.7717/peerj.9177 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 López et al. title: A new species of freshwater crab of the genus Qianguimon Huang, 2018 (Decapoda: Brachyura: Potamidae) from Guangxi, Southern China link: https://peerj.com/articles/9194 last-modified: 2020-05-20 description: A new species of freshwater crab of the genus QianguimonHuang, 2018, is described from Guangxi Zhuang Autonomous Region, southern China. It can be distinguished from congeners by the following characters: male first gonopods bent inward at about 45° at base of terminal segment, carapace regions distinct and rugged and the female vulva opening inwards and downwards. In addition, molecular evidence derived from the 16S rRNA gene supported the species described in this study as a new species of Qianguimon. creator: Song-Bo Wang creator: Ya-Nan Zhang creator: Jie-Xin Zou uri: https://doi.org/10.7717/peerj.9194 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: Enhancement of specific T-lymphocyte responses by monocyte-derived dendritic cells pulsed with E2 protein of human papillomavirus 16 and human p16INK4A link: https://peerj.com/articles/9213 last-modified: 2020-05-20 description: IntroductionProphylactic vaccines are already available for prevention of human papillomavirus (HPV) infection. However, we still await development of therapeutic vaccines with high efficiency for stimulating specific T lymphocytes to clear HPV infection.ObjectiveThis study investigates the potential for subunits of human p16INK4a protein and E2 protein of HPV16 to stimulate dendritic cells and enhance the specific response of T lymphocytes against HPV-infected cells.MethodologyImmunogenic epitopes of HPV16 E2 and p16INK4a proteins were predicted through the common HLA class I and II alleles present in the Thai population. Then, monocyte-derived dendritic cells (MDCs) were pulsed with HPV16 E2 and/or p16INK4a protein s and their maturity assessed. MDCs pulsed with either or both of these proteins at optimal concentrations were used for activation of autologous T lymphocytes and IFN-γ production was measured for specific response function.ResultsHPV16 E2 and p16INK4a proteins contain various immunogenic epitopes which can be presented by antigen-presenting cells via both HLA class I and II molecules. The stimulation of MDCs with either HPV16 E2 or p16INK4a proteins increased percentages and mean fluorescence intensity (MFI) of CD83+ MDCs in a dose-dependent manner. An optimum concentration of 250 ng/mL and 150 ng/mL of HPV16 E2 and p16INK4a proteins, respectively, stimulated MDCs via the MAPK pathway (confirmed by use of MAPK inhibitors). T lymphocytes could be activated by MDCs pulsed with these proteins, leading to high percentages of both CD4+ IFN-γ+ T lymphocytes and CD8+ IFN-γ+ T lymphocytes. The production of IFN-γ was higher in co-cultures containing MDCs pulsed with HPV16 E2 protein than those pulsed with p16INK4a. Interestingly, MDCs pulsed with a combination of HPV16 E2 and p16INK4a significantly increased IFN-γ production of T lymphocytes. The IFN-γ production was inhibited by both HLA class I and II blockade, particularly in co-cultures with MDCs pulsed with a combination of HPV16 E2 and p16INK4a.ConclusionsThis suggests that MDCs pulsed with both proteins enhances specific response of both CD4+ and CD8+ T lymphocytes. This study might provide a strategy for further in vivo study of stimulation of T lymphocytes for therapy of HPV-associated cancer. creator: Nuchsupha Sunthamala creator: Neeranuch Sankla creator: Jureeporn Chuerduangphui creator: Piyawut Swangphon creator: Wanchareeporn Boontun creator: Supakpong Ngaochaiyaphum creator: Weerayut Wongjampa creator: Tipaya Ekalaksananan creator: Chamsai Pientong uri: https://doi.org/10.7717/peerj.9213 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Sunthamala et al. title: Non-parallel changes in songbird migration timing are not explained by changes in stopover duration link: https://peerj.com/articles/8975 last-modified: 2020-05-19 description: Shifts in the timing of animal migration are widespread and well-documented; however, the mechanism underlying these changes is largely unknown. In this study, we test the hypothesis that systematic changes in stopover duration—the time that individuals spend resting and refueling at a site—are driving shifts in songbird migration timing. Specifically, we predicted that increases in stopover duration at our study site could generate increases in passage duration—the number of days that a study site is occupied by a particular species—by changing the temporal breadth of observations and vise versa. We analyzed an uninterrupted 46-year bird banding dataset from Massachusetts, USA using quantile regression, which allowed us to detect changes in early-and late-arriving birds, as well as changes in passage duration. We found that median spring migration had advanced by 1.04 days per decade; that these advances had strengthened over the last 13 years; and that early-and late-arriving birds were advancing in parallel, leading to negligible changes in the duration of spring passage at our site (+0.07 days per decade). In contrast, changes in fall migration were less consistent. Across species, we found that median fall migration had delayed by 0.80 days per decade, and that changes were stronger in late-arriving birds, leading to an average increase in passage duration of 0.45 days per decade. Trends in stopover duration, however, were weak and negative and, as a result, could not explain any changes in passage duration. We discuss, and provide some evidence, that changes in population age-structure, cryptic geographic variation, or shifts in resource availability are consistent with increases in fall passage duration. Moreover, we demonstrate the importance of evaluating changes across the entire phenological distribution, rather than just the mean, and stress this as an important consideration for future studies. creator: Nicholas N. Dorian creator: Trevor L. Lloyd-Evans creator: J. Michael Reed uri: https://doi.org/10.7717/peerj.8975 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Dorian et al. title: Recent advances in biochar application for water and wastewater treatment: a review link: https://peerj.com/articles/9164 last-modified: 2020-05-19 description: In the past decade, researchers have carried out a massive amount of research on the application of biochar for contaminants removal from aqueous solutions. As an emerging sorbent with great potential, biochar has shown significant advantages such as the broad sources of feedstocks, easy preparation process, and favorable surface and structural properties. This review provides an overview of recent advances in biochar application in water and wastewater treatment, including a brief discussion of the involved sorption mechanisms of contaminants removal, as well as the biochar modification methods. Furthermore, environmental concerns of biochar that need to be paid attention to and future research directions are put forward to promote the further application of biochar in practical water and wastewater treatment. creator: Xiaoqing Wang creator: Zizhang Guo creator: Zhen Hu creator: Jian Zhang uri: https://doi.org/10.7717/peerj.9164 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: The ability of oriental magpies (Pica serica) to solve baited multiple-string problems link: https://peerj.com/articles/9200 last-modified: 2020-05-19 description: BackgroundBaited multiple-string problems are commonly used in avian laboratory studies to evaluate complex cognition. Several bird species possess the ability to use a string pull for obtaining food.MethodsWe initially tested and trained 11 magpies to determine whether the oriental magpie (Pica sericia) possesses the ability to solve baited multiple-string problems. Eight of the birds obtained the bait by pulling, and were selected for formal multiple-string tasks in the second stage. Second stage tests were divided into seven tasks based on string configurations.ResultsOnly two magpies were able to solve two tasks: one solved the task of parallel strings, and the other solved the task of slanted strings with the bait farther from the middle point between the two strings and selected the short string in the task of long-short strings. When faced with more difficult tasks (i.e., the task of slanted strings with the bait closer to the middle point between the two strings, the task with two crossing strings, and the task of continuity and discontinuity), the birds initially observed the tasks and chose instead to adopt simpler strategies based on the proximity principle, side bias strategies and trial-and-error learning. Our results indicate that the oriental magpie had a partial understanding of the principle of multiple-string problems but adopted simpler strategies. creator: Lin Wang creator: Jinxin Guo creator: Heng jiu Tian creator: Jinling Sui uri: https://doi.org/10.7717/peerj.9200 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: The association of EGF rs2237051 variant, serum EGF levels and generalized aggressive periodontitis: a preliminary study link: https://peerj.com/articles/9212 last-modified: 2020-05-19 description: BackgroundEpidermal growth factor (EGF) is a pro-inflammatory small peptide that stimulates cell growth, proliferation and differentiation through binding to its receptor. EGF rs2237051 and serum EGF levels have been demonstrated to be related with a variety of diseases, including several tumors and inflammatory diseases. Therefore, this study aims to investigate the association of the EGF rs2237051 variant and serum EGF levels in Chinese patients with generalized aggressive periodontitis (GAgP).Material and MethodsA case-control study was conducted among 216 patients with GAgP and 138 healthy controls. The clinical parameters of plaque index, probing depth, attachment loss and bleeding index were recorded. The EGF rs2237051 polymorphism was genotyped using time-of-flight mass spectrometry, and serum EGF levels were determined. Logistic and linear regression models were used to investigate the association between the genotypes of EGF rs2237051, serum EGF levels and GAgP risk.ResultsThe AA genotype of EGF rs2237051 showed higher risk for GAgP than the combined genotypes GG and AG (adjusted OR = 1.65, 95% CI [1.06–2.57]). Increased serum EGF levels were associated with GAgP (adjusted OR = 1.18, 95% CI [1.14–1.22]). Moreover, the serum EGF level for the AA genotype was significantly higher than that for the AG/GG genotypes in patients with GAgP (adjusted β = 4.70, 95% CI [2.09–7.31]).ConclusionWe demonstrated that EGF rs2237051 variant and the increased level of serum EGF were associated with the risk of GAgP, the serum EGF was up-regulated in patients with GAgP. It was indicated that serum EGF might be a biomarker of GAgP and EGF rs2237051 may be related to the genetic background of GAgP. creator: Xian’e Wang creator: Wenjing Li creator: Li Xu creator: Ruifang Lu creator: Huanxin Meng uri: https://doi.org/10.7717/peerj.9212 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: The case for extended lifespan in cooperatively breeding mammals: a re-appraisal link: https://peerj.com/articles/9214 last-modified: 2020-05-19 description: Recent comparative studies have suggested that cooperative breeding is associated with increases in maximum lifespan among mammals, replicating a pattern also seen in birds and insects. In this study, we re-examine the case for increased lifespan in mammalian cooperative breeders by analysing a large dataset of maximum longevity records. We did not find any consistent, strong evidence that cooperative breeders have longer lifespans than other mammals after having controlled for variation in body mass, mode of life and data quality. The only possible exception to this general trend is found in the African mole-rats (the Bathyergid family), where all members are relatively long-lived, but where the social, cooperatively breeding species appear to be much longer-lived than the solitary species. However, solitary mole-rat species have rarely been kept in captivity or followed longitudinally in the wild and so it seems likely that their maximum lifespan has been underestimated when compared to the highly researched social species. Although few subterranean mammals have received much attention in a captive or wild setting, current data instead supports a causal role of subterranean living on lifespan extension in mammals. creator: Jack Thorley uri: https://doi.org/10.7717/peerj.9214 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Thorley title: Plasma antioxidants and oxidative stress status in obese women: correlation with cardiopulmonary response link: https://peerj.com/articles/9230 last-modified: 2020-05-19 description: IntroductionA high body fat coupled with low cardiopulmonary fitness and an increase in oxidative stress has been connoted as contributing factors in developing cardiovascular comorbidities. This study aimed to investigate the correlation between antioxidants and oxidative stress status with cardiopulmonary responses in women of different body mass index (BMI).Subjects and MethodsEighty female adults were recruited and divided into three groups; normal weight (n = 23), overweight (n = 28) and obese (n = 29), according to their BMI. Blood samples were obtained prior to cardiopulmonary exercise testing. Plasma samples were separated by centrifugation and analysed for enzymatic antioxidant activity including catalase, glutathione peroxidase and superoxide dismutase. Non-enzymatic antioxidant activities were assessed using 2, 2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical scavenging and ferric reducing ability of plasma (FRAP) assays. To evaluate the oxidative stress status of subjects, levels of reactive oxygen species and malondialdehyde, the by-product of lipid peroxidation, were measured. Cardiopulmonary responses were analysed using cardiopulmonary exercise testing (CPET) which involved 15 various parameters such as peak oxygen consumption, metabolic equivalents and respiratory exchange ratio.ResultsThe obese group had significantly lower ABTS radical scavenging and FRAP activities than the normal weight group. A higher catalase activity was observed in the obese group than the normal weight group. Spearman’s correlation showed an inverse relationship between catalase and peak oxygen consumption, while partial correlation analysis showed inverse correlations between superoxide dismutase and respiratory frequency, ABTS activity and oxygen pulse, and between ABTS activity and cardiac output.ConclusionOur results demonstrate a lower cardiovascular fitness and antioxidant capacity in obese women; the higher catalase activity may be a compensatory mechanism. The negative correlations found between these two parameters may indicate the potential effect of antioxidants on the cardiopulmonary system and deserve further analysis in a larger population. Nevertheless, this study provides the basis for future studies to further explore the relationships between redox status and cardiopulmonary responses. This can potentially be used to predict future risk of developing diseases associated with oxidative stress, especially pulmonary and cardiovascular diseases. creator: Dyg Mastura Adenan creator: Zulkarnain Jaafar creator: Jaime Jacqueline Jayapalan creator: Azlina Abdul Aziz uri: https://doi.org/10.7717/peerj.9230 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Adenan et al. title: Expression and protease characterization of a conserved protein YgjD in Vibrio harveyi link: https://peerj.com/articles/9061 last-modified: 2020-05-18 description: The glycopeptidase GCP and its homologue proteins are conserved and essential for survival of bacteria. The ygjD gene (Glycopeptidase homologue) was cloned from Vibrio harveyi strain SF-1. The gene consisted of 1,017 bp, which encodes a 338 amino acid polypeptide. The nucleotide sequence similarity of the ygjD gene with that of V. harveyi FDAARGOS 107 was 95%. The ygjD gene also showed similarities of 68%, 67% and 50% with those of Salmonella enterica, Escherichia coli and Bacillus cereus. The ygjD gene was expressed in E. coli BL21 (DE3) and the recombinant YgjD was purified by Ni2+ affinity chromatography column. The purified YgjD showed a specific 37 kDa band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and exhibited protease activities of 59,000 units/mg, 53,700 units/mg and 8,100 units/mg, respectively, on N-Acetyl-L-tyrosine ethyl ester monohydrate (ATEE), N-Benzoyl-L-tyrosine ethyl ester (BTEE) and N-Benzoyl-DL-arginine-4-nitroanilide hydrochloride (BAPNA) substrates. When the conserved amino acids of His111, Glu113 and His115 in the YgjD were replaced with alanine, respectively, the protease activities of the mutants were partly decreased. The two conserved His111 and His115 of YgjD were mutated and the protein lost the protease activity, which implied that the two amino acid played very important roles in maintaining its protease activity. The addition of the purified YgjD to the culture medium of V. harveyi strain SF-1 can effectively promote the bacteria growth. These results indicated that the protease activities may be involved in the survival of bacteria. creator: Yayuan Zhang creator: Jixiang Chen creator: Yonggang Wang creator: Yanlin Li creator: Wenhong Rui creator: Jiyi Zhang creator: Dan Luo uri: https://doi.org/10.7717/peerj.9061 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: The influence of surrounding land cover on wetland habitat conditions: a case study of inland wetlands in South Korea link: https://peerj.com/articles/9101 last-modified: 2020-05-18 description: Wetland ecosystems have been globally degraded and lost due to rapid urbanization and climate change. An assessment of national scale inventory, including wetland types and conditions, is urgently required to understand the big picture of endangered wetlands, such as where they are and how they look like. We analyzed the spatial patterns of each inland wetland type (brackish wetland was included) in South Korea and the relative importance of land cover categories on wetland conditions. The wetlands were grouped into four dominant types (riverine, lake, mountain, and human-made) according to their topography. Riverine wetlands constituted the largest area (71.3%). The relative ratio of wetlands in a well-conserved condition (i.e., “A” rank) was highest in riverine wetlands (23.8%), followed by mountain wetlands (22.1%). The higher proportion of grasslands was related to a better condition ranking, but the increasing bareland area had a negative impact on wetland conditions. We also found that wetlands located near wetland protected areas tend to be in a better condition compared to remote sites. Our results further support the importance of the condition of surrounding areas for wetland conservation. creator: Ran-Young Im creator: Taekyu Kim creator: Chung-Yeol Baek creator: Chang-Su Lee creator: Song-Hyun Kim creator: Jung-Hwan Lee creator: Ji Yoon Kim creator: Gea-Jae Joo uri: https://doi.org/10.7717/peerj.9101 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Im et al. title: Mitochondrial DNA variation in the Italian Heavy Draught Horse link: https://peerj.com/articles/8996 last-modified: 2020-05-15 description: BackgroundIn the last decades, Italy as well as other developed countries have registered a decrease in the population size of many local horse breeds. The continuous crossbreeding has determined the dilution of genetic heritage of several native breeds. The Italian Heavy Draught Horse (IHD) is the only autochthonous Italian coldblooded horse among these breeds; therefore, it represents a resource to be preserved. In 1927, the first generation of this breed was officially created by crossing different Heavy Draught horses with local mares and recorded in a Studbook.MethodologyTo provide the first comprehensive overview of the genetic diversity of Italian Heavy Draught horses from Central Italy, we produced and phylogenetically analysed 52 mitochondrial DNA (mtDNA) control-region sequences. Furthermore, we evaluated data available from GenBank (N = 568) to have a more complete scenario and to understand the relationships with other European Heavy Draught horse breeds.ResultsAmong the IHD samples that were analysed, we identified ten of the 17 haplogroups described in modern horses. Most of these sequences fell into L, G, and M lineages, thus showing the overall mtDNA legacy of the ancestral mares that were probably used at the initial stages of breeding selections a long time ago. The high mitochondrial haplotype diversity (Hd = 0.969) found in our samples reflected the multiple maternal origins of the horses. Our results highlighted a considerable percentage of haplotypes shared especially with Bardigiano and Hungarian Heavy Draught breeds. Furthermore, both the presence of four unique haplotypes detected in our samples and their absence among all equine mitochondrial published data demonstrate a mitochondrial peculiarity that needs to be further investigated and preserved with careful breeding practices. creator: Hovirag Lancioni creator: Irene Cardinali creator: Andrea Giontella creator: Maria Teresa Antognoni creator: Arianna Miglio uri: https://doi.org/10.7717/peerj.8996 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Lancioni et al. title: A new genus of sinogaleaspids (Galeaspida, stem-Gnathostomata) from the Silurian Period in Jiangxi, China link: https://peerj.com/articles/9008 last-modified: 2020-05-15 description: Galeaspids are an endemic clade of jawless stem-gnathostomes known as ostracoderms. Their existence illuminates how specific characteristics developed in jawed vertebrates. Sinogaleaspids are of particular interest among the galeaspids but their monophyly is controversial because little is known about Sinogaleaspis xikengensis. Newly discovered sinogaleaspids from the Lower Silurian of Jiangxi, China provide a wealth of data and diagnostic features used to establish the new genus, Rumporostralis gen. nov., for Sinogaleaspis xikengensis. A morphological study showed that the sensory canal system of sinogaleaspids had mosaic features similar to those of three known galeaspids. There are 3–8 pairs of transverse canals in the Sinogaleaspidae, which suggests that the sensory canal system of galeaspid probably had a grid distribution with transverse canals arranged throughout the cephalic division. Phylogenetic analysis of Galeaspida supports the monophyly of the Sinogaleaspidae, consisting of Sinogaleaspis, Rumporostralis, and Anjiaspis. However, Shuyu and Meishanaspis form another monophyletic group, Shuyuidae fam. nov., which is outside all other eugaleaspidiforms. We propose a cladistically-based classification of Galeaspida based on our analysis. creator: Xianren Shan creator: Min Zhu creator: Wenjin Zhao creator: Zhaohui Pan creator: Pingli Wang creator: Zhikun Gai uri: https://doi.org/10.7717/peerj.9008 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Shan et al. title: The chemistry and histology of sexually dimorphic mental glands in the freshwater turtle, Mauremys leprosa link: https://peerj.com/articles/9047 last-modified: 2020-05-15 description: Despite evidence from anatomy, behavior and genomics indicating that the sense of smell in turtles is important, our understanding of chemical communication in this group is still rudimentary. Our aim was to describe the microanatomy of mental glands (MGs) in a freshwater turtle, Mauremys leprosa (Geoemydidae), and to assess the chemical composition of their secretions with respect to variation among individuals and between sexes. MGs are paired sac-like organs on the gular region of the neck and are dimorphic in this species with males having fully functional holocrine glands while those of females appear non-secretory and vestigial. In adult males, the glandular epithelium of the inner portion of the gland provides exocytotic products as well as cellular debris into the lumen of the gland. The contents of the lumen can be secreted through the narrow duct portion of the gland ending in an orifice on the surface of the skin. Females have invaginated structures similar in general outline to male glands, but lack a glandular epithelium. Using gas chromatography coupled to mass spectrometry, we identified a total of 61 compounds in mental gland secretions, the most numerous being carboxylic acids, carbohydrates, alkanes, steroids and alcohols. The number of compounds per individual varied widely (mean (median) ± SD = 14.54 (13) ± 8.44; min = 3; max = 40), but only cholesterol was found in all samples. We found that the relative abundances of only six chemicals were different between the sexes, although males tended to have larger amounts of particular compounds. Although the lipid fraction of mental gland secretions is rich in chemical compounds, most occur in both sexes suggesting that they are metabolic byproducts with no role in chemical signaling. However, the relative amounts of some compounds tended to be higher in males, with significantly larger amounts of two carboxylic acids and one steroid, suggesting their putative involvement in chemical communication. creator: Alejandro Ibáñez creator: Albert Martínez-Silvestre creator: Dagmara Podkowa creator: Aneta Woźniakiewicz creator: Michał Woźniakiewicz creator: Maciej Pabijan uri: https://doi.org/10.7717/peerj.9047 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Ibáñez et al. title: Spirometric traits show quantile-dependent heritability, which may contribute to their gene-environment interactions with smoking and pollution link: https://peerj.com/articles/9145 last-modified: 2020-05-15 description: Background“Quantile-dependent expressivity” refers to a genetic effect that is dependent upon whether the phenotype (e.g., spirometric data) is high or low relative to its population distribution. Forced vital capacity (FVC), forced expiratory volume in 1 second (FEV1), and the FEV1/FVC ratio are moderately heritable spirometric traits. The aim of the analyses is to test whether their heritability (h2) is constant over all quantiles of their distribution.MethodsQuantile regression was applied to the mean age, sex, height and smoking-adjusted spirometric data over multiple visits in 9,993 offspring-parent pairs and 1,930 sibships from the Framingham Heart Study to obtain robust estimates of offspring-parent (βOP), offspring-midparent (βOM), and full-sib regression slopes (βFS). Nonparametric significance levels were obtained from 1,000 bootstrap samples. βOPs were used as simple indicators of quantile-specific heritability (i.e., h2 = 2βOP/(1+rspouse), where rspouse was the correlation between spouses).ResultsβOP ± standard error (SE) decreased by 0.0009 ± 0.0003 (P = 0.003) with every one-percent increment in the population distribution of FEV1/FVC, i.e., βOP ± SE were: 0.182 ± 0.031, 0.152 ± 0.015; 0.136 ± 0.011; 0.121 ± 0.013; and 0.099 ± 0.013 at the 10th, 25th, 50th, 75th, and 90th percentiles of the FEV1/FVC distribution, respectively. These correspond to h2 ± SEs of 0.350 ± 0.060 at the 10th, 0.292 ± 0.029 at the 25th, 0.262 ± 0.020 at the 50th, 0.234 ± 0.025 at the 75th, and 0.191 ± 0.025 at the 90th percentiles of the FEV1/FVC ratio. Maximum mid-expiratory flow (MMEF) h2 ± SEs increased 0.0025 ± 0.0007 (P = 0.0004) with every one-percent increment in its distribution, i.e.: 0.467 ± 0.046, 0.467 ± 0.033, 0.554 ± 0.038, 0.615 ± 0.042, and 0.675 ± 0.060 at the 10th, 25th, 50th, 75th, and 90th percentiles of its distribution. This was due to forced expiratory flow at 75% of FVC (FEF75%), whose quantile-specific h2 increased an average of 0.0042 ± 0.0008 for every one-percent increment in its distribution. It is speculated that previously reported gene-environment interactions may be partially attributable to quantile-specific h2, i.e., greater heritability in individuals with lower FEV1/FVC due to smoking or airborne particles exposure vs. nonsmoking, unexposed individuals.ConclusionHeritabilities of FEV1/FVC, MMEF, and FEF75% from quantile-regression of offspring-parent and sibling spirometric data suggest their quantile-dependent expressivity. creator: Paul T. Williams uri: https://doi.org/10.7717/peerj.9145 license: https://creativecommons.org/publicdomain/zero/1.0/ rights: title: Depigmenting potential of lichen extracts evaluated by in vitro and in vivo tests link: https://peerj.com/articles/9150 last-modified: 2020-05-15 description: Melanin is the main pigment of human skin, playing the primary role of protection from ultraviolet radiation. Alteration of the melanin production may lead to hyperpigmentation diseases, with both aesthetic and health consequences. Thus, suppressors of melanogenesis are considered useful tools for medical and cosmetic treatments. A great interest is focused on natural sources, aimed at finding safe and quantitatively available depigmenting substances. Lichens are thought to be possible sources of this kind of compounds, as the occurrence of many phenolic molecules suggests possible effects on phenolase enzymes involved in melanin synthesis, like tyrosinase. In this work, we used four lichen species, Cetraria islandica Ach., Flavoparmelia caperata Hale, Letharia vulpina (L.) Hue, and Parmotrema perlatum (Hudson) M. Choisy, to obtain extracts in solvents of increasing polarity, viz. chloroform, chloroform-methanol, methanol, and water. Cell-free, tyrosinase inhibition experiments showed highest inhibition for L. vulpina methanol extract, followed by C. islandica chloroform-methanol one. Comparable results for depigmenting activities were observed by means of in vitro and in vivo systems, such as MeWo melanoma cells and zebrafish larvae. Our study provides first evidence of depigmenting effects of lichen extracts, from tyrosinase inhibition to cell and in vivo models, suggesting that L. vulpina and C. islandica extracts deserve to be further studied for developing skin-whitening products. creator: Paola Malaspina creator: Erica Catellani creator: Bruno Burlando creator: Daniele Brignole creator: Laura Cornara creator: Miriam Bazzicalupo creator: Simona Candiani creator: Valentina Obino creator: Vincenzo De Feo creator: Lucia Caputo creator: Paolo Giordani uri: https://doi.org/10.7717/peerj.9150 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Malaspina et al. title: Comparing race, gender, age, and career categories in recognizing and grouping tasks link: https://peerj.com/articles/9156 last-modified: 2020-05-15 description: The purpose of our research was to compare how participants weighed age, gender, race, and career categories in recognizing and grouping tasks. In Study 1, we used a category recognition task to compare participants’ speeds in recognizing information from different categories. The results showed that participants recognized the gender information most quickly, followed by career, race, and age information. In Study 2, a categorization task was used to compare participants’ category preferences. The results showed that the career category had the greatest weight, and the gender category had the lowest weight. Two targets who had different career identities were more possible considered as belonging to different groups than two targets with different gender, race or age identities. Our results have implications in understanding the weight of different categories, with gender and career category are the most important category that affects perception and evaluation. creator: Jingjing Song creator: Lin Li uri: https://doi.org/10.7717/peerj.9156 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Song and Li title: Personalized analysis of breast cancer using sample-specific networks link: https://peerj.com/articles/9161 last-modified: 2020-05-15 description: Breast cancer is a disease with high heterogeneity. Cancer is not usually caused by a single gene, but by multiple genes and their interactions with others and surroundings. Estimating breast cancer-specific gene–gene interaction networks is critical to elucidate the mechanisms of breast cancer from a biological network perspective. In this study, sample-specific gene–gene interaction networks of breast cancer samples were established by using a sample-specific network analysis method based on gene expression profiles. Then, gene–gene interaction networks and pathways related to breast cancer and its subtypes and stages were further identified. The similarity and difference among these subtype-related (and stage-related) networks and pathways were studied, which showed highly specific for subtype Basal-like and Stages IV and V. Finally, gene pairwise interactions associated with breast cancer prognosis were identified by a Cox proportional hazards regression model, and a risk prediction model based on the gene pairs was established, which also performed very well on an independent validation data set. This work will help us to better understand the mechanism underlying the occurrence of breast cancer from the sample-specific network perspective. creator: Ke Zhu creator: Cong Pian creator: Qiong Xiang creator: Xin Liu creator: Yuanyuan Chen uri: https://doi.org/10.7717/peerj.9161 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zhu et al. title: Application of High Resolution Melt analysis (HRM) for screening haplotype variation in a non-model plant genus: Cyclopia (Honeybush) link: https://peerj.com/articles/9187 last-modified: 2020-05-15 description: AimThis study has three broad aims: to (a) develop genus-specific primers for High Resolution Melt analysis (HRM) of members of Cyclopia Vent., (b) test the haplotype discrimination of HRM compared to Sanger sequencing, and (c) provide an example of using HRM to detect novel haplotype variation in wild C. subternata Vogel. populations.LocationThe Cape Floristic Region (CFR), located along the southern Cape of South Africa.MethodsPolymorphic loci were detected through a screening process of sequencing 12 non-coding chloroplast DNA segments across 14 Cyclopia species. Twelve genus-specific primer combinations were designed around variable cpDNA loci, four of which failed to amplify under PCR; the eight remaining were applied to test the specificity, sensitivity and accuracy of HRM. The three top performing HRM Primer combinations were then applied to detect novel haplotypes in wild C. subternata populations, and phylogeographic patterns of C. subternata were explored.ResultsWe present a framework for applying HRM to non-model systems. HRM accuracy varied across the PCR products screened using the genus-specific primers developed, ranging between 56 and 100%. The nucleotide variation failing to produce distinct melt curves is discussed. The top three performing regions, having 100% specificity (i.e. different haplotypes were never grouped into the same cluster, no false negatives), were able to detect novel haplotypes in wild C. subternata populations with high accuracy (96%). Sensitivity below 100% (i.e. a single haplotype being clustered into multiple unique groups during HRM curve analysis, false positives) was resolved through sequence confirmation of each cluster resulting in a final accuracy of 100%. Phylogeographic analyses revealed that wild C. subternata populations tend to exhibit phylogeographic structuring across mountain ranges (accounting for 73.8% of genetic variation base on an AMOVA), and genetic differentiation between populations increases with distance (p < 0.05 for IBD analyses).ConclusionsAfter screening for regions with high HRM clustering specificity—akin to the screening process associated with most PCR based markers—the technology was found to be a high throughput tool for detecting genetic variation in non-model plants. creator: Nicholas C. Galuszynski creator: Alastair J. Potts uri: https://doi.org/10.7717/peerj.9187 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Galuszynski and Potts title: The prevalence of low back pain among female hospital staff at childbearing age link: https://peerj.com/articles/9199 last-modified: 2020-05-15 description: BackgroundLow back pain (LBP) is considered the most common work-related musculoskeletal disorder among female healthcare workers. The aim of this study is to compare the prevalence of LBP and non-ergonomic risk factors between female nurses, office workers, and patient transporters, and the effect of pain on job performance.MethodsBased on semi-structured interviews, we conducted a cross-sectional study on Jordanian female hospital workers between January and July, 2017.ResultsWe included 209 participants with a mean age of 35.57 ± 8 years from four Jordanian medical centers. Nurses have significantly higher frequency of LBP (82.5%; p = 0.05) compared to both office workers (67.5%) and patient transporters (68.6%). The mean difference in pain score using Visual Analogue Scale (VAS) after treatment varied significantly (p = 0.003), since it was 28.2 (±35.4) for office workers, compared to 22.8 (±26.5) for nurses and 6.5 (±33.7) for patient transporters. A higher frequency of nurses reported that LBP affected their job performance (64.9%; p = 0.013), and 43.3% of them reported having previous sick leaves due to LBP (p = 0.008).ConclusionsLBP is common among female hospital workers, with significantly higher prevalence among female nurses when compared to other female hospital staff. creator: Fadi Al-Hadidi creator: Isam Bsisu creator: Bassem Haddad creator: Saif Aldeen AlRyalat creator: Mamoun Shaban creator: Nada Matani creator: Sondos Dehidi creator: Yasmeen Khater creator: Rana Shahrouri creator: Tasnim Al Muzayen creator: Hashem Al Hawamdeh uri: https://doi.org/10.7717/peerj.9199 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Al-Hadidi et al. title: fRNAkenseq: a fully powered-by-CyVerse cloud integrated RNA-sequencing analysis tool link: https://peerj.com/articles/8592 last-modified: 2020-05-14 description: BackgroundDecreasing costs make RNA sequencing technologies increasingly affordable for biologists. However, many researchers who can now afford sequencing lack access to resources necessary for downstream analysis. This means that even as algorithms to process RNA-Seq data improve, many biologists still struggle to manage the sheer volume of data produced by next generation sequencing (NGS) technologies. Scalable bioinformatics tools that exploit multiple platforms are needed to democratize bioinformatics resources in the sequencing era. This is essential for equipping many research groups in the life sciences with the tools to process the increasingly unwieldy datasets they produce.MethodsOne strategy to address this challenge is to develop a modern generation of sequence analysis tools capable of seamless data sharing and communication. Such tools will provide interoperability through offerings of interlinked resources. Systems of interlinked, scalable resources, which often incorporate cloud data storage, are broadly referred to as cyberinfrastructure. Cyberinfrastructure integrated tools will help researchers to robustly analyze large scale datasets by efficiently sharing data burdens across a distributed architecture. Additionally, interoperability will allow emerging tools to cross-adapt features of existing tools. It is important that these tools are designed to be easy to use for biologists.ResultsWe introduce fRNAkenseq, a powered-by-CyVerse RNA sequencing analysis tool that exhibits interoperability with other resources and meets the needs of biologists for comprehensive, easy to use RNA sequencing analysis. fRNAkenseq leverages a complex set of Application Programming Interfaces (APIs) associated with the NSF-funded cyberinfrastructure project, CyVerse, to execute FASTQ-to-differential expression RNA-Seq analyses. Integrating across bioinformatics platforms, fRNAkenseq also exploits cloud integration and cross-talk with another CyVerse associated tool, CoGe. fRNAkenseq offers novel features for the biologist such as more robust and comprehensive pipelines for enrichment than those currently available by default in a single tool, whether they are cloud-based or local installation. Importantly, cross-talk with CoGe allows fRNAkenseq users to execute RNA-Seq pipelines on an inventory of 47,000 archived genomes stored in CoGe or upload their own draft genome. creator: Allen Hubbard creator: Matthew Bomhoff creator: Carl J. Schmidt uri: https://doi.org/10.7717/peerj.8592 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Hubbard et al. title: Transcriptomic changes across the life cycle of Trypanosoma cruzi II link: https://peerj.com/articles/8947 last-modified: 2020-05-14 description: Trypanosoma cruzi is a flagellated protozoan that causes Chagas disease; it presents a complex life cycle comprising four morphological stages: epimastigote (EP), metacyclic trypomastigote (MT), cell-derived trypomastigote (CDT) and amastigote (AM). Previous transcriptomic studies on three stages (EPs, CDTs and AMs) have demonstrated differences in gene expressions among them; however, to the best of our knowledge, no studies have reported on gene expressions in MTs. Therefore, the present study compared differentially expressed genes (DEGs), and signaling pathway reconstruction in EPs, MTs, AMs and CDTs. The results revealed differences in gene expressions in the stages evaluated; these differences were greater between MTs and AMs-PTs. The signaling pathway that presented the highest number of DEGs in all the stages was associated with ribosomes protein profiles, whereas the other related pathways activated were processes related to energy metabolism from glucose, amino acid metabolism, or RNA regulation. However, the role of autophagy in the entire life cycle of T. cruzi and the presence of processes such as meiosis and homologous recombination in MTs (where the expressions of SPO11 and Rad51 plays a role) are crucial. These findings represent an important step towards the full understanding of the molecular basis during the life cycle of T. cruzi. creator: Lissa Cruz-Saavedra creator: Gustavo A. Vallejo creator: Felipe Guhl creator: Juan David Ramírez uri: https://doi.org/10.7717/peerj.8947 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Cruz-Saavedra et al. title: An interaction mechanism for the maintenance of fission–fusion dynamics under different individual densities link: https://peerj.com/articles/8974 last-modified: 2020-05-14 description: Animals often show high consistency in their social organisation despite facing changing environmental conditions. Especially in shoaling fish, fission–fusion dynamics that describe for which periods individuals are solitary or social have been found to remain unaltered even when density changed. This compensatory ability is assumed to be an adaptation towards constant predation pressure, but the mechanism through which individuals can actively compensate for density changes is yet unknown. The aim of the current study is to identify behavioural patterns that enable this active compensation. We compared the fission–fusion dynamics of two populations of the live-bearing Atlantic molly (Poecilia mexicana) that live in adjacent habitats with very different predator regimes: cave mollies that inhabit a low-predation environment inside a sulfidic cave with a low density of predatory water bugs (Belostoma sp.), and mollies that live directly outside the cave (henceforth called “surface” mollies) in a high-predation environment. We analysed their fission–fusion dynamics under two different fish densities of 12 and 6 fish per 0.36 m2. As expected, surface mollies spent more time being social than cave mollies, and this difference in social time was a result of surface mollies being less likely to discontinue social contact (once they had a social partner) and being more likely to resume social contact (once alone) than cave mollies. Interestingly, surface mollies were also less likely to switch among social partners than cave mollies. A random walk simulation predicted each population to show reduced social encounters in the low density treatment. While cave mollies largely followed this prediction, surface mollies maintained their interaction probabilities even at low density. Surface mollies achieved this by a reduction in the size of a convex polygon formed by the group as density decreased. This may allow them to largely maintain their fission–fusion dynamics while still being able to visit large parts of the available area as a group. A slight reduction (21%) in the area visited at low densities was also observed but insufficient to explain how the fish maintained their fission–fusion dynamics. Finally, we discuss potential movement rules that could account for the reduction of polygon size and test their performance. creator: David Bierbach creator: Stefan Krause creator: Pawel Romanczuk creator: Juliane Lukas creator: Lenin Arias-Rodriguez creator: Jens Krause uri: https://doi.org/10.7717/peerj.8974 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Bierbach et al. title: Full-length transcriptome and targeted metabolome analyses provide insights into defense mechanisms of Malus sieversii against Agrilus mali link: https://peerj.com/articles/8992 last-modified: 2020-05-14 description: Malus sieversii is the wild progenitor for many cultivars of domesticated apple and an important germplasm resource for breeding. However, this valuable species faces a significant threat in the areas north of the Tianshan Mountains in China, by the invasion of Agrilus mali, a destructive pest of apple trees belonging to the family Buprestidae. Our preliminary study has has shown that there may be resistance to this insect in M. sieversii plants in the field, but the corresponding molecular mechanisms remain unclear. In this study, we compared the response of insect-resistant and insect-susceptible plants of M. sieversii to insect feeding using full-length transcriptome and targeted metabolome. 112,103 non-chimeric full-length reads (FLNC) totaling 10.52 Gb of data were generating with Pacific Biosciences SingleMolecule, Real-Time (PacBio SMRT) sequencing. A total of 130.06 Gb data of long reads were acquired with an Illumina HiSeq. Function annotation indicated that the different expressed genes (DEGs) were mainly involved in signal transduction pathway of plant hormones and in the synthesis of compounds such as terpenes, quinones, flavonoids, and jasmonic acid. Through targeted metabolome analysis resistant strains showed higher levels of trans-cinnamic acid, caffeine and ferulic acid after pest infestation. This study helps to decipher the transcriptional changes and related signaling paths in M. sieversii after an insect feeding, which lays a foundation for further research on molecular mechanisms of insect resistance in apples. creator: Chuang Mei creator: Jie Yang creator: Peng Yan creator: Ning Li creator: Kai Ma creator: Aisajan Mamat creator: Liqun Han creator: Qinglong Dong creator: Ke Mao creator: Fengwang Ma creator: Jixun Wang uri: https://doi.org/10.7717/peerj.8992 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Mei et al. title: Individual behavioral type captured by a Bayesian model comparison of cap making by sponge crabs link: https://peerj.com/articles/9036 last-modified: 2020-05-14 description: ‘Animal personality’ is considered to be developed through complex interactions of an individual with its surrounding environment. How can we quantify the ‘personality’ of an individual? Quantifying intra- and inter-individual variability of behavior, or individual behavioral type, appears to be a prerequisite in the study of animal personality. We propose a statistical method from a predictive point of view to measure the appropriateness of our assumption of ‘individual’ behavior in repeatedly measured behavioral data from several individuals. For a model case, we studied the sponge crab Lauridromia dehaani known to make and carry a ‘cap’ from a natural sponge for camouflage. Because a cap is most likely to be rebuilt and replaced repeatedly, we hypothesized that each individual crab would grow a unique behavioral type and it would be observed under an experimentally controlled environmental condition. To test the hypothesis, we conducted behavioral experiments and employed a new Bayesian model-based comparison method to examine whether crabs have individual behavioral types in the cap making behavior. Crabs were given behavioral choices by using artificial sponges of three different sizes. We modeled the choice of sponges, size of the trimmed part of a cap, size of the cavity of a cap, and the latency to produce a cap, as random variables in 26 models, including hierarchical models specifying the behavioral types. In addition, we calculated the marginal-level widely applicable information criterion (mWAIC) values for hierarchical models to evaluate and compared them with the non-hierarchical models from the predictive point of view. As a result, the crabs of less than about 9 cm in size were found to make caps from the sponges. The body size explained the behavioral variables namely, choice, trimmed cap characteristics, and cavity size, but not latency. Furthermore, we captured the behavioral type as a probabilistic distribution structure of the behavioral data by comparing WAIC. Our statistical approach is not limited to behavioral data but is also applicable to physiological or morphological data when examining whether some group structure exists behind fluctuating empirical data. creator: Keita Harada creator: Naoki Hayashi creator: Katsushi Kagaya uri: https://doi.org/10.7717/peerj.9036 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Harada et al. title: Identification of two functional xyloglucan galactosyltransferase homologs BrMUR3 and BoMUR3 in brassicaceous vegetables link: https://peerj.com/articles/9095 last-modified: 2020-05-14 description: Xyloglucan (XyG) is the predominant hemicellulose in the primary cell walls of most dicotyledonous plants. Current models of these walls predict that XyG interacts with cellulose microfibrils to provide the wall with the rigidity and strength necessary to maintain cell integrity. Remodeling of this network is required to allow cell elongation and plant growth. In this study, homologs of Arabidopsis thaliana MURUS3 (MUR3), which encodes a XyG-specific galactosyltransferase, were obtained from Brassica rapa (BrMUR3) to Brassica oleracea (BoMUR3). Genetic complementation showed that BrMUR3 and BoMUR3 rescue the phenotypic defects of the mur3-3 mutant. Xyloglucan subunit composition analysis provided evidence that BrMUR3 and BoMUR3 encode a galactosyltransferase, which transfers a galactose residue onto XyG chains. The detection of XXFG and XLFG XyG subunits (restoration of fucosylated side chains) in mur3-3 mutants overexpressing BrMUR3 or BoMUR3 show that MUR3 from Brassica to Arabidopsis are comparable as they add Gal to the third xylosyl residue of the XXXG subunit. Our results provide additional information for functional dissection and evolutionary analysis of MUR3 genes derived from brassicaceous species. creator: Meng Wang creator: Zongchang Xu creator: Shuaiqiang Guo creator: Gongke Zhou creator: Malcolm ONeill creator: Yingzhen Kong uri: https://doi.org/10.7717/peerj.9095 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: Metabolomics window into the role of acute kidney injury after coronary artery bypass grafting in diabetic nephropathy progression link: https://peerj.com/articles/9111 last-modified: 2020-05-14 description: IntroductionMetabolomics has emerged as a valuable tool to discover novel biomarkers and study the pathophysiology of diabetic nephropathy (DN). However, the effect of postoperative acute kidney injury (AKI) on diabetes mellitus (DM) to chronic DN progression has not been evaluated from the perspective of metabolomics.MethodsA group of type 2 diabetes mellitus (T2DM) inpatients, who underwent off-pump coronary artery bypass grafting (CABG), were enrolled in our study. According to whether postoperative AKI occurred, patients were grouped in either the AKI group (AKI, n = 44) or the non-AKI group (NAKI, n = 44). Urine samples were collected from these patients before and 24 h after operation. Six patients from the AKI group and six patients from the NAKI group were chosen as the pilot cohort for untargeted metabolomics analysis, with the goal of identifying postoperative AKI-related metabolites. To understand the possible role of these metabolites in the chronic development of renal injury among T2DM patients, trans-4-hydroxy-L-proline and azelaic acid were quantified by targeted metabolomics analysis among 38 NAKI patients, 38 AKI patients, 46 early DN patients (DN-micro group), and 34 overt DN patients (DN-macro group).ResultsUntargeted metabolomics screened 61 statistically distinguishable metabolites in postoperative urine samples, compared with preoperative urine samples. Via Venn diagram analysis, nine of 61 were postoperative AKI-related metabolites, including trans-4-hydroxy-L-proline, uridine triphosphate, p-aminobenzoate, caffeic acid, adrenochrome, δ-valerolactam, L-norleucine, 5′-deoxy-5′-(methylthio) adenosine, and azelaic acid. By targeted metabolomics analysis, the level of trans-4-hydroxy-L-proline increased gradually from the NAKI group to the AKI, DN-micro, and DN-macro groups. For azelaic acid, the highest level was found in the NAKI and DN-micro groups, followed by the DN-macro group. The AKI group exhibited the lowest level of azelaic acid.ConclusionsThe detection of urinary trans-4-hydroxy-L-proline after AKI could be treated as an early warning of chronic DN progression and might be linked to renal fibrosis. Urinary azelaic acid can be used to monitor renal function noninvasively in DM and DN patients. Our results identified markers of AKI on DM and the chronic progression of DN. In addition, the progression of DN was associated with AKI-like episodes occurring in DM. creator: Jiayi Wang creator: Wenzhe Yan creator: Xiang Zhou creator: Yu Liu creator: Chengyuan Tang creator: Youming Peng creator: Hong Liu creator: Lin Sun creator: Li Xiao creator: Liyu He uri: https://doi.org/10.7717/peerj.9111 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: Genome-wide identification and characterization of TCP family genes in Brassica juncea var. tumida link: https://peerj.com/articles/9130 last-modified: 2020-05-14 description: BackgroundTeosinte branched1/Cycloidea/proliferating cell factors (TCPs) are plant-specific transcription factors widely involved in leaf development, flowering, shoot branching, the circadian rhythm, hormone signaling, and stress responses. However, the TCP function in Brassica juncea var. tumida, the tumorous stem mustard, has not yet been reported. This study identified and characterized the entire TCP family members in B. juncea var. tumida.MethodsWe identified 62 BjTCP genes from the B. juncea var. tumida genome and analyzed their phylogenetic relationship, gene structure, protein motifs, chromosome location, and expression profile in different tissues.ResultsOf the 62 BjTCP genes we identified in B. juncea var. tumida, containing 34 class I and 28 class II subfamily members, 61 were distributed on 18 chromosomes. Gene structure and conserved motif analysis showed that the same clade genes displayed a similar exon/intron gene structure and conserved motifs. Cis-acting element results showed that the same clade genes also had a similar cis-acting element; however, subtle differences implied a different regulatory pathway. The BjTCP18s members were low-expressed in Dayejie strains and the unswelling stage of Yonganxiaoye strains. Treatment with gibberellin (GA) and salicylic acid (SA) showed that GA and SA affect the expression levels of multiple TCP genes.ConclusionWe performed the first genome-wide analysis of the TCP gene family of B. juncea var. tumida. Our results have provided valuable information for understanding the classification and functions of TCP genes in B. juncea var. tumida. creator: Jing He creator: Xiaohong He creator: Pingan Chang creator: Huaizhong Jiang creator: Daping Gong creator: Quan Sun uri: https://doi.org/10.7717/peerj.9130 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 He et al. title: Drivers of beta diversity in modern and ancient reef-associated soft-bottom environments link: https://peerj.com/articles/9139 last-modified: 2020-05-14 description: Beta diversity, the compositional variation among communities, is often associated with environmental gradients. Other drivers of beta diversity include stochastic processes, priority effects, predation, or competitive exclusion. Temporal turnover may also explain differences in faunal composition between fossil assemblages. To assess the drivers of beta diversity in reef-associated soft-bottom environments, we investigate community patterns in a Middle to Late Triassic reef basin assemblage from the Cassian Formation in the Dolomites, Northern Italy, and compare results with a Recent reef basin assemblage from the Northern Bay of Safaga, Red Sea, Egypt. We evaluate beta diversity with regard to age, water depth, and spatial distance, and compare the results with a null model to evaluate the stochasticity of these differences. Using pairwise proportional dissimilarity, we find very high beta diversity for the Cassian Formation (0.91 ± 0.02) and slightly lower beta diversity for the Bay of Safaga (0.89 ± 0.04). Null models show that stochasticity only plays a minor role in determining faunal differences. Spatial distance is also irrelevant. Contrary to expectations, there is no tendency of beta diversity to decrease with water depth. Although water depth has frequently been found to be a key factor in determining beta diversity, we find that it is not the major driver in these reef-associated soft-bottom environments. We postulate that priority effects and the biotic structuring of the sediment may be key determinants of beta diversity. creator: Vanessa Julie Roden creator: Martin Zuschin creator: Alexander Nützel creator: Imelda M. Hausmann creator: Wolfgang Kiessling uri: https://doi.org/10.7717/peerj.9139 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Roden et al. title: A low caffeine dose improves maximal strength, but not relative muscular endurance in either heavier-or lighter-loads, or perceptions of effort or discomfort at task failure in females link: https://peerj.com/articles/9144 last-modified: 2020-05-14 description: BackgroundThe body of literature considering caffeine as an ergogenic aid has primarily considered typically aerobic based exercise, male participants and moderate-to large-caffeine doses. With this in mind the aim of this project was to explore the effects of a low-caffeine dose upon maximal voluntary contraction (MVC) and muscular endurance (time to task failure, TTF) at heavier-and lighter-loads.MethodsNineteen physically active, habitual caffeine consuming females randomly performed four testing conditions; two with a low-dose of caffeine (100 mg equating to mean = 1.5 ± 0.18 mg·kg−1) and two placebo conditions, where they performed a maximal strength test (MVC) knee extension at 45° followed by a task of relative muscular endurance (sustained isometric contraction for TTF) using either heavier-(70% MVC) and lighter-(30% MVC) loads. Each participant performed each load condition following both caffeine and placebo consumption. Immediately following cessation of the muscular endurance test participants were asked to report their rating of perceived effort (RPE) and rating of perceived discomfort (RPD).ResultsAnalyses revealed a significant effect for caffeine upon MVC compared to placebo (p = 0.007). We also found a significantly greater TTF for the lighter-compared to the heavier-load condition (p < 0.0001); however, there was no significant effect comparing caffeine to placebo (p = 0.2368), but insufficient precision of estimates to infer equivalence in either lighter-(p = 0.750) or heavier-load (p = 0.262) conditions. There were no statistically significant effects for caffeine compared with placebo, or lighter-compared with heavier-loads, for RPE and RPD (all p > 0.05). RPE was statistically equivalent between caffeine and placebo for both lighter-(p = 0.007) and heavier-load (p = 0.002) conditions and RPD for heavier-(p = 0.006) but not lighter-load (p = 0.136).DiscussionThis is the first study to demonstrate a positive effect on strength from a low caffeine dose in female participants. However, it is unclear whether caffeine positively impacts upon relative muscular endurance in either heavier-or lighter-loads. Further, both RPE and RPD appear to be relatively similar during isometric tasks performed to task failure independently of caffeine supplementation or load. These findings may have implications for persons wishing to avoid side-effects or withdrawal symptoms associated with larger caffeine doses whilst still attaining the positive strength responses. creator: Georgina Waller creator: Melissa Dolby creator: James Steele creator: James P. Fisher uri: https://doi.org/10.7717/peerj.9144 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Waller et al. title: Effects of rumen bypass melatonin feeding (RBMF) on milk quality and mastitis of Holstein cows link: https://peerj.com/articles/9147 last-modified: 2020-05-14 description: Cow mastitis is a major problem frequently encountered by dairy farmers and it is manifested by the high number of somatic cells and the low quality of the milk. The conventional treatment for mastitis is use of antibiotics. In the current study, a new approach is applied to target this disorder: rumen bypass melatonin feeding (RBMF). The RBMF significantly reduced milk somatic cell count and improved milk nutritional values with the elevated protein, fat and dry matter levels. This approach also suppresses the stress and proinflammatory responses of the cows indicated by the reduced serum cortisol, TNF-α and IL-6 and increased IL-10 levels. Importantly, the beneficial effects of RBMF have lasted for several days after termination of the treatment. The effects of melatonin on the mastitis are probably attributed to the antioxidant and anti-inflammatory activities of melatonin. Considering the none or low toxicity of melatonin to organisms and the no invasive nature of this approach, we recommend that RBMF could be used in large scale in the dairy farming to target the cow mastitis. creator: Songyang Yao creator: Hao Wu creator: Hui Ma creator: Yao Fu creator: Wenjuan Wei creator: Tiankun Wang creator: Shengyu Guan creator: Hai Yang creator: Xiubo Li creator: Jiangpeng Guo creator: Yongqiang Lu creator: Lu Zhang creator: Changwang He creator: Yi Chang creator: Guoshi Liu uri: https://doi.org/10.7717/peerj.9147 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yao et al. title: Inventory statistics meet big data: complications for estimating numbers of species link: https://peerj.com/articles/8872 last-modified: 2020-05-13 description: We point out complications inherent in biodiversity inventory metrics when applied to large-scale datasets. The number of units of inventory effort (e.g., days of inventory effort) in which a species is detected saturates, such that crucial numbers of detections of rare species approach zero. Any rare errors can then come to dominate species richness estimates, creating upward biases in estimates of species numbers. We document the problem via simulations of sampling from virtual biotas, illustrate its potential using a large empirical dataset (bird records from Cape May, NJ, USA), and outline the circumstances under which these problems may be expected to emerge. creator: Ali Khalighifar creator: Laura Jiménez creator: Claudia Nuñez-Penichet creator: Benedictus Freeman creator: Kate Ingenloff creator: Daniel Jiménez-García creator: Town Peterson uri: https://doi.org/10.7717/peerj.8872 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Khalighifar et al. title: The mitochondrial genome of Acrobeloides varius (Cephalobomorpha) confirms non-monophyly of Tylenchina (Nematoda) link: https://peerj.com/articles/9108 last-modified: 2020-05-13 description: The infraorder Cephalobomorpha is a diverse and ecologically important nematode group found in almost all terrestrial environments. In a recent nematode classification system based on SSU rDNA, Cephalobomorpha was classified within the suborder Tylenchina with Panagrolaimomorpha, Tylenchomorpha and Drilonematomorpha. However, phylogenetic relationships among species within Tylenchina are not always consistent, and the phylogenetic position of Cephalobomorpha is still uncertain. In this study, in order to examine phylogenetic relationships of Cephalobomorpha with other nematode groups, we determined the complete mitochondrial genome sequence of Acrobeloides varius, the first sequenced representative of Cephalobomorpha, and used this sequence for phylogenetic analyses along with 101 other nematode species. Phylogenetic analyses using amino acid and nucleotide sequence data of 12 protein-coding genes strongly support a sister relationship between the two cephalobomorpha species A. varius and Acrobeles complexus (represented by a partial mt genome sequence). In this mitochondrial genome phylogeny, Cephalobomorpha was sister to all chromadorean species (excluding Plectus acuminatus of Plectida) and separated from Panagrolaimomorpha and Tylenchomorpha, rendering Tylenchina non-monophyletic. Mitochondrial gene order among Tylenchina species is not conserved, and gene clusters shared between A. varius and A. complexus are very limited. Results from phylogenetic analysis and gene order comparison confirms Tylenchina is not monophyletic. To better understand phylogenetic relationships among Tylenchina members, additional mitochondrial genome information is needed from underrepresented taxa representing Panagrolaimomorpha and Cephalobomorpha. creator: Taeho Kim creator: Yucheol Lee creator: Hyun-Jong Kil creator: Joong-Ki Park uri: https://doi.org/10.7717/peerj.9108 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kim et al. title: Nonalcoholic fatty liver disease, a potential risk factor of non-specific ST-T segment changes: data from a cross-sectional study link: https://peerj.com/articles/9090 last-modified: 2020-05-13 description: BackgroundNon-specific ST-T segment changes are prevalent and are proven risk factors for early onset of cardiovascular diseases. They can increase all-cause mortality by 100∼200% and are candidate for early signs of cardiovascular changes. Nonalcoholic fatty liver disease (NAFLD) is prevalent worldwide and is one facet of a multisystem disease that confers substantial increases morbidity and mortality of nonalcoholic fatty liver-related cardiovascular diseases. It is unclear whether NAFLD is associated with non-specific ST-T changes warning early signs of cardiovascular changes. Therefore, we investigated this association.MethodsA cross-sectional study was designed that included a sample consisting of 32,922 participants who underwent health examinations. Participants with missing information, excessive alcohol intake, viral hepatitis, chronic liver disease or established cardiovascular diseases were excluded. Electrocardiograms were used for analysis of non-specific ST-T segment changes. NAFLD was diagnosed by ultrasonographic detection of hepatic steatosis without other liver diseases. A multivariable logistic regression model was served to calculate the OR and 95% CI for non-specific ST-T segment changes.ResultsThe prevalence of non-specific ST-T segment changes was 6.5% in participants with NAFLD, however, the prevalence of NAFLD was 42.9% in participants with non-specific ST-T segment changes. NAFLD was independently associated with non-specific ST-T segment changes (OR: 1.925, 95% CI: 1.727-2.143, P < 0.001). After adjusting for age, sex, heart rate, hypertension, body mass index, fasting glucose, total cholesterol, triglycerides, HDL-C, NAFLD remained an independent risk factor of non-specific ST-T segment changes (OR: 1.289, 95% CI: 1.122-1.480).ConclusionNon-specific ST-T segment changes were independently associated with the presence of NAFLD after adjusting for potential confounders. creator: Li Xiao creator: Tao Bai creator: Junchao Zeng creator: Rui Yang creator: Ling Yang uri: https://doi.org/10.7717/peerj.9090 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Xiao et al. title: Application of 16S rRNA gene sequencing in Helicobacter pylori detection link: https://peerj.com/articles/9099 last-modified: 2020-05-13 description: Helicobacter pylori is one of the major stomach microbiome components, promoting development of inflammation and gastric cancer in humans. H. pylori has a unique ability to transform into a coccoidal form which is difficult to detect by many diagnostic methods, such as urease activity detection, and even histopathological examination. Here we present a comparison of three methods for H. pylori identification: histological assessment (with eosin, hematoxylin, and Giemsa staining), polymerase chain reaction (PCR) detection of urease (ureA specific primers), and detection by 16S rRNA gene sequencing. The study employed biopsies from the antral part of the stomach (N = 40). All samples were assessed histologically which revealed H. pylori in eight patients. Bacterial DNA isolated from the bioptates was used as a template for PCR reaction and 16S rRNA gene sequencing that revealed H. pylori in 13 and in 20 patients, respectively. Thus, 16S rRNA gene sequencing was the most sensitive method for detection of H. pylori in stomach biopsy samples. creator: Aleksander Szymczak creator: Stanisław Ferenc creator: Joanna Majewska creator: Paulina Miernikiewicz creator: Jan Gnus creator: Wojciech Witkiewicz creator: Krystyna Dąbrowska uri: https://doi.org/10.7717/peerj.9099 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Szymczak et al. title: The biomarkers of key miRNAs and target genes associated with acute myocardial infarction link: https://peerj.com/articles/9129 last-modified: 2020-05-13 description: BackgroundAcute myocardial infarction (AMI) is considered one of the most prominent causes of death from cardiovascular disease worldwide. Knowledge of the molecular mechanisms underlying AMI remains limited. Accurate biomarkers are needed to predict the risk of AMI and would be beneficial for managing the incidence rate. The gold standard for the diagnosis of AMI, the cardiac troponin T (cTnT) assay, requires serial testing, and the timing of measurement with respect to symptoms affects the results. As attractive candidate diagnostic biomarkers in AMI, circulating microRNAs (miRNAs) are easily detectable, generally stable and tissue specific.MethodsThe Gene Expression Omnibus (GEO) database was used to compare miRNA expression between AMI and control samples, and the interactions between miRNAs and mRNAs were analysed for expression and function. Furthermore, a protein-protein interaction (PPI) network was constructed. The miRNAs identified in the bioinformatic analysis were verified by RT-qPCR in an H9C2 cell line. The miRNAs in plasma samples from patients with AMI (n = 11) and healthy controls (n = 11) were used to construct receiver operating characteristic (ROC) curves to evaluate the clinical prognostic value of the identified miRNAs.ResultsWe identified eight novel miRNAs as potential candidate diagnostic biomarkers for patients with AMI. In addition, the predicted target genes provide insight into the molecular mechanisms underlying AMI. creator: Qi Wang creator: Bingyan Liu creator: Yuanyong Wang creator: Baochen Bai creator: Tao Yu creator: Xian–ming Chu uri: https://doi.org/10.7717/peerj.9129 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: Dental fear association between mothers and adolescents—a longitudinal study link: https://peerj.com/articles/9154 last-modified: 2020-05-13 description: AimTo assess the longitudinal association between adolescents’ and their mothers’ dental fear.Study DesignA longitudinal questionnaire survey study.MethodsA randomized sample of 12-year-old adolescents were selected from local Hong Kong schools. Adolescents and their mothers self-completed the Modified Dental Anxiety Scale (MDAS). The sociodemographic background of the mothers and the oral health habits of the adolescents were also collected and these measurements were repeated at 15- and 18-years-old. Non-parametric tests (Mann–Whitney U test/Kruskall Wallis test) were used to test associations between MDAS dental fear items and independent variables. Logistic regression (adjusted for family’s sociodemographic background and adolescent’s oral health habits) was performed to evaluate the longitudinal association between adolescents’ and mothers’ dental fear.ResultsA total of 212 mother-child pairs were recruited at baseline (12-year-old adolescents). In the first and second follow-ups (15- and 18-years-old), 195 and 182 mother-child pairs completed the survey. Significant associations between mother’s and child’s scores in “feeling about having their teeth scraped and polished”, “having teeth drilled”, and ‘having an injection in the gum’ were found when adolescents were 12- years-old (P < 0.01) and 18-years-old (P < 0.05), but not at 15-years-old.ConclusionAdolescents’ and mothers’ dental fear is associated at 12-years-old and 18-years-old, but not at 15-years-old, which is likely specific to the Hong Kong context but may be extrapolated to other industrialized countries with caution. creator: Hai Ming Wong creator: Yu Yuan Zhang creator: Antonio Perfecto creator: Colman P.J. McGrath uri: https://doi.org/10.7717/peerj.9154 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wong et al. title: Thecodont tooth attachment and replacement in bolosaurid parareptiles link: https://peerj.com/articles/9168 last-modified: 2020-05-13 description: Permian bolosaurid parareptiles are well-known for having complex tooth crowns and complete tooth rows in the jaws, in contrast to the comparatively simple teeth and frequent replacement gaps in all other Paleozoic amniotes. Analysis of the specialized dentition of the bolosaurid parareptiles Bolosaurus from North America and Belebey from Russia, utilizing a combination of histological and tomographic data, reveals unusual patterns of tooth development and replacement. The data confirm that bolosaurid teeth have thecodont implantation with deep roots, the oldest known such example among amniotes, and independently evolved among much younger archosauromorphs (including dinosaurs and crocodilians) and among synapsids (including mammals). High-resolution CT scans were able to detect the density boundary between the alveolar bone and the jawbone, as confirmed by histology, and revealed the location and size of developing replacement teeth in the pulp cavity of functional teeth. Evidence provided by the paratype dentary of Belebey chengi indicates that replacement teeth are present along the whole tooth row at slightly different stages of development, with the ontogenetically more developed teeth anteriorly, suggesting that tooth replacement was highly synchronized. CT data also show tooth replacement is directly related to the presence of lingual pits in the jaw, and that migration of tooth buds occurs initially close to these resorption pits to a position immediately below the functional tooth within its pulp cavity. The size and complex shape of the replacement teeth in the holotype of Bolosaurus grandis indicate that the replacement teeth can develop within the pulp cavity to an advanced stage while the previous generation remains functional for an extended time, reminiscent of the condition seen in other amniotes with occluding dentitions, including mammals. creator: Adam J. Snyder creator: Aaron R.H. LeBlanc creator: Chen Jun creator: Joseph J. Bevitt creator: Robert R. Reisz uri: https://doi.org/10.7717/peerj.9168 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Snyder et al. title: Changes in the gut microbiota during Asian particolored bat (Vespertilio sinensis) development link: https://peerj.com/articles/9003 last-modified: 2020-05-12 description: BackgroundThe gut microbiota is closely linked to host development, diet and health and is influenced by both the host and the environment. Although many studies have focused on the dynamics of the gut microbiota during development in captive animals, few studies have focused on the dynamics of the gut microbiota during development in wild animals, especially for the order Chiroptera.MethodsIn this study, we characterized the gut microbiota of the wild Asian particolored bat (Vespertilio sinensis) from 1 day to 6 weeks after birth. We explored the changes in their gut microbial community compositions, examined possible influencing factors, and predicted the feeding transition period.ResultsThe gut microbiota changed during the development of V. sinensis. The alpha diversity of the bats’ gut microbiota gradually increased but did not change significantly from the 1st day to the 4th week after birth; however, the alpha diversity decreased significantly in week 5, then stabilized. The beta diversity differed slightly in weeks 4–6. In week 4, the fecal samples showed the highest diversity in bacterial community composition. Thus, we predicted that the potential feeding transition period for V. sinensis may occur during week 4. Redundancy analysis showed that age and body mass index significantly affected the compositional changes of the gut microbiota in Asian particolored bats.ConclusionThe gut microbiota changed during the development of V. sinensis. We suggest that changes in the alpha and beta diversity during week 4 after birth indicate a potential feeding transition, highlighting the importance of diet in the gut microbiota during the development of V. sinensis. creator: Zhongwei Yin creator: Keping Sun creator: Aoqiang Li creator: Deyi Sun creator: Zhongle Li creator: Guohong Xiao creator: Jiang Feng uri: https://doi.org/10.7717/peerj.9003 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Yin et al. title: Extent, patterns and demographic correlates for physical fighting among school-attending adolescents in Namibia: examination of the 2013 Global School-based Health Survey link: https://peerj.com/articles/9075 last-modified: 2020-05-12 description: BackgroundPhysical fighting is particularly detrimental for young people, often affecting other areas of their developing lives, such as relationships with friends and family and participating in risky behaviors. We aim to quantify the amount of problematic physical fighting in Namibian adolescents and identify modifiable risk factors for intervention.MethodsWe used the Namibia 2013 Global School-based Student Health Survey (GSHS). This survey collects health-related information on school-attending adolescents in grades 7 to 12. We defined physical fighting as having participated in at least two physical fights in the 12 months prior to responding to the survey. Factors that may be associated with physical fighting were identified a prior based on the literature and included age, sex, anxiety, suicide planning, truancy, physical activity, bullying victimization, presence of supportive parental figures, presence of helpful peers, extent of social network, and food insecurity. Multivariable logistic regression models were created to identify factors associated with physical fighting.ResultsA total of 4,510 adolescents were included in the study. A total of 52.7% female. 16.9% of adolescents reported engaging in at least two physical fights in the previous year. Factors associated with an increased odds of physical fighting included having a suicide plan, anxiety, truancy, food deprivation and being bullied. Increased age and loneliness were associated with a decreased odds of physical fighting.ConclusionThis study identifies problematic physical fighting among adolescents in Namibia. We recommend public health and school-based programming that simultaneously targets risk behaviours and conflict resolution to reduce rates of physical fighting. creator: Laura E. Davis creator: Anne Abio creator: Michael Lowery Wilson creator: Masood Ali Shaikh uri: https://doi.org/10.7717/peerj.9075 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Davis et al. title: EEG based assessment of stress in horses: a pilot study link: https://peerj.com/articles/8629 last-modified: 2020-05-12 description: As has been hypothesized more than 20 years ago, data derived from Electroencephalography (EEG) measurements can be used to distinguish between behavioral states associated with animal welfare. In our current study we found a high degree of correlation between the modulation index of phase related amplitude changes in the EEG of horses (n = 6 measurements with three different horses, mare and gelding) and their facial expression, measured by the use of the horse grimace scale. Furthermore, the pattern of phase amplitude coupling (PAC) was significantly different between a rest condition and a stress condition in horses. This pilot study paves the way for a possible use of EEG derived PAC as an objective tool for the assessment of animal welfare. Beyond that, the method might be useful to assess welfare aspects in the clinical setting for human patients, as for example in the neonatal intensive care unit. creator: Nora V. de Camp creator: Mechthild Ladwig-Wiegard creator: Carola I.E. Geitner creator: Jürgen Bergeler creator: Christa Thöne-Reineke uri: https://doi.org/10.7717/peerj.8629 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 de Camp et al. title: Phylogenetic study of extirpated Korean leopard using mitochondrial DNA from an old skin specimen in South Korea link: https://peerj.com/articles/8900 last-modified: 2020-05-12 description: The leopard, Panthera pardus, is a threatened species in its range throughout the world. Although, historically, the Korean Peninsula had a high population density of leopards, they were extirpated from South Korea by 1970, leaving almost no genetic specimens. Traditionally, Korean leopards are classified as Panthera pardus orientalis; however, their classification is based only on locality and morphology. Therefore, there is a need for genetic studies to identify the phylogenetic status of Korean leopards at the subspecies level. Presently, no extant wild specimen is available from South Korea; therefore, we extracted genetic material from the old skin of a leopard captured in Jirisan, South Korea in the 1930s and conducted the first phylogenetic study of the South Korean leopard. A total of 726 bp of mitochondrial DNA, including segments of the NADH5 and control region, were amplified by PCR. A phylogenetic analysis of the fragment, along with sequences of nine leopard subspecies from GenBank revealed that the extinct South Korean leopard belonged to the Asian leopard group and in the same clade as the Amur leopard (Panthera pardus orientalis). Thus, the leopard that inhabited South Korea in the past was of the same subspecies as the Amur leopard population currently inhabiting the transboundary region of Russia, China, and North Korea. These results emphasize the importance of conserving the endangered wild Amur leopard population (estimated to be about 60–80 individuals) in Russia and China, for future restoration of leopards in the Korean Peninsula. creator: Jee Yun Hyun creator: Jang Hyuk Cho creator: Puneet Pandey creator: Mi-Sook Min creator: Kyung Seok Kim creator: Hang Lee uri: https://doi.org/10.7717/peerj.8900 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Hyun et al. title: Exploring environmental coverages of species: a new variable contribution estimation methodology for rulesets from the genetic algorithm for rule-set prediction link: https://peerj.com/articles/8968 last-modified: 2020-05-12 description: Variable contribution estimation for, and determination of variable importance within, ecological niche models (ENMs) remain an important area of research with continuing challenges. Most ENM algorithms provide normally exhaustive searches through variable space; however, selecting variables to include in models is a first challenge. The estimation of the explanatory power of variables and the selection of the most appropriate variable set within models can be a second challenge. Although some ENMs incorporate the variable selection rubric inside the algorithms, there is no integrated rubric to evaluate the variable importance in the Genetic Algorithm for Ruleset Production (GARP). Here, we designed a novel variable selection methodology based on the rulesets generated from a GARP experiment. The importance of the variables in a GARP experiment can be estimated based on the consideration of the prevalence of each environmental variable in the dominant presence rules of the best subset of models and its coverage. We tested the performance of this variable selection method based on simulated species with both weak and strong responses to simulated environmental covariates. The variable selection method generally performed well during the simulations with over 2/3 of the trials correctly identifying most covariates. We then predict the distribution of Toxostoma rufum (a bird with a cosmopolitan distribution) in the continental United States (US) and apply our variable selection procedure as a real-world example. We found that the distribution of T. rufum could be accurately modeled with 13 or 10 of 21 variables, using an UI cutoff of 0.5 or 0.25, respectively, arriving at parsimonious environmental coverages with good model accuracy. We also provide tools to simulate species distributions for testing ENM approaches using R. creator: Anni Yang creator: Juan Pablo Gomez creator: Jason K. Blackburn uri: https://doi.org/10.7717/peerj.8968 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Yang et al. title: EZH2/H3K27Me3 and phosphorylated EZH2 predict chemotherapy response and prognosis in ovarian cancer link: https://peerj.com/articles/9052 last-modified: 2020-05-12 description: BackgroundEZH2 acts as an oncogene through canonical pathway EZH2/H3K27Me3 and uncanonical pathway pAkt1/pS21EZH2 in many solid tumors including ovarian cancer. However, the clinical value of EZH2/H3K27Me3 and pAkt1/pS21EZH2 remain unclear. In the current study, we aim to investigate the correlation between these two pathways to clinical-pathological parameters and prognosis.MethodsEZH2, H3K27Me3, pAkt1 and pS21EZH2 expression were evaluated by tissue micro-array and immunohistochemistry in a cohort of ovarian cancer patients. The results were analyzed based on clinical characteristics and survival outcomes.ResultsEZH2, H3K27Me3, pAkt1 and pS21EZH2 were universally expressed in ovarian cancer specimens with a positive expression rate of 81.54% (53/65), 88.89% (48/54), 63.07% (41/65) and 75.38% (49/65). EZH2-pS21EZH2 (Spearman r = 0.580, P < 0.0001) and pS21EZH2-pAkt1 (Spearman r = 0.546, P < 0.0001) were closely correlated while EZH2- H3K27Me3 were less closely correlated (Spearman r = 0.307, P = 0.002). Low pS21EZH2 associated with better chemotherapy response (OR = 0.184; 95% CI [0.052–0.647], P = 0.008) according to logistic regression with an area under the curve of 0.789 (specificity 89.36%, sensitivity 68.42%) by ROC analysis and predicted improved progression-free survival (HR = 0.453; 95% CI [0.229–0.895], P = 0.023) as indicated by multivariate cox regression. A combination of EZH2low/H3K27Me3low status predicted better chemotherapy response (OR = 0.110; 95% CI [0.013–0.906], P = 0.040) and better progression-free survival (HR = 0.388; 95% CI [0.164–0.917], P = 0.031). The results suggested that EZH2/H3K27Me3 and pEZH2 predicted chemotherapy response and progression-free survival in ovarian cancer. creator: Si Sun creator: Qiang Yang creator: E Cai creator: Bangxing Huang creator: Feiquan Ying creator: Yiping Wen creator: Jing Cai creator: Ping Yang uri: https://doi.org/10.7717/peerj.9052 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Sun et al. title: Direct aortic route versus transaxillary route for transcatheter aortic valve replacement: a systematic review and meta-analysis link: https://peerj.com/articles/9102 last-modified: 2020-05-12 description: BackgroundThe transfemoral route is contraindicated in nearly 10% of transcatheter aortic valve replacement (TAVR) candidates because of unsuitable iliofemoral vessels. Transaxillary (TAx) and direct aortic (DAo) routes are the principal nonfemoral TAVR routes; however, few studies have compared their outcomes.MethodsWe performed a systematic review and meta-analysis to compare the rates of mortality, stroke, and other adverse events of TAx and DAo TAVR. The study was prospectively registered with PROSPERO (registration number: CRD42017069788). We searched Medline, PubMed, Embase, and Cochrane databases for studies reporting the outcomes of DAo or TAx TAVR in at least 10 patients. Studies that did not use the Valve Academic Research Consortium definitions were excluded. We included studies that did not directly compare the two approaches and then pooled rates of events from the included studies for comparison.ResultsIn total, 31 studies were included in the quantitative meta-analysis, with 2,883 and 2,172 patients in the DAo and TAx TAVR groups, respectively. Compared with TAx TAVR, DAo TAVR had a lower Society of Thoracic Surgery (STS) score, shorter fluoroscopic time, and less contrast volume use. The 30-day mortality rates were significantly higher in the DAo TAVR group (9.6%, 95% confidence interval (CI) = [8.4–10.9]) than in the TAx TAVR group (5.7%, 95% CI = [4.8–6.8]; P for heterogeneity <0.001). DAo TAVR was associated with a significantly lower risk of stroke in the overall study population (2.6% vs. 5.8%, P for heterogeneity <0.001) and in the subgroup of studies with a mean STS score of ≥8 (1.6% vs. 6.2%, P for heterogeneity = 0.005). DAo TAVR was also associated with lower risks of permanent pacemaker implantation (12.3% vs. 20.1%, P for heterogeneity = 0.009) and valve malposition (2.0% vs. 10.2%, P for heterogeneity = 0.023) than was TAx TAVR.ConclusionsDAo TAVR increased 30-day mortality rate compared with TAx TAVR; by contrast, TAx TAVR increased postoperative stroke, permanent pacemaker implantation, and valve malposition risks compared with DAo TAVR. creator: Hsiu-An Lee creator: I-Li Su creator: Shao-Wei Chen creator: Victor Chien-Chia Wu creator: Dong-Yi Chen creator: Pao-Hsien Chu creator: An-Hsun Chou creator: Yu-Ting Cheng creator: Pyng-Jing Lin creator: Feng-Chun Tsai uri: https://doi.org/10.7717/peerj.9102 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Lee et al. title: Early language competence, but not general cognitive ability, predicts children’s recognition of emotion from facial and vocal cues link: https://peerj.com/articles/9118 last-modified: 2020-05-12 description: The ability to accurately identify and label emotions in the self and others is crucial for successful social interactions and good mental health. In the current study we tested the longitudinal relationship between early language skills and recognition of facial and vocal emotion cues in a representative UK population cohort with diverse language and cognitive skills (N = 369), including a large sample of children that met criteria for Developmental Language Disorder (DLD, N = 97). Language skills, but not non-verbal cognitive ability, at age 5–6 predicted emotion recognition at age 10–12. Children that met the criteria for DLD showed a large deficit in recognition of facial and vocal emotion cues. The results highlight the importance of language in supporting identification of emotions from non-verbal cues. Impairments in emotion identification may be one mechanism by which language disorder in early childhood predisposes children to later adverse social and mental health outcomes. creator: Sarah Griffiths creator: Shaun Kok Yew Goh creator: Courtenay Fraiser Norbury uri: https://doi.org/10.7717/peerj.9118 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Griffiths et al. title: Analyzing bacterial community in pit mud of Yibin Baijiu in China using high throughput sequencing link: https://peerj.com/articles/9122 last-modified: 2020-05-12 description: “Yibin Baijiu” (YB) is a special Chinese strong-aroma Baijiu (CSAB) that originated in Yibin, a city in western China. YB is fermented in cellars lined with pit mud (PM), the microbiota in which may affect YB quality. In this study, high throughput sequencing of the 16S rRNA gene was used to demonstrate the bacterial community structure and diversity in PM of YB. In addition, the physicochemical characteristics of PM were also analyzed, including moisture content, pH, and available phosphorous, ammonia nitrogen, and humic acid levels. Results showed that Firmicutes was the dominant phylum in all PM samples with abundance > 70.0%, followed by Euryarchaeota (11.3%), Bacteroidetes (6.5%), Synergistetes (3.0%), Actinobacteria (1.4%), and Proteobacteria (1.2%). Furthermore, 14 different genera with average relative abundance of > 1% were detected. The Chao1 and Shannon indexes did not vary significantly between the sub-layer and middle-layer PM (P > 0.05). However, Linear discriminant analysis Effect Size (LEfSe) analysis showed that the relative abundance of Lactobacillus in the sub-layer PM was significantly higher than in middle-layer PM. pH differed significantly (P < 0.05) between the two groups. Canonical correspondence analysis revealed that bacterial community in PM correlated significantly with available phosphorous content and pH. Our study provides basic data for further elucidating the diversity of microbiota in the PM of YB and the potential mechanism of Baijiu production. creator: Li Chen creator: Yuzhu Li creator: Lei Jin creator: Li He creator: Xiaolin Ao creator: Shuliang Liu creator: Yong Yang creator: Aiping Liu creator: Shujuan Chen creator: Likou Zou uri: https://doi.org/10.7717/peerj.9122 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: Community science participants gain environmental awareness and contribute high quality data but improvements are needed: insights from Bumble Bee Watch link: https://peerj.com/articles/9141 last-modified: 2020-05-12 description: Bumble Bee Watch is a community science program where participants submit photos of bumble bees from across Canada and the United States for expert verification. The data can be used to help better understand bumble bee biology and aid in their conservation. Yet for community science programs like this to be successful and sustainable, it is important to understand the participant demographics, what motivates them, and the outcomes of their participation, as well as areas that are working well or could be improved. It is also important to understand who verifies the submissions, who uses the data and their views on the program. Of the surveyed users, most participate to contribute to scientific data collection (88%), because of a worry about bees and a desire to help save them (80%), to learn more about species in their property (63%) or region (56%), and because of a personal interest (59%). About 77% report increased awareness of species diversity, while 84% report improvement in their identification skills. We found that 81% had at least one college or university degree. There were more respondents from suburban and rural areas than urban areas, but area did not affect numbers of submissions. While half were between 45 and 64 years of age, age did not influence motivation or number of submissions. Respondents were happy with the program, particularly the website resources, the contribution to knowledge and conservation efforts, the educational values, and the ability to get identifications. Areas for improvement included app and website functionality, faster and more detailed feedback, localized resources, and more communication. Most respondents participate rarely and have submitted fewer than ten records, although about five percent are super users who participate often and submit more than fifty records. Suggested improvements to the program may increase this participation rate. Indeed, increased recruitment and retention of users in general is important, and advertising should promote the outcomes of participation. Fifteen experts responded to a separate survey and were favorable of the program although there were suggestions on how to improve the verification process and the quality of the submitted data. Suggested research questions that could be asked or answered from the data included filling knowledge gaps (species diversity, ranges, habitat, phenology, floral associations, etc.), supporting species status assessments, effecting policy and legislation, encouraging habitat restoration and management efforts, and guiding further research. However, only about half have used data from the project to date. Further promotion of Bumble Bee Watch and community science programs in general should occur amongst academia, conservationists, policy makers, and the general public. This would help to increase the number and scope of submissions, knowledge of these species, interest in conserving them, and the overall program impact. creator: Victoria J. MacPhail creator: Shelby D. Gibson creator: Sheila R. Colla uri: https://doi.org/10.7717/peerj.9141 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 MacPhail et al. title: Long-term outcomes of repaired and unrepaired truncus arteriosus: 20-year, single-center experience in Thailand link: https://peerj.com/articles/9148 last-modified: 2020-05-12 description: BackgroundTruncus arteriosus (TA) is a complex congenital heart disease that carries morbidities in the first year of life. Previous authors have reported an operative mortality of 50%. In this report, we aim to report on the survival of patients with TA in our medical center in the recent era.MethodsA retrospective review of all patients diagnosed with TA in Siriraj Hospital, Thailand from August 1995 to March 2018 was performed. Patients with single ventricle, hemiTA were excluded. The characteristics and outcomes of repaired and unrepaired TA patients with a known recent functional status in 2018 were reviewed. Operative mortality risks were analyzed using a multivariate model.ResultsA total of 74 patients (median age at referral: 70 days) were included in the cohort. One-third of the patients had associated anomalies including DiGeorge syndrome (13.5%). Anatomical repair was not performed in 22 patients (29.7%). The median age at time of repair for the 52 patients was 133 days (range: 22 days to 16.7 years). Complex TA was 10%. Early mortality occurred in 16 patients (30.8%). Five patients (9.6%) had late deaths at 0.3–1.2 years. Significant mortality risk was weight at time of operation <4 kg (HR 3.05, 95% CI [1.05–8.74], p-value 0.041). Of the 31 operation survivors, 17 required re-intervention within 0.4–11.4 years. Eight patients had reoperation at 8.7 years (range: 2.7–14.6 years) post-repair. Freedom from reoperation was 93%, 70.4%, and 31%, at 5, 10, and 15 years, respectively. All late survivors were in functional class I–II. Of the 22 unrepaired TA patients, 11 patients (50%) died (median age: 13.6 years; range: 14 days–32.8 years). Survival of unrepaired TA patients was 68.2%, 68.2%, and 56.8, at 5, 10, and 15 years of age, respectively. At the end of study, 11 survivors of TA with palliative treatment had a recent mean oxygen saturation value of 84.1 ± 4.8% and a mean weight for height of 81.4 ± 12.7%, which were significantly lower than those of 31 late-survivors who had undergone anatomical repair.ConclusionContemporary survival rates of patients with TA following operation in the center has been gradually improved over time. Most of the operative mortality occurs in the early postoperative period. Compared to patients with TA who had palliative treatment, operative survivors have a better functional status even though they carry a risk for re-intervention. creator: Ekkachai Dangrungroj creator: Chodchanok Vijarnsorn creator: Prakul Chanthong creator: Paweena Chungsomprasong creator: Supaluck Kanjanauthai creator: Kritvikrom Durongpisitkul creator: Jarupim Soongswang creator: Kriangkrai Tantiwongkosri creator: Thaworn Subtaweesin creator: Somchai Sriyoschati uri: https://doi.org/10.7717/peerj.9148 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Dangrungroj et al. title: High type I collagen density fails to increase breast cancer stem cell phenotype link: https://peerj.com/articles/9153 last-modified: 2020-05-12 description: Breast cancer is a highly frequent and lethal malignancy which metastasis and relapse frequently associates with the existence of breast cancer stem cells (CSCs). CSCs are undifferentiated, aggressive and highly resistant to therapy, with traits modulated by microenvironmental cells and the extracellular matrix (ECM), a biologically complex and dynamic structure composed mainly by type I collagen (Col-I). Col-I enrichment in the tumor-associated ECM leads to microenvironment stiffness and higher tumor aggressiveness and metastatic potential. While Col-I is also known to induce tumor stemness, it is unknown if such effect is dependent of Col-I density. To answer this question, we evaluated the stemness phenotype of MDA-MB-231 and MCF-7 human breast cancer cells cultured within gels of varying Col-I densities. High Col-I density increased CD44+CD24− breast cancer stem cell (BCSC) immunophenotype but failed to potentiate Col-I fiber alignment, cell self-renewal and clonogenicity in MDA-MB-231 cells. In MCF-7 cells, high Col-I density decreased total levels of variant CD44 (CD44v). Common to both cell types, high Col-I density induced neither markers related to CSC nor those related with mechanically-induced cell response. We conclude that high Col-I density per se is not sufficient to fully develop the BCSC phenotype. creator: Iuri C. Valadão creator: Ana Carolina L. Ralph creator: François Bordeleau creator: Luciana M. Dzik creator: Karen S.C. Borbely creator: Murilo V. Geraldo creator: Cynthia A. Reinhart-King creator: Vanessa M. Freitas uri: https://doi.org/10.7717/peerj.9153 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Valadão et al. title: Exploring the interactions among SNARC effect, finger counting direction and embodied cognition link: https://peerj.com/articles/9155 last-modified: 2020-05-12 description: The Spatial Numerical Association of Response Code (SNARC) is the preferential association between smaller/larger magnitudes and left/right side, respectively. Some evidence suggest a link between SNARC and a left-to-right finger counting habit. We asked 268 participants to show how they use the hands to count from 1 to 10. By means of this ecological task, 80% of the sample use first the right hand (to count from 1 to 5) and the majority of them use a palm-up posture. In Experiment 2 (N = 46) right-starters were asked to categorize 1-to-5 magnitudes as even or odd, using the left and right hand. Stimuli were presented both as Arabic numbers and by means of left and right hand photographs in palm-up and palm-down posture. Results confirmed the expected SNARC effect in the Arabic condition. With hand images we found that right hand responses were better for larger than for smaller magnitudes (SNARC, mainly for left hand palm-up stimuli), showing that the SNARC can be generalized to different codes. Finally, the interactions between magnitudes and left/right hand images in palm-up and palm-down posture suggest that embodied cognition can influence numerical processing. creator: Giulia Prete creator: Luca Tommasi uri: https://doi.org/10.7717/peerj.9155 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Prete and Tommasi title: Ontogeny of a sexually selected structure in an extant archosaur Gavialis gangeticus (Pseudosuchia: Crocodylia) with implications for sexual dimorphism in dinosaurs link: https://peerj.com/articles/9134 last-modified: 2020-05-12 description: Despite strong evidence for sexual selection in various display traits and other exaggerated structures in large extinct reptiles, such as dinosaurs, detecting sexual dimorphism in them remains difficult. Their relatively small sample sizes, long growth periods, and difficulties distinguishing the sexes of fossil specimens mean that there are little compelling data on dimorphism in these animals. The extant gharial (Gavialis gangeticus) is a large and endangered crocodylian that is sexually dimorphic in size, but males also possesses a sexually selected structure, the ghara, which has an osteological correlate in the presence of a fossa associated with the nares. This makes the species a unique model for potentially assessing dimorphism in fossil lineages, such as dinosaurs and pterosaurs, because it is a large, slow-growing, egg-laying archosaur. Here we assess the dimorphism of G. gangeticus across 106 specimens and show that the presence of a narial fossa diagnoses adult male gharials. Males are larger than females, but the level of size dimorphism, and that of other cranial features, is low and difficult to detect without a priori knowledge of the sexes, even with this large dataset. By extension, dimorphism in extinct reptiles is very difficult to detect in the absence of sex specific characters, such as the narial fossa. creator: David Hone creator: Jordan C. Mallon creator: Patrick Hennessey creator: Lawrence M. Witmer uri: https://doi.org/10.7717/peerj.9134 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Hone et al. title: Luteolin attenuates doxorubicin-induced cardiotoxicity by modulating the PHLPP1/AKT/Bcl-2 signalling pathway link: https://peerj.com/articles/8845 last-modified: 2020-05-11 description: BackgroundLuteolin (LUT) is a flavonoid found in vegetables and fruits that has diverse functions. Doxorubicin (DOX) is an anthracycline antibiotic that is frequently used for the treatment of various cancers. Unfortunately, the clinical efficacy of DOX is limited by its dose-related cardiotoxicity. In this study, we aimed to investigate the potential mechanism through which LUT attenuates cardiotoxicity in vivo.MethodsWe evaluated the body weight, heart weight, electrocardiogram, and pathological changes before and after administration of LUT. Moreover, the effects of LUT (50 mg/kg in the low dose group, 100 mg/kg in the high dose group) on biochemical parameters (brain natriuretic peptide, creatine kinase MB, cardiac troponin T, and dehydrogenation of lactate enzyme) and oxidative stress parameters (malondialdehyde and superoxide dismutase) were studied in the sera of cardiotoxicity model rats. We also identified the apoptotic mediators whose expression was induced by LUT by quantitative real-time reverse transcription-polymerase chain reaction (RT-qPCR) evaluation. In addition, we used network analysis to predict DOX-induced cardiotoxicity and protection afforded by LUT. Western blotting was used to detect the expression of associated proteins.ResultsLUT significantly improved DOX-induced cardiotoxicity in a dose-dependent fashion. LUT ameliorated DOX-induced weight loss and heart weight changes, as well as changes in biochemical parameters and oxidative stress parameters in heart injury model rats. LUT’s protective effect was observed via regulation of the apoptotic markers Bcl-2, Bax, and caspase-3 mRNA and protein expression levels. Network analysis showed that the AKT/Bcl-2 signalling pathway was activated; specifically, the PH domain leucine-rich repeats protein phosphatase 1 (phlpp1) was involved in the AKT/Bcl-2 signal pathway. LUT inhibited the activity of phlpp1 leading to positive regulation of the AKT/Bcl-2 pathway, which attenuated doxorubicin-induced cardiotoxicity.ConclusionsThese results demonstrate that LUT exerted protective effects against DOX-induced cardiotoxicity in vivo by alleviating oxidative stress, suppressing phlpp1 activity, and activating the AKT/Bcl-2 signalling pathway. creator: YanDong Zhang creator: ChengYuan Ma creator: ChunShui Liu creator: Feng Wei uri: https://doi.org/10.7717/peerj.8845 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: The relationship between adolescent obesity and pelvis dimensions in adulthood: a retrospective longitudinal study link: https://peerj.com/articles/8951 last-modified: 2020-05-11 description: BackgroundThe effect of fat tissue on a developing individual is fundamentally different from the effect on an adult. Several changes caused by obesity during sexual maturation have an irreversible and severe negative effect (lower fertility, reduced final height, type 2 diabetes mellitus) even for those who have subsequently lost weight. Our study was focused on monitoring the skeletal structure substantially influenced by sex hormones—the pelvis. The adult pelvis is strongly sexually dimorphic, which is not the case for the juvenile pelvis; skeletal differences between sexes are not so prominent and start to manifest with the onset of puberty. Evidence from animal models and case studies of treatment of gender dysphoria suggests that estrogens have a stimulatory effect on the growth plates present on the pelvis, leading to morphological change. Male obesity, especially in puberty, is connected with hypogonadism, manifesting in low levels of testosterone, and high levels of estrogens. The goal of our study was to evaluate the influence of obesity during adolescence on the morphology of the adult pelvis in the context of androgen and estrogen status.Sample and MethodsOur sample consists of 238 individuals (144 females, 94 males) observed after an 8 year follow-up (mean age during enrollment 15.2 years, follow-up 23.3 years). Anthropometry and body composition using bioimpedance analysis (BIA) were obtained. During the follow-up, saliva samples from male participants were also collected to estimate testosterone and estradiol levels using the salivary ELISA kit (Salimetrics LLC, State College, PA, USA).ResultsThe body fat (percentage of body fat estimated using BIA) was strongly positively associated with relative pelvic breadths in adulthood (males r = 0.64; females r = 0.56, both with p < 0.001). Adulthood pelvic breadth was a highly sensitive (0.81) and specific (0.74) retrospective marker of obesity during adolescence. The complex regression model (with reduction of dimensionality) including testosterone, estradiol to testosterone ratio and body fat (adolescent and adulthood) was able to describe 54.8% variability of pelvic breadth among males.DiscussionWe observed that adults with a history of obesity from adolescence tend to have a wider dimension of the bony pelvis in adulthood. Based on the parameters of the adult pelvis, the history of obesity can be determined with satisfactory sensitivity and specificity (<70%). One of the explanations for this observation can be the influence of relatively elevated estrogens levels connected with excessive adiposity leading to a wider pelvis. However, the biomechanical stress connected with elevated body mass also has to be considered, as does the influence of physical activity and gait pattern on the skeletal build. creator: Jan M. Novak creator: Jaroslav Bruzek creator: Hana Zamrazilova creator: Marketa Vankova creator: Martin Hill creator: Petr Sedlak uri: https://doi.org/10.7717/peerj.8951 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Novak et al. title: Discordance of vancomycin minimum inhibitory concentration for methicillin-resistant Staphylococcus aureus at 2 μg/mL between Vitek II, E-test, and Broth Microdilution link: https://peerj.com/articles/8963 last-modified: 2020-05-11 description: BackgroundVancomycin, the first line antibiotic for methicillin-resistant Staphylococcus aureus (MRSA) bacteremia, is often administered inappropriately when MIC is greater than 2 µg/mL, including ‘susceptible’ strains. This study assessed the discordance of vancomycin minimum inhibitory concentration (MIC) for methicillin-resistant Staphylococcus aureus (MRSA).MethodsIn total, 229 MRSA isolates from blood cultures collected between 2009 and 2015 at a tertiary hospital in Taiwan were examined. The MICs of vancomycin were measured using Vitek 2, E-test, and standard broth microdilution at the level of 2 µg/mL.ResultsThe geometric mean of the MICs of hospital-acquired MRSA was higher than that of community-acquired MRSA (P < 0.001), with the exact agreement rates (with broth microdilution) at 2 µg/mL being 53.6% in Vitek 2 and 86.7% in E-test. Overall, E-test (98.1%) had more categorical accordance than did Vitek 2 (94.0%; P = 0.026). Vitek 2 had a tendency to overestimate MRSA in high-MIC isolates, whereas E-test inclined underestimation in low-MIC isolates. Surprisingly, the discordance rates of MRSA vancomycin MICs were higher in hospital-acquired isolates (13.3%–17.0%) than in community-acquired isolates (6.2%–7.0%).ConclusionThe Infectious Diseases Society of America recommends the use of alternative antimicrobial agents when vancomycin MIC is ≥ 2 µg/mL; in this study, only 53.6% of the isolates tested using Vitek 2 showed a high MIC in the broth microdilution method. Accurate identification of the resistance profile is a key component of antimicrobial stewardship programs. Therefore, to reduce inappropriate antibiotic use and mitigate the emergence of resistant strains, we recommend using complementary tests such as E-test or Broth microdilution to verify the MIC before administering second-line antibiotics.Strengths(1) We compared the categorical agreement between different methods measuring MRSA MICs level. (2) Physicians should incorporate this information and consider a complementary test to verify the appropriateness of the decision of shifting vancomycin to second-line antibiotic treatment to improve patients’ prognosis. (3) MRSA-vancomycin MICs at a cutoff of 2 µg/mL obtained using Vitek II exhibited a higher sensitivity level and negative predictive value than those obtained using E-test in the prediction of categorical agreement with standard broth microdilution.Limitation(1) Our research was based on a single hospital-based study. (2) The MRSA strains in this study were stored for more than 12 months after isolation. (3) We did not collect information on clinical prognosis. creator: Chien-Feng Kuo creator: Chon Fu Lio creator: Hsiang-Ting Chen creator: Yu-Ting Tina Wang creator: Kevin Sheng-Kai Ma creator: Yi Ting Chou creator: Fu-Chieh Chang creator: Shin-Yi Tsai uri: https://doi.org/10.7717/peerj.8963 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kuo et al. title: Clustering of multiple health risk behaviors and its association with diabetes in a Southern Chinese adult population: a cross-sectional study link: https://peerj.com/articles/9025 last-modified: 2020-05-11 description: BackgroundIdentifying the clustering patterns of health risk behaviors (HRBs) within individuals and their health impacts are essential to develop lifestyle promotion strategies. This study aimed to explore the clustering of a range of HRBs and the associations between such identified clusters and diabetes in Southern Chinese adults.MethodsData from 5,734 adults aged 35–75 years and underwent health examinations from November 2012 to December 2013 at a tertiary hospital in Guangzhou were analyzed. Behavioral characteristics, including smoking, alcohol use, physical activity, and sleep duration and quality, were measured by questionnaires. Latent class analysis was conducted by gender to identify HRBs clustering patterns, and logistic regression models were used to estimate the associations between behavioral patterns and diabetes.ResultsThree distinct behavioral clusters emerged in both genders. Male classes were defined as: (1) healthy lifestyle (Class 1, 62.9%); (2) cumulate harmful habits (Class 2, 27.1%); (3) poor sleep and risky habits (Class 3, 10.0%). Female classes were: (1) healthy lifestyle (Class 1, 83.0%); (2) inactive, daytime dysfunction (Class 2, 5.7%); (3) poor sleep habits (Class 3, 11.3%). Individuals of Class 2 and Class 3 showed a higher likelihood of diabetes across genders (multivariable-adjusted ORs [95% CIs], 2.03 [1.49–2.76] and 2.61 [1.78–3.81] among males, 2.64 [1.16–5.98] and 1.81 [1.07–3.06] among females) when compared with those of Class 1.ConclusionsOur data provided additional evidence of HRBs clustering among adults, and such clustering was associated with an increased risk of diabetes. These findings have implications for identifying vulnerable subgroups and developing diabetes prevention programs. creator: Guanrong Zhang creator: Caibing Luo creator: Ying Cui creator: Yifan Lu creator: Yang Yang uri: https://doi.org/10.7717/peerj.9025 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: The spindle assembly checkpoint and speciation link: https://peerj.com/articles/9073 last-modified: 2020-05-11 description: A mechanism is proposed by which speciation may occur without the need to postulate geographical isolation of the diverging populations. Closely related species that occupy overlapping or adjacent ecological niches often have an almost identical genome but differ by chromosomal rearrangements that result in reproductive isolation. The mitotic spindle assembly checkpoint normally functions to prevent gametes with non-identical karyotypes from forming viable zygotes. Unless gametes from two individuals happen to undergo the same chromosomal rearrangement at the same place and time, a most improbable situation, there has been no satisfactory explanation of how such rearrangements can propagate. Consideration of the dynamics of the spindle assembly checkpoint suggest that chromosomal fission or fusion events may occur that allow formation of viable heterozygotes between the rearranged and parental karyotypes, albeit with decreased fertility. Evolutionary dynamics calculations suggest that if the resulting heterozygous organisms have a selective advantage in an adjoining or overlapping ecological niche from that of the parental strain, despite the reproductive disadvantage of the population carrying the altered karyotype, it may accumulate sufficiently that homozygotes begin to emerge. At this point the reproductive disadvantage of the rearranged karyotype disappears, and a single population has been replaced by two populations that are partially reproductively isolated. This definition of species as populations that differ from other, closely related, species by karyotypic changes is consistent with the classical definition of a species as a population that is capable of interbreeding to produce fertile progeny. Even modest degrees of reproductive impairment of heterozygotes between two related populations may lead to speciation by this mechanism, and geographical isolation is not necessary for the process. creator: Robert C. Jackson creator: Hitesh B. Mistry uri: https://doi.org/10.7717/peerj.9073 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Jackson and Mistry title: Assessment and simulation of land use and land cover change impacts on the land surface temperature of Chaoyang District in Beijing, China link: https://peerj.com/articles/9115 last-modified: 2020-05-11 description: Rapid urbanization is changing the existing patterns of land use land cover (LULC) globally, which is consequently increasing the land surface temperature (LST) in many regions. The present study is focused on estimating current and simulating future LULC and LST trends in the urban environment of Chaoyang District, Beijing. Past patterns of LULC and LST were identified through the maximum likelihood classification (MLC) method and multispectral Landsat satellite images during the 1990–2018 data period. The cellular automata (CA) and stochastic transition matrix of the Markov model were applied to simulate future (2025) LULC and LST changes, respectively, using their past patterns. The CA model was validated for the simulated and estimated LULC for 1990–2018, with an overall Kappa (K) value of 0.83, using validation modules in IDRISI software. Our results indicated that the cumulative changes in built-up to vegetation area were 74.61 km2 (16.08%) and 113.13 km2 (24.38%) from 1990 to 2018. The correlation coefficient of land use and land cover change (LULCC), including vegetation, water bodies and built-up area, had values of r =  − 0.155 (p > 0.005), −0.809 (p = 0.000), and 0.519 (p > 0.005), respectively. The results of future analysis revealed that there will be an estimated 164.92 km2 (−12%) decrease in vegetation area, while an expansion of approximately 283.04 km2 (6% change) will occur in built-up areas from 1990 to 2025. This decrease in vegetation cover and expansion of settlements would likely cause a rise of approximately ∼10.74 °C and ∼12.66 °C in future temperature, which would cause a rise in temperature (2025). The analyses could open an avenue regarding how to manage urban land cover patterns to enhance the resilience of cities to climate warming. This study provides scientific insights for environmental development and sustainability through efficient and effective urban planning and management in Beijing and will also help strengthen other research related to the UHI phenomenon in other parts of the world. creator: Muhammad Amir Siddique creator: Liu Dongyun creator: Pengli Li creator: Umair Rasool creator: Tauheed Ullah Khan creator: Tanzeel Javaid Aini Farooqi creator: Liwen Wang creator: Boqing Fan creator: Muhammad Awais Rasool uri: https://doi.org/10.7717/peerj.9115 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Amir Siddique et al. title: Higher carbon sequestration potential and stability for deep soil compared to surface soil regardless of nitrogen addition in a subtropical forest link: https://peerj.com/articles/9128 last-modified: 2020-05-11 description: BackgroundLabile carbon input could stimulate soil organic carbon (SOC) mineralization through priming effect, resulting in soil carbon (C) loss. Meanwhile, labile C could also be transformed by microorganisms in soil as the processes of new C sequestration and stabilization. Previous studies showed the magnitude of priming effect could be affected by soil depth and nitrogen (N). However, it remains unknown how the soil depth and N availability affect the amount and stability of the new sequestrated C, which complicates the prediction of C dynamics.MethodsA 20-day incubation experiment was conducted by adding 13C labeled glucose and NH4NO3 to study the effects of soil depth and nitrogen addition on the net C sequestration. SOC was fractioned into seven fractions and grouped into three functional C pools to assess the stabilization of the new sequestrated C.ResultsOur results showed that glucose addition caused positive priming in both soil depths, and N addition significantly reduced the priming effect. After 20 days of incubation, deep soil had a higher C sequestration potential (48% glucose-C) than surface soil (43% glucose-C). The C sequestration potential was not affected by N addition in both soil depths. Positive net C sequestration was observed with higher amount of retained glucose-C than that of stimulated mineralized SOC for both soil depths. The distribution of new sequestrated C in the seven fractions was significantly affected by soil depth, but not N addition. Compared to deep soil, the new C in surface soil was more distributed in the non-protected C pool (including water extracted organic C, light fraction and sand fraction) and less distributed in the clay fraction. These results suggested that the new C in deep soil was more stable than that in surface soil. Compared to the native SOC for both soil depths, the new sequestrated C was more distributed in non-protected C pool and less distributed in biochemically protected C pool (non-hydrolyzable silt and clay fractions). The higher carbon sequestration potential and stability in deep soil suggested that deep soil has a greater role on C sequestration in forest ecosystems. creator: Chang Liao creator: Dong Li creator: Lin Huang creator: Pengyun Yue creator: Feng Liu creator: Qiuxiang Tian uri: https://doi.org/10.7717/peerj.9128 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liao et al. title: Phylogenetic relationships in the southern African genus Drosanthemum (Ruschioideae, Aizoaceae) link: https://peerj.com/articles/8999 last-modified: 2020-05-08 description: BackgroundDrosanthemum, the only genus of the tribe Drosanthemeae, is widespread over the Greater Cape Floristic Region in southern Africa. With 114 recognized species, Drosanthemum, together with the highly succulent and species-rich tribe Ruschieae, constitute the ‘core ruschioids’ in Aizoaceae. Within Drosanthemum, nine subgenera have been described based on flower and fruit morphology. Their phylogenetic relationships, however, have not yet been investigated, hampering understanding of monophyletic entities and patterns of geographic distribution.MethodsUsing chloroplast and nuclear DNA sequence data, we performed network- and tree-based phylogenetic analyses of 73 species of Drosanthemum with multiple accessions for widespread species. A well-curated, geo-referenced occurrence dataset comprising the 134 genetically analysed and 863 further accessions was used to describe the distributional ranges of intrageneric lineages and the genus as a whole.ResultsPhylogenetic inference supports nine clades within Drosanthemum, seven of which group in two major clades, while the remaining two show ambiguous affinities. The nine clades are generally congruent to previously described subgenera within Drosanthemum, with exceptions such as cryptic species. In-depth analyses of sequence patterns in each gene region were used to reveal phylogenetic affinities inside the retrieved clades in more detail. We observe a complex distribution pattern including widespread, species-rich clades expanding into arid habitats of the interior (subgenera Drosanthemum p.p., Vespertina, Xamera) that are genetically and morphologically diverse. In contrast, less species-rich, genetically less divergent, and morphologically unique lineages are restricted to the central Cape region and more mesic conditions (Decidua, Necopina, Ossicula, Quastea, Quadrata, Speciosa). Our results suggest that the main lineages arose from an initial rapid radiation, with subsequent diversification in some clades. creator: Sigrid Liede-Schumann creator: Guido W. Grimm creator: Nicolai M. Nürk creator: Alastair J. Potts creator: Ulrich Meve creator: Heidrun E.K. Hartmann uri: https://doi.org/10.7717/peerj.8999 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liede-Schumann et al. title: Hellebrigenin anti-pancreatic cancer effects based on apoptosis and autophage link: https://peerj.com/articles/9011 last-modified: 2020-05-08 description: Hellebrigenin is a natural product found in the toad skin secretions and plants of Urginea, including Hellebores and Kalanchoe genera. It has been shown to be active against Leishmania chagasi promastigotes and Trypanosoma cruzi trypomastigotes and also reported to play an anti-tumor effect on several cancer cell lines in vitro, including pancreatic cancer. This study is aimed to investigate the effects of Hellebrigenin on pancreatic carcinoma cells, SW1990 and BxPC-3 in vitro and its molecular mechanism involved in antitumor activities. Our results showed that Hellebrigenin effectively inhibited the proliferation of SW1990 and BxPC-3 cells in dose- and time-dependent manner. Flow cytometry results showed that Hellebrigenin induced the G0/G1 arrest in both of SW1990 and BxPC-3 cells and promoted cell early apoptosis and autophagy according to morphological observation. Immunofluorescence staining results further confirmed that cell apoptosis and autophagy also increased upon the Hellebrigenin treatment. Moreover, higher dose of Hellebrigenin further increased the cell apoptosis rate while decrease the mitochondrial membrane potential 24 h after treatment. The autophagy rate increased 48 h after treatment with significant difference (P < 0.05). Western blot analysis showed that the expression of caspase 3, 7, cleaved caspase 7, Atg 12, LC3 proteins were increased in SW1990 cell after treatment with Hellebrigenin. In addition, increasing expression of caspase 3, 7, 9, PARP, cleaved caspase 3, 7, 9, PARP, the sub basic protein of the PI3K family, Beclin-1, LC 3, Atg 3, 5, 12, 16 L were also observed after BxPC-3 cells treated with Hellebrigenin. In summary, this study reported for the first time that Hellebrigenin effectively induced autophagy and apoptosis especially the early apoptosis in SW1990 and BxPC-3 cells. creator: Xiaolu Wei creator: Jing He creator: Bo Gao creator: Lingyu Han creator: Yingqiu Mao creator: Haiyu Zhao creator: Nan Si creator: Hongjie Wang creator: Jian Yang creator: Baolin Bian uri: https://doi.org/10.7717/peerj.9011 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wei et al. title: Characteristics of the soil microbial community in the forestland of Camellia oleifera link: https://peerj.com/articles/9117 last-modified: 2020-05-08 description: Characterizing soil microbial community is important for forest ecosystem management and microbial utilization. The microbial community in the soil beneath Camellia oleifera, an important woody edible oil tree in China, has not been reported before. Here, we used Illumina sequencing of 16S and ITS rRNA genes to study the species diversity of microorganisms in C. oleifera forest land in South China. The results showed that the rhizosphere soil had higher physicochemical properties, enzyme activities and microbial biomass than did the non-rhizosphere soil. The rhizosphere soil microorganisms had a higher carbon source utilization capacity than the non-rhizosphere soil microorganisms, and attained the highest utilization capacity in summer. The soil microbial community of C. oleifera was characterized by rich ester and amino acid carbon sources that played major roles in the principal functional components of the community. In summer, soil microbes were abundant in species richness and very active in community function. Rhizosphere microorganisms were more diverse than non-root systems in species diversity, which was associated with soil pH, Available phosphorous (AP) and Urease (URE). These results indicated that microbial resources were rich in rhizosphere soil. A priority should be given to the rhizosphere microorganisms in the growing season in developing and utilizing soil microorganisms in C. oleifera plantation. It is possible to promote the growth of C. oleifera by changing soil microbial community, including carbon source species, pH, AP, and URE. Our findings provide valuable information to guide microbial isolation and culturing to manage C. oleifera land. creator: Peng Zhang creator: Zhiyi Cui creator: Mengqing Guo creator: Ruchun Xi uri: https://doi.org/10.7717/peerj.9117 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: Impact of electrocardiographic morphology on clinical outcomes in patients with non-ST elevation myocardial infarction receiving coronary angiography and intervention: a retrospective study link: https://peerj.com/articles/8796 last-modified: 2020-05-07 description: BackgroundThe impact of electrocardiography (ECG) morphology on clinical outcomes in patients with non-ST segment elevation myocardial infarction (NSTEMI) receiving percutaneous coronary intervention (PCI) is unknown. This study investigated whether different ST morphologies had different clinical outcomes in patients with NSTEMI receiving PCI.MethodsThis retrospective study analyzed record-linked data of 362 patients who had received PCI for NSTEMI between January 2008 and December 2010. ECG revealed ST depression in 67 patients, inverted T wave in 91 patients, and no significant ST-T changes in 204 patients. The primary endpoint was long-term all-cause mortality. The secondary endpoint was long-term cardiac death and non-fatal major adverse cardiac events.ResultsCompared to those patients whose ECG showed an inverted T wave and non-specific ST-T changes, patients whose ECG showed ST depression had more diabetes mellitus, advanced chronic kidney disease (CKD) and left main artery disease, as well as more in-hospital mortality, cardiac death and pulmonary edema during hospitalization. Patients with ST depression had a significantly higher rate of long-term total mortality and cardiac death. Finally, multiple stepwise Cox regression analysis showed that an advanced Killip score, age, advanced CKD, prior percutaneous transluminal coronary angioplasty and ST depression were independent predictors of the primary endpoint.ConclusionsAmong NSTEMI patients undergoing coronary angiography, those with ST depression had more in-hospital mortality and cardiac death. Long-term follow-up of patients with ST depression consistently reveals poor outcomes. creator: Chiung-Jen Wu creator: Kuo-Ho Yeh creator: Hui-Ting Wang creator: Wen-Hao Liu creator: Huang-Chung Chen creator: Han-Tan Chai creator: Wen-Jung Chung creator: Shukai Hsueh creator: Chien-Jen Chen creator: Hsiu-Yu Fang creator: Yung-Lung Chen uri: https://doi.org/10.7717/peerj.8796 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wu et al. title: Integrated analysis of lymphocyte infiltration-associated lncRNA for ovarian cancer via TCGA, GTEx and GEO datasets link: https://peerj.com/articles/8961 last-modified: 2020-05-07 description: BackgroundAbnormal expression of long non-coding RNAs (lncRNA) play a significant role in the incidence and progression of high-grade serous ovarian cancer (HGSOC), which is a leading cause of mortality among gynecologic malignant tumor patients. In this study, our aim is to identify lncRNA-associated competing endogenous RNA (ceRNA ) axes that could define more reliable prognostic parameters of HGSOC, and to investigate the lncRNAs’ potential mechanism of in lymphocyte infiltration.MethodsThe RNA-seq and miRNA expression profiles were downloaded from The Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) database; while for obtaining the differentially expressed lncRNAs (DELs), miRNAs (DEMs), and genes (DEGs), we used edgeR, limma and DESeq2. After validating the RNA, miRNA and gene expressions, using integrated three RNA expression profiles (GSE18520, GSE27651, GSE54388) and miRNA profile (GSE47841) from the Gene Expression Omnibus (GEO) database, we performed Gene Ontology (GO) and Kyoto Encyclopedia of Gene and Genome (KEGG) pathway analyses through ClusterProfiler. The prognostic value of these genes was determined with Kaplan–Meier survival analysis and Cox regression analysis. The ceRNA network was constructed using Cytoscape. The correlation between lncRNAs in ceRNA network and immune infiltrating cells was analyzed by using Tumor IMmune Estimation Resource (TIMER), and gene markers of tumor-infiltrating immune cells were identified using Spearman’s correlation after removing the influence of tumor purity.ResultsA total of 33 DELs (25 upregulated and eight downregulated), 134 DEMs (76 upregulated and 58 downregulated), and 1,612 DEGs (949 upregulated and 663 downregulated) were detected that could be positively correlated with overall survival (OS) of HGSOC. With the 1,612 analyzed genes, we constructed a ceRNA network, which indicated a pre-dominant involvement of the immune-related pathways. Furthermore, our data revealed that LINC00665 influenced the infiltration level of macrophages and dendritic cells (DCs). On the other hand, FTX and LINC00665, which may play their possible roles through the ceRNA axis, demonstrated a potential to inhibit Tregs and prevent T-cell exhaustion.ConclusionWe defined several prognostic biomarkers for the incidence and progression of HGSOC and constructed a network for ceRNA axes; among which three were indicated to have a positive correlation with lymphocyte infiltration, namely: FTX-hsa-miR-150-5p-STK11, LINC00665-hsa-miR449b-5p-VAV3 and LINC00665-hsa-miR449b-5p-RRAGD. creator: Meijing Wu creator: Xiaobin Shang creator: Yue Sun creator: Jing Wu creator: Guoyan Liu uri: https://doi.org/10.7717/peerj.8961 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wu et al. title: Association of heart rate trajectories with the risk of adverse outcomes in a community-based cohort in Taiwan link: https://peerj.com/articles/8987 last-modified: 2020-05-07 description: Heart rate trajectory patterns integrate information regarding multiple heart rate measurements and their changes with time. Different heart rate patterns may exist in one population, and these are associated with different outcomes. Our study investigated the association of adverse outcomes with heart rate trajectory patterns. This was a prospective cohort study based on the Chin-Shan Community Cardiovascular Cohort in Taiwan. A total of 3,015 Chinese community residents aged > 35 years were enrolled in a prospective investigation of cardiovascular risk factors and outcomes from 1990 to 2013.The primary outcome was all-cause mortality, and the secondary outcome was a composite of coronary artery disease and cerebrovascular accidents. The following trajectory patterns were identified: stable, 61%; decreased, 5%; mildly increased, 32%; and markedly increased, 2%. During follow-up (median, 13.9 years), 557 participants died and 217 experienced secondary outcomes. The adjusted hazard ratios of primary and secondary outcomes for participants with a markedly increased trajectory pattern were 1.80 (95% CI [1.18–2.76]) and 1.45 (95% CI [0.67–3.12]), respectively, compared to those for participants with a stable trajectory pattern. A markedly increased heart rate trajectory pattern may be associated with all-cause mortality risks. Heart rate trajectory patterns demonstrated the utility of repeated heart rate measurements for risk assessment. creator: Cheng-Chun Wei creator: Pei-Chun Chen creator: Hsiu-Ching Hsu creator: Ta-Chen Su creator: Hung-Ju Lin creator: Ming-Fong Chen creator: Yuan-Teh Lee creator: Kuo-Liong Chien uri: https://doi.org/10.7717/peerj.8987 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wei et al. title: Understanding PM2.5 concentration and removal efficiency variation in urban forest park—Observation at human breathing height link: https://peerj.com/articles/8988 last-modified: 2020-05-07 description: To increase our knowledge of PM2.5 concentrations near the surface in a forest park in Beijing, an observational study measured the concentration and composition of PM2.5 in Beijing Olympic Forest Park from 2014 to 2015. This study analyzed the meteorological factors and removal efficiency at 1.5 m above the ground (human breathing height) over the day in the forest. The results showed that the average concentrations of PM2.5 near the surface peaked at 07:00–09:30 and reached their lowest at 12:00–15:00. Besides, the results showed that the annual concentration of PM2.5 in the forest was highest during winter, followed by spring and fall, and was lowest during summer. The main chemical components of PM2.5 near the surface in the forest were SO42− and NO3−, which accounted for 68.72% of all water-soluble ions that we observed. The concentration of PM2.5 in the forest had a significant positive correlation with relative humidity and a significant negative correlation with temperature. The removal efficiency near the surface showed no significant variation through the day or year. In the forest, the highest removal efficiency occurred between 07:00 and 09:30 in summer, while the lowest occurred between 09:30 and 12:00 in winter. creator: Guoxin Yan creator: Zibo Yu creator: Yanan Wu creator: Jiakai Liu creator: Yu Wang creator: Jiexiu Zhai creator: Ling Cong creator: Zhenming Zhang uri: https://doi.org/10.7717/peerj.8988 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yan et al. title: Tetrandrine alleviates cerebral ischemia/reperfusion injury by suppressing NLRP3 inflammasome activation via Sirt-1 link: https://peerj.com/articles/9042 last-modified: 2020-05-07 description: Background & AimsTetrandrine (Tet) has been reported to have anti-inflammatory effects and protect from the ischemic strokes. The NLRP3 inflammasome plays a key role in cerebral ischemia/reperfusion (I/R)-induced inflammatory lesions. However, the molecular mechanisms of Tet related to the progression of cerebral ischemia are still unclear. Therefore, the aim of this study was to investigate the possible effects of Tet on cerebral ischemia and the related mechanisms involved in NLRP3 inflammasome.MethodsC57BL/6J mice used as a cerebral I/R injury model underwent middle cerebral artery occlusion (MCAO) for 2 h following reperfusion for 24 h. Tet (30 mg/kg/day, i.p.) was administered for seven days and 30 min before and after MCAO. Their brain tissues were evaluated for NLRP3 inflammasome and Sirtuin-1 (Sirt-1) expression. An intracerebroventricular injection of Sirt-1 siRNA was administered to assess the activation of the NLRP3 inflammasome.ResultsTet significantly reduced the neurological deficits, infarction volume, and cerebral water content in MCAO mice. Moreover, it inhibited I/R-induced over expression of NLRP3, cleaved caspase-1, interleukin (IL)-1β, IL-18, and Sirt-1. Sirt-1 knockdown with siRNA greatly blocked the Tet-induced reduction of neurological severity score and infarct volume, and reversed the inhibition of NLRP3 inflammasome activation.ConclusionOur results demonstrate that Tet has benefits for cerebral I/R injury, which are partially related to the suppression of NLRP3 inflammasome activation via upregulating Sirt-1. creator: Jun Wang creator: Ming Guo creator: Ruojia Ma creator: Maolin Wu creator: Yamei Zhang uri: https://doi.org/10.7717/peerj.9042 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Wang et al. title: Public perspectives and media reporting of wolf reintroduction in Colorado link: https://peerj.com/articles/9074 last-modified: 2020-05-07 description: In the state of Colorado, a citizen ballot initiative to reintroduce gray wolves (Canis lupus) is eliciting polarization and conflict among multiple stakeholder and interest groups. Given this complex social landscape, we examined the social context surrounding wolf reintroduction in Colorado as of 2019. We used an online survey of 734 Coloradans representative in terms of age and gender, and we sampled from different regions across the state, to examine public beliefs and attitudes related to wolf reintroduction and various wolf management options. We also conducted a content analysis of media coverage on potential wolf reintroduction in 10 major daily Colorado newspapers from January 2019, when the signature-gathering effort for the wolf reintroduction initiative began, through the end of January 2020, when the initiative was officially added to the ballot. Our findings suggest a high degree of social tolerance or desire for wolf reintroduction in Colorado across geographies, stakeholder groups, and demographics. However, we also find that a portion of the public believes that wolves would negatively impact their livelihoods, primarily because of concerns over the safety of people and pets, loss of hunting opportunities, and potential wolf predation on livestock. These concerns—particularly those related to livestock losses—are strongly reflected in the media. We find that media coverage has focused only on a few of the many perceived positive and negative impacts of wolf reintroduction identified among the public. Our findings highlight the need to account for this diversity of perspectives in future decisions and to conduct public outreach regarding likely impacts of wolf reintroduction. creator: Rebecca Niemiec creator: Richard E.W. Berl creator: Mireille Gonzalez creator: Tara Teel creator: Cassiopeia Camara creator: Matthew Collins creator: Jonathan Salerno creator: Kevin Crooks creator: Courtney Schultz creator: Stewart Breck creator: Dana Hoag uri: https://doi.org/10.7717/peerj.9074 license: https://creativecommons.org/publicdomain/zero/1.0/ rights: title: Resampling-based methods for biologists link: https://peerj.com/articles/9089 last-modified: 2020-05-07 description: Ecological data often violate common assumptions of traditional parametric statistics (e.g., that residuals are Normally distributed, have constant variance, and cases are independent). Modern statistical methods are well equipped to handle these complications, but they can be challenging for non-statisticians to understand and implement. Rather than default to increasingly complex statistical methods, resampling-based methods can sometimes provide an alternative method for performing statistical inference, while also facilitating a deeper understanding of foundational concepts in frequentist statistics (e.g., sampling distributions, confidence intervals, p-values). Using simple examples and case studies, we demonstrate how resampling-based methods can help elucidate core statistical concepts and provide alternative methods for tackling challenging problems across a broad range of ecological applications. creator: John R. Fieberg creator: Kelsey Vitense creator: Douglas H. Johnson uri: https://doi.org/10.7717/peerj.9089 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Fieberg et al. title: Evaluation of a novel syndromic surveillance system for the detection of the 2007 melamine-related nephrotoxicosis foodborne outbreak in dogs and cats in the United States link: https://peerj.com/articles/9093 last-modified: 2020-05-07 description: The 2007 nephrotoxicosis outbreak associated with melamine and cyanuric acid adulteration of pet foods in the United States sparked an urgent need for a nationwide companion animal surveillance program. In 2016, we introduced a syndromic surveillance system based on a novel epidemiological algorithm, the proportionate diagnostic outcome ratio (PDOR). The PDOR procedure was validated using simulated outbreaks of foodborne illness (i.e., aflatoxicosis and gastrointestinal illness) in dogs and cats. In this study, we further evaluated the PDOR procedure using the 2007 melamine-related outbreak of nephrotoxicosis. The performance of the PDOR procedure was assessed by the time to alert and positive predictive value (PPV). Electronic medical records of dogs and cats seen at networked primary care veterinary hospitals across the United States were retrieved from a centralized database. The data of four relevant syndromic components: elevated serum creatinine concentration, vomiting, anorexia, and lethargy from July 28, 2006 to May 31, 2007 were prospectively analyzed using the PDOR algorithm. The results showed that the alerts generated from the analysis of elevated serum creatinine concentration could have led to an early detection of this nephrotoxicosis foodborne outbreak and were well matched to the reported timeline of the outbreak. Additionally, we also observed variations in the performance of the PDOR procedure across age of animals and syndromic components, with the PPVs ranged from 0.61 to 1.0. Combined with the findings from previous evaluations using simulated outbreak scenarios, this study provided additional evidence that the PDOR procedure can be applied in syndromic surveillance to effectively and accurately detect various types of foodborne illness outbreaks in companion animals. However, the interpretations of and responses to alerts require an understanding of clinical veterinary medicine and relevant syndromic knowledge, and should not be based solely on quantitative measures. creator: Hsin-Yi Weng creator: Mark A.L. Gaona creator: Philip H. Kass uri: https://doi.org/10.7717/peerj.9093 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Weng et al. title: Genome-based development of 15 microsatellite markers in fluorescent multiplexes for parentage testing in captive tigers link: https://peerj.com/articles/8939 last-modified: 2020-05-06 description: As one of the most endangered species, tiger (Panthera tigris) inbreeding has become an urgent issue to address. Using a microsatellite (short tandem repeat, STR) identification system, paternity testing may be helpful to avoid inbreeding in captive breeding programs. In this study, we developed a genome-based identification system named tiger pedigree identification multiplex system (TPI-plex). By analyzing the entire tiger genome, 139,967 STR loci were identified and 12.76% of these displayed three to six alleles among three re-sequenced individual tiger genomes. A total of 204 candidate STRs were identified and screened with a reference population containing 31 unrelated captive tigers. Of these, 15 loci were chosen for inclusion in the multiplex panel. The mean allele number and mean expected heterozygosity (He) were 7.3333 and 0.7789, respectively. The cumulative probability of exclusion (CPE) and total probability of discrimination power (TDP) reached 0.999999472 and 0.999999999999995, respectively. The results showed that the TPI-plex system can be applied in routine pedigree identification for captive tigers. We also added a sex identification marker named TAMEL into the TPI-plex for sex determination. creator: Xiao Zhao creator: Qiguan Qiu creator: Chang Li creator: Dongke Fu creator: Xuesong Hu creator: Shengjie Gao creator: Yugang Zhu creator: Haofang Mu creator: Runping Wang creator: Huanming Yang creator: Bo Li uri: https://doi.org/10.7717/peerj.8939 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zhao et al. title: Cloning, sequencing, and expression analysis of 32 NAC transcription factors (MdNAC) in apple link: https://peerj.com/articles/8249 last-modified: 2020-05-06 description: BackgroundNAC transcription factors play important roles in the regulation of plant growth, development, abiotic and biotic stress responses. The transcriptional level of MdNACs in different tissues and under various biotic and abiotic stress treatments was determined to provide a solid foundation for studying the function of MdNACs.MethodsThirty-two full-length cDNA sequences of Md NACs were isolated by homologous comparison and RT-PCR confirmation, and the obtained cDNA sequences and the deduced amino acid sequences were analyzed with bioinformatics methods. The prediction of subcellular locations of MdNAC proteins was performed using CELLO v.2.5, PSORT, and SoftBerry ProtComp 9.0. Expression levels of MdNACs were detected in 16 different tissues using an array. Expression patterns of MdNACs were detected in response to Alternaria alternata apple pathotype (AAAP) infection using RNA-seq, and the expression of MdNACs was analyzed under NaCl and mannitol treatments using RT-qPCR.ResultsThe sequencing results produced 32 cDNAs (designated as MdNAC24-39, MdNAC54-65, and MdNAC67-70 with GenBank accession No. MG099861–MG099876, MG099891–MG099902, and MG099904–MG099907, respectively). Phylogenetic analysis revealed that MdNAC34 belonged to the ATAF group, MdNAC63 belonged to the AtNAC3 group, MdNAC24, MdNAC26-30, MdNAC32-33, MdNAC35, MdNAC37-39, MdNAC56-57, MdNAC59-62, MdNAC64-65, and MdNAC67-70 belonged to the NAM group, and MdNAC25, MdNAC36, MdNAC54-55, and MdNAC58 belonged to the VND group. Predictions of subcellular localization showed that MdNAC24-27, MdNAC29-30, MdNAC33-37, MdNAC39, MdNAC54-65, and MdNAC67-70 proteins were located in the nucleus, MdNAC28 proteins were located in the cytoplasm, MdNAC31-32 proteins were located in the nucleus and cytoplasm, and MdNAC38 proteins were located in the nucleus and plasma membrane. Array results indicated that 32 MdNACs were expressed in all examined tissues at various expression levels. RNA-seq results showed that expression levels of MdNAC26-28, MdNAC33-34, MdNAC60, MdNAC62-65, and MdNAC68 were induced, but MdNAC24, MdNAC32, and MdNAC58 were down-regulated in response to AAAP infection. Under salt treatment, MdNAC24, MdNAC27, MdNAC29, MdNAC34, MdNAC37, MdNAC39, MdNAC54, MdNAC59, and MdNAC63 transcription levels were induced. Under mannitol treatment, MdNAC32 and MdNAC54 transcription levels were induced, but MdNAC24, MdNAC28, MdNAC30, MdNAC33, MdNAC35, MdNAC37, MdNAC55, MdNAC56, MdNAC58, and MdNAC59 were down-regulated. Taken together, the results indicated that the cloned MdNAC genes were expressed constitutively in all examined tissues. These genes were up-regulated or down-regulated in response to AAAP infection and to salt or mannitol, which suggested they may be involved in the regulation of growth, development, and stress response in apple. creator: Huifeng Li creator: Kun Ran creator: Qinglong Dong creator: Qiang Zhao creator: Song Shi uri: https://doi.org/10.7717/peerj.8249 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Li et al. title: Predicting the consequences of global warming on Gentiana lutea germination at the edge of its distributional and ecological range link: https://peerj.com/articles/8894 last-modified: 2020-05-06 description: BackgroundTemperature is the main environmental factor controlling seed germination; it determines both the percentage and the rate of germination. According to the Intergovernmental Panel on Climate Change, the global mean surface temperature could increase of approximately 2–4 °C by 2090–2099. As a consequence of global warming, the period of snow cover is decreasing on several mountain areas. Thermal time approach can be used to characterise the seed germination of plants and to evaluate the germination behaviour under the climate change scenarios. In this study, the effect of different cold stratification periods on seed dormancy release and germination of Gentiana lutea subsp. lutea, a taxon listed in Annex V of the Habitats Directive (92/43/EEC), was evaluated. Furthermore, the thermal requirements and the consequences of the temperature rise for seed germination of this species were estimated. In addition, a conceptual representation of the thermal time approach is presented.MethodsSeeds of G. lutea subsp. lutea were harvested from at least 50 randomly selected plants in two representative localities of the Gennargentu massif (Sardinia). Germination tests were carried out under laboratory conditions and the responses at 5, 10, 15, 20, 25 and 30 °C were recorded. Different cold stratification pre-treatments at 1 ± 1 °C (i.e. 0, 15, 30, 60 and 90 days) were applied. Successively, the base temperature (Tb) and the number of thermal units (θ, °Cd) for germination were estimated. Additionally, this study examined the consequences of an increase in temperatures based on the Representative Concentration Pathways (RPC) scenarios.ResultsThe results indicated that from 0 to 30 days of cold stratification, the germination was null or very low. After 60 and 90 days of cold stratification the seed dormancy was removed; however, 25 and 30 °C negatively affected the germination capacity of non-dormant seeds. Seeds cold-stratified for 90 days showed a lower Tb than those stratified for 60 days. However, 60 and 90 days of cold stratification did not cause great variations in the thermal time units. Analysing the RPC scenarios, we detected that the number of days useful for dormancy release of seeds of G. lutea may be less than 30 days, a condition that does not permit an effective dormancy release.ConclusionsWe conclude that seeds of G. lutea need at least 60 days of cold stratification to remove dormancy and promote the germination. The thermal time model developed in this work allowed us to identify the thermal threshold requirements of seed germination of this species, increasing the knowledge of a plant threatened by global warming. Our results emphasise the need for further studies aiming at a better characterisation of germination efficiency, especially for species that require cold stratification. This would improve the knowledge on the germination mechanisms of adaptation to different future global warming conditions. creator: Alba Cuena-Lombraña creator: Marco Porceddu creator: Caterina Angela Dettori creator: Gianluigi Bacchetta uri: https://doi.org/10.7717/peerj.8894 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Cuena-Lombraña et al. title: Evaluating the stress-response of dental students to the dental school environment link: https://peerj.com/articles/8981 last-modified: 2020-05-06 description: Introduction and ObjectiveDentists experience high amounts of professional stress beginning with their student years in dental school. This stress, given its early onset, may negatively impact the personal and professional lives of these individuals, as well as the quality of their clinical work. We sought to create an objective scale to evaluate the levels of stress in students at different stages of their education, as well as in practicing physicians.Materials and MethodsThirty dental students participated in this study, with 10 students each selected from junior, mid-senior, and senior classes. They were randomly divided into two groups in which one group was subjected to stressors while the other group was not. JINS MEME ES_R (JINS) smart glasses and Garmin Vivoactive 3 smartwatches were used to obtain data, including electrooculography (EOG), heart rate (HR), and accelerometer (ACC) and gyroscope (GYRO) feedback, while the subjects performed a dental exercise on a phantom tooth.ResultsThe heart rates of more experienced students were lower than those of the junior students. The EOG, ACC, and GYRO signals showed multiple differences in the measurement of amplitudes and frequency of episodes.ConclusionOur pilot results show that electronic tools, like smart glasses with software and sensors, are useful for monitoring the stress levels of dental students in preclinical operating conditions. We would like to further assess the stress levels in students performing dental procedures on phantom teeth and in later clinical interactions with patients. creator: Katarzyna Mocny-Pachońska creator: Rafał Doniec creator: Agata Trzcionka creator: Marek Pachoński creator: Natalia Piaseczna creator: Szymon Sieciński creator: Oleksandra Osadcha creator: Patrycja Łanowy creator: Marta Tanasiewicz uri: https://doi.org/10.7717/peerj.8981 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Mocny-Pachońska et al. title: Identification of key miRNAs in the progression of hepatocellular carcinoma using an integrated bioinformatics approach link: https://peerj.com/articles/9000 last-modified: 2020-05-06 description: BackgroudIt has been shown that aberrant expression of microRNAs (miRNAs) and transcriptional factors (TFs) is tightly associated with the development of HCC. Therefore, in order to further understand the pathogenesis of HCC, it is necessary to systematically study the relationship between the expression of miRNAs, TF and genes. In this study, we aim to identify the potential transcriptomic markers of HCC through analyzing common microarray datasets, and further establish the differential co-expression network of miRNAs–TF–mRNA to screen for key miRNAs as candidate diagnostic markers for HCC.MethodWe first downloaded the mRNA and miRNA expression profiles of liver cancer from the GEO database. After pretreatment, we used a linear model to screen for differentially expressed genes (DEGs) and miRNAs. Further, we used weighed gene co-expression network analysis (WGCNA) to construct the differential gene co-expression network for these DEGs. Next, we identified mRNA modules significantly related to tumorigenesis in this network, and evaluated the relationship between mRNAs and TFs by TFBtools. Finally, the key miRNA was screened out in the mRNA–TF–miRNA ternary network constructed based on the target TF of differentially expressed miRNAs, and was further verified with external data set.ResultsA total of 465 DEGs and 215 differentially expressed miRNAs were identified through differential genes expression analysis, and WGCNA was used to establish a co-expression network of DEGs. One module that closely related to tumorigenesis was obtained, including 33 genes. Next, a ternary network was constructed by selecting 256 pairs of mRNA–TF pairs and 100 pairs of miRNA–TF pairs. Network mining revealed that there were significant interactions between 18 mRNAs and 25 miRNAs. Finally, we used another independent data set to verify that miRNA hsa-mir-106b and hsa-mir-195 are good classifiers of HCC and might play key roles in the progression of HCC.ConclusionOur data indicated that two miRNAs—hsa-mir-106b and hsa-mir-195—are identified as good classifiers of HCC. creator: Qi Zheng creator: Xiaoyong Wei creator: Jun Rao creator: Cuncai Zhou uri: https://doi.org/10.7717/peerj.9000 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zheng et al. title: Anthropocene geochemistry of metals in sediment cores from the Laurentian Great Lakes link: https://peerj.com/articles/9034 last-modified: 2020-05-06 description: Geochemical analyses applied to lake sedimentary records can reveal the history of pollution by metals and the effects of remedial efforts. Lakes provide ideal environments for geochemical studies because they have steady deposition of fine grained material suitable for fixation of pollutants. The Laurentian Great Lakes are the most studied system in this field, and they have well-preserved chronological profiles. To date, this important system has been considered in parts for inorganic geochemistry, hampering basin-wide conclusions regarding metal contamination. We filled spatial and temporal gaps in a comprehensive geochemical analysis of 11 sediment cores collected from all five Great Lakes. Hierarchical cluster analysis of all Great Lakes samples divided the metal analytes into five functional groups: (1) carbonate elements; (2) metals and oxides with diverse natural sources, including a subgroup of analytes known to be anthropogenically enriched (Cd, Pb, Sn, Zn, and Sb); (3) common crustal elements; (4) metals related to coal and nuclear power generation; and (5) all of the co-occurring rare earth elements. Two contamination indices (Igeo and EF) applied to sedimentary metals indicated that Na, Co, Mn, Cd, Pb, Ta, and Cu were each, at some point during the Anthropocene, the most enriched metal pollutants in Great Lakes sediments. Land uses correlated with the metal analytes, such as increases in contaminant metals with the rise in catchment population and increases in carbonate elements (e.g. Ca) with agriculture. Certain contamination trends were observed basin-wide, such as for the atmospheric pollutant Pb, which followed a rise associated with fossil fuel combustion and a decline following the ban of leaded gasoline. Other trends were lake-specific, such as recent high concentrations of Na in Lake Superior, likely due to road salt applications, and a late-20th-century peak in Ca associated with algal whiting events in Lake Ontario. Some metals exceeded guidelines for sediment quality, in some cases prior to European settlement of the basin, indicating that a paleolimnological context is important for appropriate management of sediment contamination. The Great Lakes are sensitive to environmental changes such as pollution by metals, and it is clear that while there has been remedial success, results from the uppermost intervals of cores indicate ongoing problems. creator: Malachi Nicholas Granmo creator: Euan D. Reavie creator: Sara P. Post creator: Lawrence M. Zanko uri: https://doi.org/10.7717/peerj.9034 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Aliff et al. title: CRISPR screening identifies M1AP as a new MYC regulator with a promoter-reporter system link: https://peerj.com/articles/9046 last-modified: 2020-05-06 description: BackgroundMYC is one of the proto-oncogenes contributing to tumorigenesis in many human cancers. Although the mechanism of MYC regulation is still not fully understood, learning about the comprehensive mechanism controlling the transcriptional activity of MYC will lead to therapeutic targets. The CRISPR/Cas9 library system is a simple and powerful screening technique. This study aims to identify new transcriptional upstream activators of MYC using the CRISPR activation library with new promoter-reporter systems.Methods and ResultsThe MYC promoter-reporter system was developed with a photoconvertible fluorescent protein, Dendra2, and named “pMYC-promoter-Dendra2.” This MYC promoter-reporter system was designed to harbor a proximal MYC promoter at (3.1 kb). Both the CRISPR activation library and pMYC-promoter-Dendra2 were induced to HEK 293T cells, and Dendra2-positive cells, that are supposed that MYC should be upregulated, were collected individually by a cell sorter. Among the 169 cells collected, 12 clones were successfully established. Then, pMYC-promoter-Dendra2 was transfected again into these 12 clones, and two of 12 clones showed Dendra2 positivity. In this procedure, the cells with non-specific autofluorescence were correctly distinguished by utilizing the photoswitchable character of Dendra2. Using extracted genomic DNA of these two Dendra2 positive clones, polymerase chain reaction (PCR) was performed to amplify the guide RNA (gRNA) containing region, which was introduced by the CRISPR activation library. Eventually, PLEKHO2, MICU, MBTPS1, and M1AP were identified, and these gRNAs were transfected individually into HEK 293T cells again using the CRISPR activation system. Only M1AP gRNA transfected cells showed Dendra2-positive fluorescence. Then, the overexpression vector for M1AP with a doxycycline-inducible vector confirmed that M1AP induced high MYC expression by real-time quantitative PCR and western blot. Furthermore, the dual-luciferase assay showed a significant increase of promoter activity, and MYC mRNA was higher in M1AP- overexpressing cells. M1AP is highly expressed in several cancers, though, a positive correlation between M1AP and MYC was observed only in human acute myeloid leukemia.ConclusionThe present study confirmed that the experimental method using the CRISPR library technology functions effectively for the identification of molecules that activate endogenous MYC. This method will help elucidate the regulatory mechanism of MYC expression, as well as supporting further drug research against malignant tumors. creator: Akiko Yamamoto creator: Morito Kurata creator: Iichiroh Onishi creator: Keisuke Sugita creator: Miwa Matsumura creator: Sachiko Ishibashi creator: Masumi Ikeda creator: Kouhei Yamamoto creator: Masanobu Kitagawa uri: https://doi.org/10.7717/peerj.9046 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yamamoto et al. title: Metallic elements and oxides and their relevance to Laurentian Great Lakes geochemistry link: https://peerj.com/articles/9053 last-modified: 2020-05-06 description: The Laurentian Great Lakes are the most studied system in lake geochemistry and have well-preserved chronological profiles. Metals play numerous critical roles in natural and anthropogenic characteristics of lake ecosystems, so patterns in the historical records of metals from sedimentary cores provide important information about environmental baselines and human impacts. Relevant studies of Great Lakes geochemistry are listed, and we follow with encyclopedic descriptions of metals and their oxides in the lakes. These descriptions include likely natural and anthropogenic sources of elements, their known history from previous paleoecological studies, and their status as potential contaminants of concern. Despite the well-studied geology of the Great Lakes catchment, sourcing elements was sometimes difficult due to materials often being moved long distances by glaciation and the global prevalence of atmospheric pollutants. We summarized available information on metals and their roles as geochemical indicators in the Great Lakes. creator: Malachi N. Granmo creator: Euan D. Reavie creator: Sara P. Post creator: Lawrence M. Zanko uri: https://doi.org/10.7717/peerj.9053 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Granmo et al. title: Diversity of cultivable protease-producing bacteria and their extracellular proteases associated to scleractinian corals link: https://peerj.com/articles/9055 last-modified: 2020-05-06 description: Protease-producing bacteria play a vital role in degrading organic nitrogen in marine environments. However, the diversity of the bacteria and extracellular proteases has seldom been addressed, especially in communities of coral reefs. In this study, 136 extracellular protease-producing bacterial strains were isolated from seven genera of scleractinian corals from Luhuitou fringing reef, and their protease types were characterized. The massive coral had more cultivable protease-producing bacteria than branching or foliose corals. The abundance of cultivable protease-producing bacteria reached 106 CFU g−1 of coral. Phylogenetic analysis of 16S rRNA gene sequences revealed that the isolates were assigned to 24 genera, from which 20 corresponded to the phyla Firmicutes and Proteobacteria. Bacillus and Fictibacillus were retrieved from all coral samples. Moreover, Vibrio and Pseudovibrio were most prevalent in massive or foliose coral Platygyra and Montipora. In contrast, 11 genera were each identified in only one isolate. Nearly all the extracellular proteases from the bacteria were serine proteases or metalloproteases; 45.83% of isolates also released cysteine or aspartic proteases. These proteases had different hydrolytic ability against different substrates. This study represents a novel insight on the diversity of cultivable protease-producing bacteria and their extracellular proteases in scleractinian corals. creator: Hongfei Su creator: Zhenlun Xiao creator: Kefu Yu creator: Qinyu Huang creator: Guanghua Wang creator: Yinghui Wang creator: Jiayuan Liang creator: Wen Huang creator: Xueyong Huang creator: Fen Wei creator: Biao Chen uri: https://doi.org/10.7717/peerj.9055 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Su et al. title: Identification of key genes associated with progression and prognosis for lung squamous cell carcinoma link: https://peerj.com/articles/9086 last-modified: 2020-05-06 description: BackgroundLung squamous cell carcinoma (LUSC) is a major subtype of lung cancer with limited therapeutic options and poor clinical prognosis.MethodsThree datasets (GSE19188, GSE33532 and GSE33479) were obtained from the gene expression omnibus (GEO) database. Differentially expressed genes (DEGs) between LUSC and normal tissues were identified by GEO2R, and functional analysis was employed using the Database for Annotation, Visualization and Integrated Discovery (DAVID) online tool. Protein–protein interaction (PPI) and hub genes were identified via the Search Tool for the Retrieval of Interacting Genes (STRING) and Cytoscape software. Hub genes were further validated in The Cancer Genome Atlas (TCGA) database. Subsequently, survival analysis was performed using the Kapla–Meier curve and Cox progression analysis. Based on univariate and multivariate Cox progression analysis, a gene signature was established to predict overall survival. Receiver operating characteristic curve was used to evaluate the prognostic value of the model.ResultsA total of 116 up-regulated genes and 84 down-regulated genes were identified. These DEGs were mainly enriched in the two pathways: cell cycle and p53 signaling way. According to the degree of protein nodes in the PPI network, 10 hub genes were identified. The mRNA expression levels of the 10 hub genes in LUSC were also significantly up-regulated in the TCGA database. Furthermore, a novel seven-gene signature (FLRT3, PPP2R2C, MMP3, MMP12, CAPN8, FILIP1 and SPP1) from the DEGs was constructed and acted as a significant and independent prognostic signature for LUSC.ConclusionsThe 10 hub genes might be tightly correlated with LUSC progression. The seven-gene signature might be an independent biomarker with a significant predictive value in LUSC overall survival. creator: Xiaohan Ma creator: Huijun Ren creator: Ruoyu Peng creator: Yi Li creator: Liang Ming uri: https://doi.org/10.7717/peerj.9086 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Ma et al. title: Specificity trumps flexibility—location-based stable associations between Symbiodiniaceae genera and Platygyra verweyi (Scleractinia; Merulinidae) link: https://peerj.com/articles/8791 last-modified: 2020-05-05 description: This study monitored symbiont communities bi-monthly in native coral cores used in a reciprocal transplantation of the coral Platygyra verweyi over two years (2014–2016) and samples of mother colonies from three locations with variable thermal regimes; our results show that associating with multiple Symbiodiniaceae genera (Cladocopium spp. and Durusdinium spp.) is not a prerequisite for symbiont shuffling. Platygyra verweyi associates with certain Symbiodiniaceae genera based on location. Results of quantitative real-time PCR indicated small-scale temporal changes in Symbiodiniaceae genera compositions from 2014 to 2016; however, these changes were not enough to invoke shuffling or switching, despite degree heating weeks exceeding 6 °C-weeks in 2014 and 4 °C-weeks in 2015, which usually resulted in substantial coral bleaching. Microsatellite analysis of the P. verweyi host showed no genetic differences among the study locations. Our results suggest that P. verweyi undergoes long-term acclimatization and/or adaptation based on microgeographic and local environmental conditionsby altering its combinations of associated Symbiodiniaceae. Results also suggest that shuffling might not be as common a phenomenon as it has been given credit for; corals thrive through specific associations, and many corals could still be vulnerable to climate change-induced stress, despite being promiscuous or able to associate with rare and background Symbiodiniaceae genera. creator: Shashank Keshavmurthy creator: Hwee Sze Tee creator: Kuo-Wei Kao creator: Jih-Terng Wang creator: Chaolun Allen Chen uri: https://doi.org/10.7717/peerj.8791 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Keshavmurthy et al. title: PHACTR1 is associated with disease progression in Chinese Moyamoya disease link: https://peerj.com/articles/8841 last-modified: 2020-05-05 description: Moyamoya disease (MMD) is a progressive stenosis at the terminal portion of internal carotid artery and frequently occurs in East Asian countries. The etiology of MMD is still largely unknown. We performed a case-control design with whole-exome sequencing analysis on 31 sporadic MMD patients and 10 normal controls with matched age and gender. Patients clinically diagnosed with MMD was determined by digital subtraction angiography (DSA). Twelve predisposing mutations on seven genes associated with the sporadic MMD patients of Chinese ancestry (CCER2, HLA-DRB1, NSD-1, PDGFRB, PHACTR1, POGLUT1, and RNF213) were identified, of which eight single nucleotide variants (SNVs) were deleterious with CADD PHRED scaled score > 15. Sanger sequencing of nine cases with disease progression and 22 stable MMD cases validated that SNV (c.13185159G>T, p.V265L) on PHACTR1 was highly associated with the disease progression of MMD. Finally, we knocked down the expression of PHACTR1 by transfection with siRNA and measured the cell survival of human coronary artery endothelial cell (HCAEC) cells. PHACTR1 silence reduced the cell survival of HCAEC cells under serum starvation cultural condition. Together, these data identify novel predisposing mutations associated with MMD and reveal a requirement for PHACTR1 in mediating cell survival of endothelial cells. creator: Yongbo Yang creator: Jian Wang creator: Qun Liang creator: Yi Wang creator: Xinhua Chen creator: Qingrong Zhang creator: Shijie Na creator: Yi Liu creator: Ting Yan creator: Chunhua Hang creator: Yichao Zhu uri: https://doi.org/10.7717/peerj.8841 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yang et al. title: Investigating the potential impact of 1.5, 2 and 3 °C global warming levels on crop suitability and planting season over West Africa link: https://peerj.com/articles/8851 last-modified: 2020-05-05 description: West African rainfed agriculture is highly vulnerable to climate variability and change. Global warming is projected to result in higher regional warming and have a strong impact on agriculture. This study specifically examines the impact of global warming levels (GWLs) of 1.5°, 2° and 3 °C relative to 1971–2000 on crop suitability over West Africa. We used 10 Coupled Model Intercomparison Project Phase5 Global Climate Models (CMIP5 GCMs) downscaled by Coordinated Regional Downscaling Experiment (CORDEX) Rossby Centre’s regional Atmospheric model version 4, RCA4, to drive Ecocrop, a crop suitability model, for pearl millet, cassava, groundnut, cowpea, maize and plantain. The results show Ecocrop simulated crop suitability spatial representation with higher suitability, observed to the south of latitude 14°N and lower suitability to its north for 1971–2000 for all crops except for plantain (12°N). The model also simulates the best three planting months within the growing season from September-August over the past climate. Projected changes in crop suitability under the three GWLs 1.5–3.0 °C suggest a spatial suitability expansion for legume and cereal crops, notably in the central southern Sahel zone; root and tuber and plantain in the central Guinea-Savanna zone. In contrast, projected decreases in the crop suitability index value are predicted to the south of 14°N for cereals, root and tuber crops; nevertheless, the areas remain suitable for the crops. A delay of between 1-3 months is projected over the region during the planting month under the three GWLs for legumes, pearl millet and plantain. A two month delay in planting is projected in the south, notably over the Guinea and central Savanna zone with earlier planting of about three months in the Savanna-Sahel zones. The effect of GWL2.0 and GWL3.0 warming in comparison to GWL1.5 °C are more dramatic on cereals and root and tuber crops, especially cassava. All the projected changes in simulated crop suitability in response to climatic variables are statistically significant at 99% confidence level. There is also an increasing trend in the projected crop suitability change across the three warming except for cowpea. This study has implications for improving the resilience of crop production to climate changes, and more broadly, to food security in West Africa. creator: Temitope Samuel Egbebiyi creator: Olivier Crespo creator: Christopher Lennard creator: Modathir Zaroug creator: Grigory Nikulin creator: Ian Harris creator: Jeff Price creator: Nicole Forstenhäusler creator: Rachel Warren uri: https://doi.org/10.7717/peerj.8851 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Egbebiyi et al. title: Responses of rhizosphere soil bacteria to 2-year tillage rotation treatments during fallow period in semiarid southeastern Loess Plateau link: https://peerj.com/articles/8853 last-modified: 2020-05-05 description: BackgroundSoil compaction can be mitigated by deep tillage and subsoiling practices following a long period of no-tillage. Fallow tillage rotation methods are frequently used to improve water availability in the soils of the southeastern Loess Plateau region of China. Rhizosphere soil bacteria are ecologically important for the transformation of matter and energy in the plant root system and can be influenced by tillage rotation treatments. However, the effect of tillage rotations on the bacterial community and structure of rhizosphere soil is not well understood.MethodsA two-year field experiment was conducted with four tillage rotation treatments, including subsoil–subsoil (SS-SS), subsoil–deep tillage (SS-DT), deep tillage–deep tillage (DT-DT), and the control treatment of no-tillage–no-tillage (NT-NT). Our study was conducted during wheat’s fallow period to investigate the abundance, diversity, and functions of rhizosphere soil bacteria using high-throughput sequencing technology.ResultsOur results showed that tillage rotation methods significantly influenced the bacterial diversity and composition of the rhizosphere soil in the plough layer (20–40 cm depth) by altering the moisture content of the soil. The metabolism, environmental information processing, and genetic information processing of the bacteria in the rhizosphere soil were affected. The most abundant phyla across all samples were Proteobacteria, Actinobacteria, Acidobacteria, Planctomycetes, Bacteroidetes, Gemmatimonadetes, Frimicutes, Chloroflexi, Nitrospirae, and Verrucomicrobia, which are classic bacterial decomposers in soil. The bacterial diversity and composition was similar for treatments causing higher soil perturbation (SS-DT and DT-DT), which disrupted the balance between aerobic and anaerobic bacteria. The less disruptive tillage methods (SS-SS and NT-NT), preserved the integrity of the soil bacteria. However, the NT-NT treatment may have led to soil compaction, particularly in the 20–40 cm layer. These results suggested that SS-SS was the most effective tillage rotation practice to accumulate soil moisture, maintain the balance between aerobic and anaerobic bacteria, and to enhance the metabolic capacity of rhizosphere soil bacteria. This method may have a significant impact on the sustainable development and farming practices of dryland agriculture. creator: Qing Xia creator: Xiaoli Liu creator: Zhiqiang Gao creator: Jianming Wang creator: Zhenping Yang uri: https://doi.org/10.7717/peerj.8853 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Xia et al. title: Transcriptomic profiling of purple broccoli reveals light-induced anthocyanin biosynthetic signaling and structural genes link: https://peerj.com/articles/8870 last-modified: 2020-05-05 description: Purple Broccoli (Brassica oleracea L. var italica) attracts growing attention as a functional food. Its purple coloration is due to high anthocyanin amounts. Light represents a critical parameter affecting anthocyanins biosynthesis. In this study, ‘Purple Broccoli’, a light-responding pigmentation cultivar, was assessed for exploring the mechanism underlying light-induced anthocyanin biosynthesis by RNA-Seq. Cyanidin, delphinidin and malvidin derivatives were detected in broccoli head samples. Shading assays and RNA-seq analysis identified the flower head as more critical organ compared with leaves. Anthocyanin levels were assessed at 0, 7 and 11 days, respectively, with further valuation by RNA-seq under head-shading and light conditions. RNA sequences were de novo assembled into 50,329 unigenes, of which 38,701 were annotated against four public protein databases. Cluster analysis demonstrated that anthocyanin/phenylpropanoid biosynthesis, photosynthesis, and flavonoid biosynthesis in cluster 8 were the main metabolic pathways regulated by light and had showed associations with flower head growth. A total of 2,400 unigenes showed differential expression between the light and head-shading groups in cluster 8, including 650 co-expressed, 373 specifically expressed under shading conditions and 1,377 specifically expressed under normal light. Digital gene expression (DGE) analysis demonstrated that light perception and the signal transducers CRY3 and HY5 may control anthocyanin accumulation. Following shading, 15 structural genes involved in anthocyanin biosynthesis were downregulated, including PAL, C4H, 4CL, CHS, CHI, F3H and DFR. Moreover, six BoMYB genes (BoMYB6-1, BoMYB6-2, BoMYB6-3, BoMYB6-4, BoMYBL2-1 and BoMYBL2-2) and three BobHLH genes (BoTT8_5-1, BoTT8_5-2 and BoEGL5-3) were critical transcription factors controlling anthocyanin accumulation under light conditions. Based on these data, a light-associated anthocyanin biosynthesis pathway in Broccoli was proposed. This information could help improve broccoli properties, providing novel insights into the molecular mechanisms underpinning light-associated anthocyanin production in purple vegetables. creator: Chunqing Liu creator: Xueqin Yao creator: Guangqing Li creator: Lei Huang creator: Zhujie Xie uri: https://doi.org/10.7717/peerj.8870 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liu et al. title: Cloning, expression and characterization of a chitinase from Paenibacillus chitinolyticus strain UMBR 0002 link: https://peerj.com/articles/8964 last-modified: 2020-05-05 description: BackgroundChitinases are enzymes which degrade β-1,4-glycosidid linkages in chitin. The enzymatic degradation of shellfish waste (containing chitin) to chitooligosaccharides is used in industrial applications to generate high-value-added products from such waste. However, chitinases are currently produced with low efficiency and poor tolerance, limiting the industrial utility. Therefore, identifying chitinases with higher enzymatic activity and tolerance is of great importance.MethodsPrimers were designed using the genomic database of Paenibacillus chitinolyticus NBRC 15660. An exochitinase (CHI) was cloned into the recombinant plasmid pET-22b (+) to form pET-22b (+)-CHI, which was transformed into Escherichia coli TOP10 to construct a genomic library. Transformation was confirmed by colony-polymerase chain reaction and electrophoresis. The target sequence was verified by sequencing. Recombinant pET-22b (+)-CHI was transformed into E. coli Rosetta-gami B (DE3) for expression of chitinase. Recombinant protein was purified by Ni-NTA affinity chromatography and enzymatic analysis was carried out.ResultsThe exochitinase CHI from P. chitinolyticus strain UMBR 0002 was successfully cloned and heterologously expressed in E. coli Rosetta-gami B (DE3). Purification yielded a 13.36-fold enrichment and recovery yield of 72.20%. The purified enzyme had a specific activity of 750.64 mU mg−1. The optimum pH and temperature for degradation of colloidal chitin were 5.0 and 45 °C, respectively. The enzyme showed high stability, retaining >70% activity at pH 4.0–10.0 and 25–45 °C (maximum of 90 min). The activity of CHI strongly increased with the addition of Ca2+, Mn2+, Tween 80 and urea. Conversely, Cu2+, Fe3+, acetic acid, isoamyl alcohol, sodium dodecyl sulfate and β-mercaptoethanol significantly inhibited enzyme activity. The oligosaccharides produced by CHI from colloidal chitin exhibited a degree of polymerization, forming N-acetylglucosamine (GlcNAc) and (GlcNAc)2 as products.ConclusionsThis is the first report of the cloning, heterologous expression and purification of a chitinase from P. chitinolyticus strain UMBR 0002. The results highlight CHI as a good candidate enzyme for green degradation of chitinous waste. creator: Cong Liu creator: Naikun Shen creator: Jiafa Wu creator: Mingguo Jiang creator: Songbiao Shi creator: Jinzi Wang creator: Yanye Wei creator: Lifang Yang uri: https://doi.org/10.7717/peerj.8964 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Liu et al. title: A long-term study on zooplankton in two contrasting cascade reservoirs (Iguaçu River, Brazil): effects of inter-annual, seasonal, and environmental factors link: https://peerj.com/articles/8979 last-modified: 2020-05-05 description: BackgroundIn reservoirs, zooplankton strongly interact with the physical and chemical characteristics of water, and this interaction is mainly influenced by climate variation and the different methods used to manage the dam water level. Therefore, the aim of this study was to evaluate how the distinct operating modes of two cascade reservoirs affected the richness, abundance, and composition of zooplankton, both spatially (intra and inter-reservoirs) and temporally (annual and seasonal). In this study, the upstream reservoir (Salto Santiago) operates using the storage method, with a water retention time (WRT) of 51 days, whereas the downstream reservoir (Salto Osório) operates using the run-of-river method, with a WRT of 16 days.MethodsZooplankton samples were collected for 16 consecutive years from the two reservoirs located on the Iguaçu River, Brazil. A total of 720 samples were collected. Four-way ANOVAs were used to determine the differences in richness and abundance of the zooplankton among years, periods, reservoirs, and environments. Multidimensional non-metric scaling (NMDS) and an analysis of similarities (ANOSIM) were used to describe similarity patterns in species composition. Finally, a canonical correspondence analysis (CCA) was used to select the environmental predictors that best explained the variation in zooplankton abundance data.ResultsWe identified a total of 115 taxa in this study, and rotifers were the richest group. In contrast, the copepods were the most abundant. The four-way ANOVA results showed significant differences in the species richness and abundance of the zooplankton among years, periods, reservoirs, and environments. The NMDS ordination and ANOSIM test indicated that the largest differences in zooplankton species composition were annual and seasonal differences. Finally, the CCA showed that these differences were mainly associated with changes in water transparency, temperature, and the chlorophyll a, phosphorus, and total dissolved solids concentrations.DiscussionInter-annual changes in zooplankton species composition showed that over time, large filters-feeders (e.g., large daphinids and calanoid copepods) were replaced by small cladocerans (e.g., bosminids) and generalist rotifers. The highest species richness was associated with the fluvial environment, whereas the highest abundance was associated with the transitional and lacustrine reservoir environments. Variations in water temperature, nutrients, and food availability explained the annual and seasonal changes in community structure, whereas variations in the water flow characteristics of the environments explained the longitudinal changes in the richness and abundance of zooplankton in reservoirs. The differences in zooplankton structure between the two reservoirs can be explained by the functional differences between the two systems, such as their WRTs and morphometrics. creator: Pablo H.S. Picapedra creator: Cleomar Fernandes creator: Juliana Taborda creator: Gilmar Baumgartner creator: Paulo V. Sanches uri: https://doi.org/10.7717/peerj.8979 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Picapedra et al. title: Rebuilding soil organic C stocks in degraded grassland by grazing exclusion: a linked decline in soil inorganic C link: https://peerj.com/articles/8986 last-modified: 2020-05-05 description: BackgroundOur study evaluated how soil organic carbon (SOC) and soil inorganic carbon (SIC) recovered over time in deep loessial soil as overgrazed grassland was fenced and restored.MethodsThe study was conducted in the Yunwu Mountain Nature Reserve in the Ningxia Autonomous Region of China. In it we compared soil data from three grazed grassland (G) sites, three sites that were fenced for 15 years (F15), and three sites that were fenced for 30 years (F30) as a so-called ‘space for time series’.Results and DiscussionWe compared SOC accumulation in soil up to 200 cm below the surface in G, F15, and F30 plots. An increase in SOC correlated with a decrease in soil pH, and decreased soil bulk density. However, SOC sequestration in fenced plots was largely offset by a decrease in SIC, which was closely correlated (r = 0.713, p = 0.001) with SOC-driven soil pH decline. We observed no significant increase in soil total carbon in the F15 or F30 sites after comparing them to G.ConclusionsOur data indicate that fencing causes the slow diffusion processes to intensify the soil property changes from increased litter return, and this slow diffusion process is still active 30 years after fencing at 100–200 cm soil depths in the studied deep loessial soil. These findings are likely applicable to similar sites. creator: Yi Zhang creator: Yingzhong Xie creator: Hongbin Ma creator: Le Jing creator: Cory Matthew creator: Jianping Li uri: https://doi.org/10.7717/peerj.8986 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Zhang et al. title: Role of Glycine max in improving drought tolerance in Zanthoxylum bungeanum link: https://peerj.com/articles/9040 last-modified: 2020-05-05 description: Intercropping may improve community stability and yield under climate change. Here, we set up a field experiment to evaluate the advantages of cultivating Z anthoxylum bungeanum with Capsicum annum, and Z. bungeanum with Glycine max as intercrops, compared with cultivating Z. bungeanum in monoculture. Effects of extreme drought stress conditions on morphological, physiological, and biochemical traits of the three crop species cultivated in the three contrasting planting systems were compared. Results showed that extreme drought conditions induced negative impacts on Z. bungeanum grown in monoculture, due to reduced growth and metabolic impairment. However, limited stomatal conductance, reduced transpiration rate (Tr), and increased water use efficiency, carotenoid content, catalase activity, and accumulation of soluble sugars in Z. bungeanum indicated its adaptive strategies for tolerance of extreme drought stress conditions. Compared with cultivation in monoculture, intercropping with C. annum had positive effects on Z. bungeanum under extreme drought stress conditions, as a result of improved crown diameter, leaf relative water content (LRWC), net photosynthetic rate, and proline content, while intercropping with G. max under extreme drought stress conditions increased net CO2 assimilation rates, LRWC, Tr, and superoxide dismutase (SOD) activity. In conclusion, Z. bungeanum has an effective defense mechanism for extreme drought stress tolerance. Intercropping with G. max enhanced this tolerance potential primarily through its physio-biochemical adjustments, rather than as a result of nitrogen fixation by G. max. creator: Zilong Li creator: Akash Tariq creator: Kaiwen Pan creator: Corina Graciano creator: Feng Sun creator: Dagang Song creator: Olusanya Abiodun Olatunji uri: https://doi.org/10.7717/peerj.9040 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Li et al. title: Linking diffuse radiation and ecosystem productivity of a desert steppe ecosystem link: https://peerj.com/articles/9043 last-modified: 2020-05-05 description: Radiation components have distinct effects on photosynthesis. In the desert steppe ecosystem, the influence of diffuse radiation on carbon fixation has not been thoroughly explored. We examined this diffusion and its effect on ecosystem productivity was examined during the growing season from 2014 to 2015 on the basis of eddy covariance measurements of CO2 exchange in a desert steppe ecosystem in northwest China. Our results indicated that the gross ecosystem production (GEP) and diffuse photosynthetically active radiation (PARdif) peaked when the clearness index (CI) was around 0.5. The maximum canopy photosynthesis (Pmax) under cloudy skies (CI < 0.7) was 23.7% greater than under clear skies (CI ≥ 0.7). When the skies became cloudy in the desert steppe ecosystem, PARdif had a greater effect on GEP. Additionally, lower vapor pressure deficits (VPD ≤ 1 kPa), lower air temperatures (Ta ≤ 20 °C), and non-stressed water conditions (REW ≥ 0.4) were more conducive for enhanced ecosystem photosynthesis under cloudy skies than under clear skies. This may be due to the comprehensive effects of VPD and Ta on stomatal conductance. We concluded that cloudiness can influence diffuse radiation components and that diffuse radiation can increase the ecosystem production of desert steppe ecosystems in northwest China. creator: Cheng Li creator: Xin Jia creator: Jingyong Ma creator: Peng Liu creator: Ruizhi Yang creator: Yujie Bai creator: Muhammad Hayat creator: Jinglan Liu creator: Tianshan Zha uri: https://doi.org/10.7717/peerj.9043 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Li et al. title: Genome-wide analysis of the superoxide dismutase (SOD) gene family in Zostera marina and expression profile analysis under temperature stress link: https://peerj.com/articles/9063 last-modified: 2020-05-05 description: Superoxide dismutases (SODs) serve as the first line of defense in the plant antioxidant enzyme system, and play a primary role in the removal of reactive oxygen species (ROS). However, our understanding of the functions of the SOD family in Zostera marina is limited. In this study, a systematic analysis was conducted on the characteristics of the SOD genes in Z. marina at the whole-genome level. Five SOD genes were identified, consisting of two Cu/ZnSODs, two FeSODs, and one MnSOD. Phylogenetic analysis showed that ZmSOD proteins could be divided into two major categories (Cu/ZnSODs and Fe-MnSODs). Sequence motifs, gene structure, and the 3D-modeled protein structures further supported the phylogenetic analysis, with each subgroup having similar motifs, exon-intron structures, and protein structures. Additionally, several cis-elements were identified that may respond to biotic and abiotic stresses. Transcriptome analysis revealed expression diversity of ZmSODs in various tissues. Moreover, qRT-PCR analysis showed that the expression level of most ZmSOD genes trended to decreased expression with the increase of temperature, indicating that heat stress inhibits expression of ZmSODs and may result in reduced ability of ZmSODs to scavenge ROS. Our results provide a basis for further functional research on the SOD gene family in Z. marina, which will help to determine the molecular mechanism of ZmSOD genes in response to environmental stress. creator: Yu Zang creator: Jun Chen creator: Ruoxi Li creator: Shuai Shang creator: Xuexi Tang uri: https://doi.org/10.7717/peerj.9063 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Zang et al. title: Comparison of defense responses of transgenic potato lines expressing three different Rpi genes to specific Phytophthora infestans races based on transcriptome profiling link: https://peerj.com/articles/9096 last-modified: 2020-05-05 description: Potato late blight, one of the most devastating diseases in potato, is caused by the oomycete Phytophthora infestans. Over 20 resistance genes have been cloned including R1, R3a, and R3b. The distinctions between defense response mechanisms mediated by different resistance genes are still unclear. Here we performed transcriptome profiling in three transgenic lines, R1, R3a, and R3b, and wild-type Desiree under inoculation with two P. infestans isolates, 89148 (race 0) and CN152 (super race), using RNA-seq. Compared with wild type, specific differentially expressed genes (DEGs) were identified in the three transgenic lines. The highest number of DEGs occurred in transgenic R3b, with 779 DEGs in response to isolate 89148 and 864 DEGs in response to infection by CN152, followed by transgenic R1 lines with 408 DEGs for isolate 89148 and 267 DEGs for CN152. Based on gene ontology, the most common GO terms (15 for 89148 and 20 for CN152) were enriched in transgenic R3a and R3b lines. This indicates that the defense pathways mediated by R3a and R3b are more similar than those mediated by R1. Further separate GO analysis of up- or down-regulated DEGs showed that the down-regulated DEGs mainly functioned in mediating the resistance of potato to P. infestans 89148 by response to stress biological process and to CN152 by oxidation reduction biological process. KEGG pathways of DNA replication, plant-pathogen interaction and pentose and glucuronate interconversions are unique for transgenic R1, R3a, and R3b lines in incompatible interactions. Quantitative real-time PCR experimental validation confirmed the induced expression of DEGs in the late blight resistance signaling pathway. Our results will lay a solid foundation for further understanding the mechanisms of plant-pathogen interactions, and provide a theoretical reference for durable resistance in potato. creator: Xiaohui Yang creator: Xiao Guo creator: Guangxia Chen creator: Daofeng Dong creator: Fang Liu creator: Yuanjun Yang creator: Yu Yang creator: Guangcun Li uri: https://doi.org/10.7717/peerj.9096 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Yang et al. title: Genetic diversity of Alternaria species associated with black point in wheat grains link: https://peerj.com/articles/9097 last-modified: 2020-05-05 description: The genus Alternaria is a widely distributed major plant pathogen that can act as a saprophyte in plant debris. Fungi of this genus frequently infect cereal crops and cause such diseases as black point and wheat leaf blight, which decrease the yield and quality of cereal products. A total of 25 Alternaria sp. isolates were collected from germ grains of various wheat cultivars from different geographic regions in Kazakhstan. We investigated the genetic relationships of the main Alternaria species related to black point disease of wheat in Kazakhstan, using the inter-primer binding site (iPBS) DNA profiling technique. We used 25 retrotransposon-based iPBS primers to identify the differences among and within Alternaria species populations, and analyzed the variation using clustering (UPGMA) and statistical approaches (AMOVA). Isolates of Alternaria species clustered into two main genetic groups, with species of A.alternata and A.tennuissima forming one cluster, and isolates of A. infectoria forming another. The genetic diversity found using retrotransposon profiles was strongly correlated with geographic data. Overall, the iPBS fingerprinting technique is highly informative and useful for the evaluation of genetic diversity and relationships of Alternaria species. creator: Ainur Turzhanova creator: Oxana N. Khapilina creator: Asem Tumenbayeva creator: Vladislav Shevtsov creator: Olesya Raiser creator: Ruslan Kalendar uri: https://doi.org/10.7717/peerj.9097 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Turzhanova et al. title: Conditioned medium from M2b macrophages modulates the proliferation, migration, and apoptosis of pulmonary artery smooth muscle cells by deregulating the PI3K/Akt/FoxO3a pathway link: https://peerj.com/articles/9110 last-modified: 2020-05-05 description: BackgroundImmunity and inflammation are considered to be central features of pulmonary artery hypertension (PAH), in which macrophages are one of the main components of inflammatory cell infiltration around the pulmonary artery. M2b macrophages, which are different from M1 and M2 macrophages, are believed to have immunomodulatory activities and produce little fibrosis. The purpose of this study was to explore the effect of M2b macrophages on pulmonary artery smooth muscle cells (PASMCs) derived from monocrotaline-induced PAH rats.MethodsPASMCs were cultured in serum-free medium, the supernatant of M0 macrophages, or the supernatant of M2b macrophages for 24 hours. Then cell proliferation was assessed by cell counting kit-8 and cell migration ability was detected by wound healing and transwell assays. The apoptosis rate of cells was determined by TUNEL staining and annexin V-PE/7-ADD staining. Western blot was used to detect the expression of Bcl-2 family proteins, cleaved caspase-9 and PI3K/Akt/FoxO3a pathway. LY294002 (a specific inhibitor of PI3K) was used to investigate its effect on PASMCs and its relationship with M2b macrophages.ResultsConditioned medium from M2b macrophages significantly inhibited the proliferation and migration of PASMCs compared with the control group and M0 macrophage group. Furthermore, conditioned medium from M2b macrophages promote PASMC apoptosis and increased the expression of pro-apoptotic proteins Bax and cleaved caspase-9, inhibited the expression of anti-apoptotic proteins Bcl-2 and Bcl-xl. Finally, conditioned medium from M2b macrophages inhibited the PI3K/Akt/FoxO3a pathway. Inhibition of PI3K/Akt/FoxO3a pathway also significantly inhibit the proliferation, migration, and apoptosis resistance of PASMCs.ConclusionConditioned medium from M2b macrophages can inhibit the proliferation, migration, and apoptosis resistance of PASMCs, which may be at least partially by deregulating the PI3K/Akt/FoxO3a pathway. creator: Suiqing Huang creator: Yuan Yue creator: Kangni Feng creator: Xiaolin Huang creator: Huayang Li creator: Jian Hou creator: Song Yang creator: Shaojie Huang creator: Mengya Liang creator: Guangxian Chen creator: Zhongkai Wu uri: https://doi.org/10.7717/peerj.9110 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Huang et al. title: Mucoactive agent use in adult UK Critical Care Units: a survey of health care professionals’ perception, pharmacists’ description of practice, and point prevalence of mucoactive use in invasively mechanically ventilated patients link: https://peerj.com/articles/8828 last-modified: 2020-05-04 description: BackgroundMechanical ventilation for acute respiratory failure is one of the most common indications for admission to intensive care units (ICUs). Airway mucus clearance is impaired in these patients medication, impaired mucociliary motility, increased mucus production etc. and mucoactive agents have the potential to improve outcomes. However, studies to date have provided inconclusive results. Despite this uncertainty, mucoactives are used in adult ICUs, although the extent of use and perceptions about place in therapy are not known.Aims and ObjectivesWe aim to describe the use of mucoactive agents in mechanically ventilated patients in UK adult critical care units. Specifically, our objectives are to describe clinicians perceptions about the use of mucoactive agents, understand the indications and anticipated benefits, and describe the prevalence and type of mucoactive agents in use.MethodsWe conducted three surveys. Firstly, a practitioner-level survey aimed at nurses, physiotherapists and doctors to elucidate individual practitioners perceptions about the use of mucoactive agents. Secondly, a critical care unit-level survey aimed at pharmacists to understand how these perceptions translate into practice. Thirdly, a point prevalence survey to describe the extent of prescribing and range of products in use. The practitioner-level survey was disseminated through the UK Intensive Care Society for completion by a multi-professional membership. The unit-level and point prevalence surveys were disseminated cthrough the UK Clinical Pharmacy Association for completion by pharmacists.ResultsThe individual practitioners survey ranked ‘thick secretions’ as the main reason for commencing mucoactive agents determined using clinical assessment. The highest ranked perceived benefit for patient centred outcomes was the duration of ventilation. Of these respondents, 79% stated that further research was important and 87% expressed support for a clinical trial. The unit-level survey found that mucoactive agents were used in 83% of units. The most highly ranked indication was again ‘thick secretions’ and the most highly ranked expected patient centred clinical benefit being improved gas exchange and reduced ventilation time. Only five critical care units provided guidelines to direct the use of mucoactive agents (4%). In the point prevalence survey, 411/993 (41%) of mechanically ventilated patients received at least one mucoactive agent. The most commonly administered mucoactives were inhaled sodium chloride 0.9% (235/993, 24%), systemic carbocisteine (161/993, 16%) and inhaled hypertonic sodium cloride (127/993, 13%).ConclusionsMucoactive agents are used extensively in mechanically ventilated adult patients in UK ICUs to manage ‘thick secretions’, with a key aim to reduce the duration of ventilation. There is widespread support for clinical trials to determine the optimal use of mucoactive agent therapy in this patient population. creator: Mark Borthwick creator: Danny McAuley creator: John Warburton creator: Rohan Anand creator: Judy Bradley creator: Bronwen Connolly creator: Bronagh Blackwood creator: Brenda O’Neill creator: Marc Chikhani creator: Paul Dark creator: Murali Shyamsundar uri: https://doi.org/10.7717/peerj.8828 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Borthwick et al. title: The impact of sleep loss on sustained and transient attention: an EEG study link: https://peerj.com/articles/8960 last-modified: 2020-05-04 description: Sleep is one of our most important physiological functions that maintains physical and mental health. Two studies examined whether discrete areas of attention are equally affected by sleep loss. This was achieved using a repeated-measures within-subjects design, with two contrasting conditions: normal sleep and partial sleep restriction of 5-h. Study 1 compared performance on a sustained attention task (Psychomotor Vigilance task; PVT) with performance on a transient attention task (Attentional Blink; AB). PVT performance, but not performance on the AB task, was impaired after sleep restriction. Study 2 sought to determine the neural underpinnings of the phenomenon, using electroencephalogram (EEG) frequency analysis, which measured activity during the brief eyes-closed resting state before the tasks. AB performance was unaffected by sleep restriction, despite clearly observable changes in brain activity. EEG results showed a significant reduction in resting state alpha oscillations that was most prominent centrally in the right hemisphere. Changes in individual alpha and delta power were also found to be related to changes in subjective sleepiness and PVT performance. Results likely reflect different levels of impairment in specific forms of attention following sleep loss. creator: Lucienne Shenfield creator: Vanessa Beanland creator: Ashleigh Filtness creator: Deborah Apthorp uri: https://doi.org/10.7717/peerj.8960 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Shenfield et al. title: Origin identification of migratory pests (European Starling) using geochemical fingerprinting link: https://peerj.com/articles/8962 last-modified: 2020-05-04 description: The European Starling (Sturnidae: Sturnus vulgaris L.) is an invasive bird in North America where it is an agricultural pest. In British Columbia (Canada), the starling population increases in orchards and vineyards in autumn, where they consume and damage ripening fruits. Starlings also cause damage in dairy farms and feedlots by consuming and contaminating food and spreading diseases. Damage can be partly mitigated by the use of scare devices, which can disperse flocks until they become habituated. Large-scale trapping and euthanizing before starlings move to fields and farms could be a practical means of preventing damage, but requires knowledge of natal origin. Within a small (20,831 km2), agriculturally significant portion of south-central British Columbia, the Okanagan-Similkameen region, we used 21 trace elements in bone tissue to discriminate the spatial distribution of juvenile starlings and to reveal the geographic origin of the problem birds in fall. Stepwise discriminant analysis of trace elements classified juveniles to their natal origin (minimum discrimination distance of 12 km) with 79% accuracy. In vineyards and orchards, the majority (55%) of problem birds derive from northern portions of the valley; and the remaining 45% of problem birds were a mixture of local and immigrant/unassigned birds. In contrast, problem birds in dairy farms and feedlots were largely immigrants/unassigned (89%) and 11% were local from northern region of the valley. Moreover, elemental signatures can separate starling populations in the Valley yielding a promising tool for identifying the geographic origin of these migratory birds. creator: Upama Khatri-Chhetri creator: John G. Woods creator: Ian R. Walker creator: P. Jeff Curtis uri: https://doi.org/10.7717/peerj.8962 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Khatri-Chhetri et al. title: Cryptic diversity among Yazoo Darters (Percidae: Etheostoma raneyi) in disjunct watersheds of northern Mississippi link: https://peerj.com/articles/9014 last-modified: 2020-05-04 description: The Yazoo Darter, Etheostoma raneyi (Percidae), is an imperiled freshwater fish species endemic to tributaries of the Yocona and Little Tallahatchie rivers of the upper Yazoo River basin, in northern Mississippi, USA. The two populations are allopatric, isolated by unsuitable lowland habitat between the two river drainages. Relevant literature suggests that populations in the Yocona River represent an undescribed species, but a lack of data prevents a thorough evaluation of possible diversity throughout the range of the species. Our goals were to estimate phylogenetic relationships of the Yazoo Darter across its distribution and identify cryptic diversity for conservation management purposes. Maximum likelihood (ML) phylogenetic analyses of the mitochondrial cytochrome b (cytb) gene returned two reciprocally monophyletic clades representing the two river drainages with high support. Bayesian analysis of cytb was consistent with the ML analysis but with low support for the Yocona River clade. Analyses of the nuclear S7 gene yielded unresolved relationships among individuals in the Little Tallahatchie River drainage with mostly low support, but returned a monophyletic clade for individuals from the Yocona River drainage with high support. No haplotypes were shared between the drainages for either gene. Additional cryptic diversity within the two drainages was not indicated. Estimated divergence between Yazoo Darters in the two drainages occurred during the Pleistocene (<1 million years ago) and was likely linked to repeated spatial shifts in suitable habitat and changes in watershed configurations during glacial cycles. Individuals from the Yocona River drainage had lower genetic diversity consistent with the literature. Our results indicate that Yazoo Darters in the Yocona River drainage are genetically distinct and that there is support for recognizing Yazoo Darter populations in the Yocona River drainage as a new species under the unified species concept. creator: Ken A. Sterling creator: Stuart V. Nielsen creator: Andrew J. Brown creator: Melvin L. Warren, Jr. creator: Brice P. Noonan uri: https://doi.org/10.7717/peerj.9014 license: https://creativecommons.org/publicdomain/zero/1.0/ rights: ©2020 Sterling et al. title: Fauna associated with shallow-water methane seeps in the Laptev Sea link: https://peerj.com/articles/9018 last-modified: 2020-05-04 description: BackgroundMethane seeps support unique benthic ecosystems in the deep sea existing due to chemosynthetic organic matter. In contrast, in shallow waters there is little or no effect of methane seeps on macrofauna. In the present study we focused on the recently described methane discharge area at the northern Laptev Sea shelf. The aim of this work was to describe the shallow-water methane seep macrofauna and to understand whether there are differences in macrobenthic community structure between the methane seep and background areas.MethodsSamples of macrofauna were taken during three expeditions of RV Akademik Mstislav Keldysh in 2015, 2017 and 2018 using 0.1 m2 grabs and the Sigsbee trawl. 21 grabs and two trawls in total were taken at two methane seep sites named Oden and C15, located at depths of 60–70 m. For control, three 0.1 m2 grabs were taken in area without methane seepage.ResultsThe abundance of macrofauna was higher at methane seep stations compared to non-seep sites. Cluster analysis revealed five station groups corresponding to control area, Oden site and C15 site (the latter represented by three groups). Taxa responsible for differences among the station groups were mostly widespread Arctic species that were more abundant in samples from methane seep sites. However, high densities of symbiotrophic siboglinids Oligobrachia sp. were found exclusively at methane seep stations. In addition, several species possibly new to science were found at several methane seep stations, including the gastropod Frigidalvania sp. and the polychaete Ophryotrocha sp. The fauna at control stations was represented only by well-known and widespread Arctic taxa. Higher habitat heterogeneity of the C15 site compared to Oden was indicated by the higher number of station groups revealed by cluster analysis and higher species richness in C15 trawl sample. The development of the described communities at the shallow-water methane seeps can be related to pronounced oligotrophic environment on the northern Siberian shelf. creator: Andrey A. Vedenin creator: Valentin N. Kokarev creator: Margarita V. Chikina creator: Alexander B. Basin creator: Sergey V. Galkin creator: Andrey V. Gebruk uri: https://doi.org/10.7717/peerj.9018 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Vedenin et al. title: Public priorities on locally-driven sea level rise planning on the East Coast of the United States link: https://peerj.com/articles/9044 last-modified: 2020-05-04 description: Sea level rise poses a substantial concern to communities worldwide. Increased inundation, storm surge, saltwater intrusion, and other impacts create challenges which will require considerable planning to address. Recognizing the broad and differing scope of sea level rise issues and the variability of policy options to address them, local planning frameworks are necessary in addition to tools and resources available from state and federal governments. To help assess priorities and preferences on sea level rise planning, a survey of 503 persons affiliated with coastal communities on the East Coast of the United States was conducted in December 2017. This survey studied key aspects locally-driven sea level rise plans, including planning priorities, funding options, methods to resolve conflict, and potential responses. Six key findings address these and other concerns to provide the foundation of a locally driven framework for public officials. creator: Adam T. Carpenter uri: https://doi.org/10.7717/peerj.9044 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Carpenter title: Intra-lake response of Arcellinida (testate lobose amoebae) to gold mining-derived arsenic contamination in northern Canada: Implications for environmental monitoring link: https://peerj.com/articles/9054 last-modified: 2020-05-04 description: Arcellinida (testate lobose amoebae) were examined from 40 near-surface sediment samples (top 0.5 cm) from two lakes impacted by arsenic (As) contamination associated with legacy gold mining in subarctic Canada. The objectives of the study are two folds: quantify the response of Arcellinida to intra-lake variability of As and other physicochemical controls, and evaluate whether the impact of As contamination derived from two former gold mines, Giant Mine (1938–2004) and Tundra Mine (1964–1968 and 1983–1986), on the Arcellinida distribution in both lakes is comparable or different. Cluster analysis and nonmetric multidimensional scaling (NMDS) were used to identify Arcellinida assemblages in both lakes, and redundancy analysis (RDA) was used to quantify the relationship between the assemblages, As, and other geochemical and sedimentological parameters. Cluster analysis and NMDS revealed four distinct arcellinidan assemblages in Frame Lake (assemblages 1–4) and two in Hambone Lake (assemblages 5 and 6): (1) Extreme As Contamination (EAC) Assemblage; (2) High calcium (HC) Assemblage; (3) Moderate As Contamination (MAC) assemblages; (4) High Nutrients (HN) Assemblage; (5) High Diversity (HD) Assemblage; and (6) Centropyxis aculeata (CA) Assemblage. RDA analysis showed that the faunal structure of the Frame Lake assemblages was controlled by five variables that explained 43.2% of the total faunal variance, with As (15.8%), Olsen phosphorous (Olsen-P; 10.5%), and Ca (9.5%) being the most statistically significant (p < 0.004). Stress-tolerant arcellinidan taxa were associated with elevated As concentrations (e.g., EAC and MAC; As concentrations range = 145.1–1336.6 mg kg−1; n = 11 samples), while stress-sensitive taxa thrived in relatively healthier assemblages found in substrates with lower As concentrations and higher concentrations of nutrients, such as Olsen-P and Ca (e.g., HC and HM; As concentrations range = 151.1–492.3 mg kg−1; n = 14 samples). In contrast, the impact of As on the arcellinidan distribution was not statistically significant in Hambone Lake (7.6%; p-value = 0.152), where the proportion of silt (24.4%; p-value = 0.005) and loss-on-ignition-determined minerogenic content (18.5%; p-value = 0.021) explained a higher proportion of the total faunal variance (58.4%). However, a notable decrease in arcellinidan species richness and abundance and increase in the proportions of stress-tolerant fauna near Hambone Lake’s outlet (e.g., CA samples) is consistent with a spatial gradient of higher sedimentary As concentration near the outlet, and suggests a lasting, albeit weak, As influence on Arcellinida distribution in the lake. We interpret differences in the influence of sedimentary As concentration on Arcellinida to differences in the predominant As mineralogy in each lake, which is in turn influenced by differences in ore-processing at the former Giant (roasting) and Tundra mines (free-milling). creator: Nawaf A. Nasser creator: R. Timothy Patterson creator: Jennifer M. Galloway creator: Hendrik Falck uri: https://doi.org/10.7717/peerj.9054 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Nasser et al. title: Transcriptional profiling to identify the key genes and pathways of pterygium link: https://peerj.com/articles/9056 last-modified: 2020-05-04 description: PurposePterygium results from a variety of biological pathways that are involved in the formation of ocular surface diseases. However, the exact pathogenesis of pterygium is still unclear. Our study focused on gene expression profiles to better understand the potential mechanisms of pterygium.MethodsRNA sequencing experiments were performed on clinical pterygium tissues and normal conjunctival tissues. To identify the hub genes for the development of pterygium, we further conducted weighted gene co-expression network analysis (WGCNA). qRT-PCR was utilized to validate the dysregulation of the most significant differentially expressed genes (DEGs) and key hub genes in the independent subjects.ResultsA total of 339 DEGs (P-adjusted < 0.05 and log2 fold change [log2FC] ≥ 1.0) were obtained that reached statistical significance with p-values < 0.05. Among them, 200 DEGs were upregulated; these genes were mainly associated with the extracellular matrix and with cell adhesion or migration. In contrast, the 139 downregulated genes were enriched for endocrine and inflammation pathways. With regard to WGCNA, five modules were assigned based on the DEG profiles, and the biological functions of each module were verified with previously published GO terms. The functions included ECM-receptor interactions, the PI3K-Akt signalling pathway and an endoplasmic reticulum (ER)-related pathway. The five hub genes with the highest connectivity in each module and the five most significant DEGs showed dysregulated expression in the independent cohort samples.ConclusionsRNA sequencing and WGCNA provided novel insights into the potential regulatory mechanisms of pterygium. The identified DEGs and hub genes, which were classified into two groups according to different functions or signalings, may provide important references for further research on the molecular biology of pterygium. creator: Yihui Chen creator: Haoyu Wang creator: Yaping Jiang creator: Xiaoyan Zhang creator: Qingzhong Wang uri: https://doi.org/10.7717/peerj.9056 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Chen et al. title: Phosphate solubilizing bacteria stimulate wheat rhizosphere and endosphere biological nitrogen fixation by improving phosphorus content link: https://peerj.com/articles/9062 last-modified: 2020-05-04 description: Phosphate (P) availability often limits biological nitrogen fixation (BNF) by diazotrophic bacteria. In soil, only 0.1% of the total P is available for plant uptake. P solubilizing bacteria can convert insoluble P to plant-available soluble P (ionic P and low molecular-weight organic P). However, limited information is available about the effects of synergistic application of diazotrophic bacteria and P solubilizing bacteria on the nitrogenase activity of rhizosphere and nifH expression of endosphere. In this study, we investigated the effects of co-inoculation with a diazotrophic bacterium (Paenibacillus beijingensis BJ-18) and a P-solubilizing bacterium (Paenibacillus sp. B1) on wheat growth, plant and soil total N, plant total P, soil available P, soil nitrogenase activity and the relative expression of nifH in plant tissues. Co-inoculation significantly increased plant biomass (length, fresh and dry weight) and plant N content (root: 27%, shoot: 30%) and P content (root: 63%, shoot: 30%). Co-inoculation also significantly increased soil total N (12%), available P (9%) and nitrogenase activity (69%) compared to P. beijingensis BJ-18 inoculation alone. Quantitative real-time PCR analysis showed co-inoculation doubled expression of nifH genes in shoots and roots. Soil nitrogenase activity and nifH expression within plant tissues correlated with P content of soil and plant tissues, which suggests solubilization of P by Paenibacillus sp. B1 increased N fixation in soils and the endosphere. In conclusion, P solubilizing bacteria generally improved soil available P and plant P uptake, and considerably stimulated BNF in the rhizosphere and endosphere of wheat seedlings. creator: Yongbin Li creator: Qin Li creator: Guohua Guan creator: Sanfeng Chen uri: https://doi.org/10.7717/peerj.9062 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Li et al. title: On the importance of spatial scales on beta diversity of coral assemblages: a study from Venezuelan coral reefs link: https://peerj.com/articles/9082 last-modified: 2020-05-04 description: Estimating variability across spatial scales has been a major issue in ecology because the description of patterns in space is extremely valuable to propose specific hypotheses to unveil key processes behind these patterns. This paper aims to estimate the variability of the coral assemblage structure at different spatial scales in order to determine which scales explain the largest variability on β-diversity. For this, a fully-nested design including a series of hierarchical-random factors encompassing three spatial scales: (1) regions, (2) localities and (3) reefs sites across the Venezuelan territory. The variability among spatial scales was tested with a permutation-based analysis of variance (Permanova) based on Bray-Curtis index. Dispersion in species presence/absence across scales (i.e., β-diversity) was tested with a PermDisp analysis based on Jaccard’s index. We found the highest variability in the coral assemblage structure between sites within localities (Pseudo-F = 5.34; p-value = 0.001, CV = 35.10%). We also found that longitude (Canonical corr = 0.867, p = 0.001) is a better predictor of the coral assemblage structure in Venezuela, than latitude (Canonical corr = 0.552, p = 0.021). Largest changes in β-diversity of corals occurred within sites (F = 2.764, df1= 35, df2 = 107, p = 0.045) and within localities (F = 4.438, df1= 6, df2 = 29, p = 0.026). Our results suggest that processes operating at spatial scales of hundreds of meters and hundreds of kilometers might both be critical to shape coral assemblage structure in Venezuela, whereas smaller scales (i.e., hundreds of meters) showed to be highly- important for the species turnover component of β-diversity. This result highlights the importance of creating scale-adapted management actions in Venezuela and likely across the Caribbean region. creator: Emy Miyazawa creator: Luis M. Montilla creator: Esteban Alejandro Agudo-Adriani creator: Alfredo Ascanio creator: Gloria Mariño-Briceño creator: Aldo Croquer uri: https://doi.org/10.7717/peerj.9082 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Miyazawa et al. title: Fast sequence-based microsatellite genotyping development workflow link: https://peerj.com/articles/9085 last-modified: 2020-05-04 description: Application of high-throughput sequencing technologies to microsatellite genotyping (SSRseq) has been shown to remove many of the limitations of electrophoresis-based methods and to refine inference of population genetic diversity and structure. We present here a streamlined SSRseq development workflow that includes microsatellite development, multiplexed marker amplification and sequencing, and automated bioinformatics data analysis. We illustrate its application to five groups of species across phyla (fungi, plant, insect and fish) with different levels of genomic resource availability. We found that relying on previously developed microsatellite assay is not optimal and leads to a resulting low number of reliable locus being genotyped. In contrast, de novo ad hoc primer designs gives highly multiplexed microsatellite assays that can be sequenced to produce high quality genotypes for 20–40 loci. We highlight critical upfront development factors to consider for effective SSRseq setup in a wide range of situations. Sequence analysis accounting for all linked polymorphisms along the sequence quickly generates a powerful multi-allelic haplotype-based genotypic dataset, calling to new theoretical and analytical frameworks to extract more information from multi-nucleotide polymorphism marker systems. creator: Olivier Lepais creator: Emilie Chancerel creator: Christophe Boury creator: Franck Salin creator: Aurélie Manicki creator: Laura Taillebois creator: Cyril Dutech creator: Abdeldjalil Aissi creator: Cecile F.E. Bacles creator: Françoise Daverat creator: Sophie Launey creator: Erwan Guichoux uri: https://doi.org/10.7717/peerj.9085 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Lepais et al. title: Antifungal activity of 8-methoxynaphthalen-1-ol isolated from the endophytic fungus Diatrype palmicola MFLUCC 17-0313 against the plant pathogenic fungus Athelia rolfsii on tomatoes link: https://peerj.com/articles/9103 last-modified: 2020-05-04 description: Thirty-four endophytic fungal isolates were obtained from the leaves of the medicinal plant Polyscias fruticosa, and their antagonistic activities against the growth of the common tomatoes plant pathogenic fungus Athelia rolfsii were initially screened using a dual culture assay. The endophytic fungus MFLUCC 17-0313, which was later molecularly identified as Diatrype palmicola, displayed the highest inhibition percentage (49.98%) in comparison to the others. This fungus was then chosen for further evaluation. Its culture broth and mycelia from a 10 L scale were separated and extracted using ethyl acetate, methanol, and hexane. Each extract was tested for antifungal activity against the same pathogen using a disc diffusion assay. Only the crude hexane extract of fungal mycelium showed antifungal activity. The hexane extract was fractioned using sephadex gel filtration chromatography and each fraction was tested for antifungal activity until the one with the highest inhibition percentage was obtained. The bioactive compound was identified as 8-methoxynaphthalen-1-ol using nuclear magnetic resonance spectroscopy and mass spectrometry. The minimum inhibition concentration of 8-methoxynaphthalen-1-ol was demonstrated at 250 µg/mL against the selected pathogen. Using the leaf assay, the solution of 8-methoxynapthalen-1-ol was tested for phytotoxic activity against A. rolfsii and was found to have no phytotoxic effects. These results showed that 8-methoxynaphthalen-1-ol has the potential for controlling the growth of A. rolfsii, the cause of Southern blight disease on tomatoes. This study may provide the foundation for future use of this compound as a biofungicide. creator: Chutima Tanapichatsakul creator: Acharavadee Pansanit creator: Sakon Monggoot creator: Siraprapa Brooks creator: Surasak Prachya creator: Prasat Kittakoop creator: Parinya Panuwet creator: Patcharee Pripdeevech uri: https://doi.org/10.7717/peerj.9103 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Tanapichatsakul et al. title: Multimode ultrasonic technique is recommended for the differential diagnosis of thyroid cancer link: https://peerj.com/articles/9112 last-modified: 2020-05-04 description: BackgroundB-mode ultrasound is one of the most commonly used imaging techniques for evaluating thyroid nodules due to its noninvasive property and excellent performance in terms of discriminating between benign and malignant nodules. However, the accuracy of differential diagnosis strongly depends on the experience of ultrasonographers. In addition to B-mode ultrasound, the elastic mode and contrast-enhanced mode have shown complimentary value in the diagnosis of thyroid nodules. The combination of multiple modes in ultrasonic techniques may effectively undermine diagnostic subjectiveness and improve accuracy. In this study, we evaluated the diagnostic value of combining the three ultrasonic modes for differentiating thyroid cancers.MethodsIn this retrospective study, we analyzed a total of 196 thyroid nodules with suspected malignancies from 185 patients who gave informed consent. Xi’an Jiaotong University granted ethical approval (No. 2018200) to carry out the study within its facilities. All the patients received ultrasonic examinations with the B mode, elastic mode and contrast-enhanced mode, followed by histopathological confirmation by fine-need aspiration or surgery. A predictive multivariate logistic regression model was selected to integrate the variety of data obtained from the three modes.ResultsThe combination of three ultrasonic techniques for differentiating malignant from benign thyroid nodules showed the highest diagnostic accuracy of 0.985 compared to the B mode alone (0.841) and the two-mode combination. The accuracy of the B mode combined with the elastic technique was 0.954, and the accuracy of the B mode combined with the contrast-enhanced technique was 0.960.DiscussionMultimode ultrasonic techniques should be recommended to patients with suspected malignant thyroid nodules in routine clinical practice. creator: Juan Wang creator: Xin He creator: Li Ma creator: Miao Li creator: Lei Sun creator: Jue Jiang creator: Qi Zhou uri: https://doi.org/10.7717/peerj.9112 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Wang et al. title: Association between dietary mineral nutrient intake, body mass index, and waist circumference in U.S. adults using quantile regression analysis NHANES 2007–2014 link: https://peerj.com/articles/9127 last-modified: 2020-05-04 description: ObjectiveMineral nutrients play an important role in maintaining material and energy metabolism. Reports on mineral nutrient intakes and body mass index (BMI) and waist circumference (WC) are rare in the United States. This study examined the relationship between BMI, WC and dietary mineral intakes.MethodWe used the data from National Health and Nutrition Examination Survey 2007–2014. Nutrient intakes were adjusted for energy according to the residual adjustment method. We used the quantile regression model to analyze the relationship between BMI, WC under different distributions and the average daily mineral intakes.ResultA total of 19,952 people were included in the study, including 9,879 men and 10,073 women (≥20 years old). The median BMI was 27.935 kg/m2 and the median WC was 97.700 cm. The results of quantile regression showed that calcium, magnesium, potassium, copper, zinc and iron intakes were negatively correlated with BMI and WC, after adjusting for age and gender. Sodium and phosphorus intakes were positively correlated with BMI, sodium intakes were positively correlated with WC. This correlation was enhanced with increasing quantiles of risk levels. In high BMI or high WC populations, mineral intakes had a greater impact on BMI and WC. The quantile regression coefficients of selenium intakes were not statistically significant at each quantile.ConclusionOur results suggested that the mineral nutrient intakes were associated with BMI and WC in American adults. However, we also need to further study the longitudinal effects of mineral intakes and obesity. creator: Shan Jiang creator: Xiaoyu Ma creator: Meng Li creator: Shoumeng Yan creator: Hantong Zhao creator: Yingan Pan creator: Changcong Wang creator: Yan Yao creator: Lina Jin creator: Bo Li uri: https://doi.org/10.7717/peerj.9127 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Jiang et al. title: Comparison of mobile and clinical EEG sensors through resting state simultaneous data collection link: https://peerj.com/articles/8969 last-modified: 2020-05-01 description: Development of mobile sensors brings new opportunities to medical research. In particular, mobile electroencephalography (EEG) devices can be potentially used in low cost screening for epilepsy and other neurological and psychiatric disorders. The necessary condition for such applications is thoughtful validation in the specific medical context. As part of validation and quality assurance, we developed a computer-based analysis pipeline, which aims to compare the EEG signal acquired by a mobile EEG device to the one collected by a medically approved clinical-grade EEG device. Both signals are recorded simultaneously during 30 min long sessions in resting state. The data are collected from 22 patients with epileptiform abnormalities in EEG. In order to compare two multichannel EEG signals with differently placed references and electrodes, a novel data processing pipeline is proposed. It allows deriving matching pairs of time series which are suitable for similarity assessment through Pearson correlation. The average correlation of 0.64 is achieved on a test dataset, which can be considered a promising result, taking the positions shift due to the simultaneous electrode placement into account. creator: Ekaterina Kutafina creator: Alexander Brenner creator: Yannic Titgemeyer creator: Rainer Surges creator: Stephan Jonas uri: https://doi.org/10.7717/peerj.8969 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Kutafina et al. title: Enhanced genome editing in human iPSCs with CRISPR-CAS9 by co-targeting ATP1a1 link: https://peerj.com/articles/9060 last-modified: 2020-05-01 description: Genome editing in human induced pluripotent stem cells (iPSCs) provides the potential for disease modeling and cell therapy. By generating iPSCs with specific mutations, researchers can differentiate the modified cells to their lineage of interest for further investigation. However, the low efficiency of targeting in iPSCs has hampered the application of genome editing. In this study we used a CRISPR-Cas9 system that introduces a specific point substitution into the sequence of the Na+/K+-ATPase subunit ATP1A1. The introduced mutation confers resistance to cardiac glycosides, which can then be used to select successfully targeted cells. Using this system, we introduced different formats of donor DNA for homology-directed repair (HDR), including single-strand DNAs, double-strand DNAs, and plasmid donors. We achieved a 35-fold increase in HDR when using plasmid donor with a 400 bp repair template. We further co-targeted ATP1A1 and a second locus of interest to determine the enrichment of mutagenesis after cardiac glycoside selection. Through this approach, INDEL rate was increased after cardiac glycoside treatment, while HDR enrichment was only observed at certain loci. Collectively, these results suggest that a plasmid donor with a 400 bp repair template is an optimal donor DNA for targeted substitution and co-targeting ATP1A1 with the second locus enriches for mutagenesis events through cardiac glycoside selection in human iPSCs. creator: Jui-Tung Liu creator: James L. Corbett creator: James A. Heslop creator: Stephen A. Duncan uri: https://doi.org/10.7717/peerj.9060 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Liu et al. title: Assessing the growth in clinical skills using a progress clinical skills examination link: https://peerj.com/articles/9091 last-modified: 2020-05-01 description: BackgroundThis study evaluates the generalizability of an eight-station progress clinical skills examination and assesses the growth in performance for six clinical skills domains among first- and second-year medical students over four time points during the academic year.MethodsWe conducted a generalizability study for longitudinal and cross-sectional comparisons and assessed growth in six clinical skill domains via repeated measures ANOVA over the first and second year of medical school.ResultsThe generalizability of the examination domain scores was low but consistent with previous studies of data gathering and communication skills. Variations in case difficulty across administrations of the examination made it difficult to assess longitudinal growth. It was possible to compare students at different training levels and the interaction of training level and growth. Second-year students outperformed first-year students, but first-year students’ clinical skills performance grew faster than second-year students narrowing the gap in clinical skills over the students’ first year of medical school.ConclusionsCase specificity limits the ability to assess longitudinal growth in clinical skills through progress testing. Providing students with early clinical skills training and authentic clinical experiences appears to result in the rapid growth of clinical skills during the first year of medical school. creator: Heather S. Laird-Fick creator: Chi Chang creator: Ling Wang creator: Carol Parker creator: Robert Malinowski creator: Matthew Emery creator: David J. Solomon uri: https://doi.org/10.7717/peerj.9091 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Laird-Fick et al. title: Enhancing flavonoid production by promiscuous activity of prenyltransferase, BrPT2 from Boesenbergia rotunda link: https://peerj.com/articles/9094 last-modified: 2020-05-01 description: Flavonoids and prenylated flavonoids are active components in medicinal plant extracts which exhibit beneficial effects on human health. Prenylated flavonoids consist of a flavonoid core with a prenyl group attached to it. This prenylation process is catalyzed by prenyltranferases (PTs). At present, only a few flavonoid-related PT genes have been identified. In this study, we aimed to investigate the roles of PT in flavonoid production. We isolated a putative PT gene (designated as BrPT2) from a medicinal ginger, Boesenbergia rotunda. The deduced protein sequence shared highest gene sequence homology (81%) with the predicted homogentisate phytyltransferase 2 chloroplastic isoform X1 from Musa acuminata subsp. Malaccensis. We then cloned the BrPT2 into pRI vector and expressed in B. rotunda cell suspension cultures via Agrobacterium-mediated transformation. The BrPT2-expressing cells were fed with substrate, pinostrobin chalcone, and their products were analyzed by liquid chromatography mass spectrometry. We found that the amount of flavonoids, namely alpinetin, pinostrobin, naringenin and pinocembrin, in BrPT2-expressing cells was higher than those obtained from the wild type cells. However, we were unable to detect any targeted prenylated flavonoids. Further in-vitro assay revealed that the reaction containing the BrPT2 protein produced the highest accumulation of pinostrobin from the substrate pinostrobin chalcone compared to the reaction without BrPT2 protein, suggesting that BrPT2 was able to accelerate the enzymatic reaction. The finding of this study implied that the isolated BrPT2 may not be involved in the prenylation of pinostrobin chalcone but resulted in high yield and production of other flavonoids, which is likely related to enzyme promiscuous activities. creator: Yvonne Jing Mei Liew creator: Yean Kee Lee creator: Norzulaani Khalid creator: Noorsaadah Abd Rahman creator: Boon Chin Tan uri: https://doi.org/10.7717/peerj.9094 license: https://creativecommons.org/licenses/by/4.0/ rights: © 2020 Liew et al. title: Time matters: genetic composition and evaluation of effective population size in temperate coastal fish species link: https://peerj.com/articles/9098 last-modified: 2020-05-01 description: BackgroundExtensive knowledge on the genetic characterization of marine organisms has been assembled, mainly concerning the spatial distribution and structuring of populations. Temporal monitoring assesses not only the stability in genetic composition but also its trajectory over time, providing critical information for the accurate forecast of changes in genetic diversity of marine populations, particularly important for both fisheries and endangered species management. We assessed fluctuations in genetic composition among different sampling periods in the western Portuguese shore in three fish species.MethodsWhite seabream Diplodus sargus, sand smelt Atherina presbyter and shanny Lipophrys pholis were chosen, because of their genetic patterns in distinct ecological environments, insight into historical and contemporary factors influencing population effective size (Ne), and degree of commercial exploitation. Samples were obtained near Lisbon between 2003 and 2014 and screened for genetic variation with mitochondrial and nuclear markers. Analyses included genealogies, genetic diversities, temporal structures and contemporary Ne.ResultsFor mtDNA no temporal structure was detected, while for nDNA significant differences were recorded between some sampling periods for the shanny and the sand smelt. Haplotype networks revealed deep genealogies, with various levels of diversification. The shanny revealed a smaller Ne/generation when compared to the other species, which, in turn, revealed no evidence of genetic drift for most study periods. These results highlight the fact that temporal variations in genetic pool composition should be considered when evaluating the population structure of fish species with long distance dispersal, which are more vulnerable to recruitment fluctuations. creator: Sara M. Francisco creator: Joana I. Robalo uri: https://doi.org/10.7717/peerj.9098 license: https://creativecommons.org/licenses/by/4.0/ rights: ©2020 Francisco and Robalo