title: PeerJ description: Articles published in PeerJ link: https://peerj.com/articles/index.rss3?journal=peerj&month=2017-01 creator: info@peerj.com PeerJ errorsTo: info@peerj.com PeerJ language: en title: Boxer crabs induce asexual reproduction of their associated sea anemones by splitting and intraspecific theft link: https://peerj.com/articles/2954 last-modified: 2017-01-31 description: Crabs of the genus Lybia have the remarkable habit of holding a sea anemone in each of their claws. This partnership appears to be obligate, at least on the part of the crab. The present study focuses on Lybia leptochelis from the Red Sea holding anemones of the genus Alicia (family Aliciidae). These anemones have not been found free living, only in association with L. leptochelis. In an attempt to understand how the crabs acquire them, we conducted a series of behavioral experiments and molecular analyses. Laboratory observations showed that the removal of one anemone from a crab induces a “splitting” behavior, whereby the crab tears the remaining anemone into two similar parts, resulting in a complete anemone in each claw after regeneration. Furthermore, when two crabs, one holding anemones and one lacking them, are confronted, the crabs fight, almost always leading to the “theft” of a complete anemone or anemone fragment by the crab without them. Following this, crabs “split” their lone anemone into two. Individuals of Alicia sp. removed from freshly collected L. leptochelis were used for DNA analysis. By employing AFLP (Fluorescence Amplified Fragments Length Polymorphism) it was shown that each pair of anemones from a given crab is genetically identical. Furthermore, there is genetic identity between most pairs of anemone held by different crabs, with the others showing slight genetic differences. This is a unique case in which one animal induces asexual reproduction of another, consequently also affecting its genetic diversity. creator: Yisrael Schnytzer creator: Yaniv Giman creator: Ilan Karplus creator: Yair Achituv uri: https://doi.org/10.7717/peerj.2954 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Schnytzer et al. title: Catchments catch all in South African coastal lowlands: topography and palaeoclimate restricted gene flow in Nymania capensis (Meliaceae)—a multilocus phylogeographic and distribution modelling approach link: https://peerj.com/articles/2965 last-modified: 2017-01-31 description: BackgroundThis study investigates orbitally-forced range dynamics at a regional scale by exploring the evolutionary history of Nymania capensis (Meliaceae) across the deeply incised landscapes of the subescarpment coastal lowlands of South Africa; a region that is home to three biodiversity hotspots (Succulent Karoo, Fynbos, and Maputaland-Pondoland-Albany hotspots).MethodsA range of methods are used including: multilocus phylogeography (chloroplast and high- and low-copy nuclear DNA), molecular dating and species distribution modelling (SDM).ResultsThe results support an ‘evolutionarily distinct catchment’ hypothesis where: (1) different catchments contain genetically distinct lineages, (2) limited genetic structuring was detected within basins whilst high structuring was detected between basins, and (3) within primary catchment populations display a high degree of genealogical lineage sorting. In addition, the results support a glacial refugia hypothesis as: (a) the timing of chloroplast lineage diversification is restricted to the Pleistocene in a landscape that has been relatively unchanged since the late Pliocene, and (b) the projected LGM distribution of suitable climate for N. capensis suggest fragmentation into refugia that correspond to the current phylogeographic populations.DiscussionThis study highlights the interaction of topography and subtle Pleistocene climate variations as drivers limiting both seed and pollen flow along these lowlands. This lends support to the region’s large-scale conservation planning efforts, which used catchments as foundational units for conservation as these are likely to be evolutionarily significant units. creator: Alastair J. Potts uri: https://doi.org/10.7717/peerj.2965 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Potts title: Influence of different types of sessile epibionts on the community structure of mobile invertebrates in an eelgrass bed link: https://peerj.com/articles/2952 last-modified: 2017-01-31 description: Eelgrass (Zostera marina) beds are known to have high ecological and economical values within coastal ecosystems of the temperate northern hemisphere although their biodiversity and functions varied greatly from sites to sites. The variation in the biomass, abundance and diversity of mobile invertebrates in eelgrass beds has been examined in relation to various abiotic and biotic factors, such as water temperature, salinity, eelgrass biomass and epiphytic microalgae presence. However, the importance of sessile epibionts, such as macroalgae and calcific spirorbid polychaetes attached to eelgrass blades, has not been the focus of previous studies. In the present study, we examined the effects of three different sessile epibionts, namely, branched red algae, filamentous green algae, and calcific spirorbid polychaetes, on the biomass and diversity of mobile invertebrates in the eelgrass beds of Akkeshi in northeastern Japan. The relationships between seven abiotic and biotic variables including three types of epibionts, and biomass of 11 dominant mobile invertebrate species as well as three community-level variables (the total biomass of mobile invertebrates, species richness and the Shannon-Wiener species diversity index) were analyzed using a linear mixed model. Our results show that branched red algae are correlated with Pontogeneia rostrata, Lacuna spp., Nereis sp., Syllis sp. and the total biomass of mobile invertebrates, filamentous green algae with P. rostrata, Ansola angustata and the species diversity of mobile invertebrates, and spirorbid polychaetes with A. angustata, Lacuna spp., Siphonacmea oblongata, Syllis sp., the species richness and diversity of mobile invertebrates. The effect size of the epibionts was similar or even higher than that of abiotic and eelgrass factors on the total biomass of mobile invertebrates, species richness, species diversity and most of dominant invertebrate populations across the taxonomic groups. Consequently, epibiotic macroalgae and spirorbid polychaetes can be good predictors of the variation in the total biomass, species richness and species diversity of mobile invertebrates and the biomass of major dominant species, especially for species that have a relatively high dependency on eelgrass blades. These results suggest that the different functional groups of sessile epibionts have significant roles in determining the biomass and diversity of mobile invertebrates in eelgrass beds. creator: Kyosuke Momota creator: Masahiro Nakaoka uri: https://doi.org/10.7717/peerj.2952 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Momota and Nakaoka title: Genomic and transcriptomic resources for assassin flies including the complete genome sequence of Proctacanthus coquilletti (Insecta: Diptera: Asilidae) and 16 representative transcriptomes link: https://peerj.com/articles/2951 last-modified: 2017-01-31 description: A high-quality draft genome for Proctacanthus coquilletti (Insecta: Diptera: Asilidae) is presented along with transcriptomes for 16 Diptera species from five families: Asilidae, Apioceridae, Bombyliidae, Mydidae, and Tabanidae. Genome sequencing reveals that P. coquilletti has a genome size of approximately 210 Mbp and remarkably low heterozygosity (0.47%) and few repeats (15%). These characteristics helped produce a highly contiguous (N50 = 862 kbp) assembly, particularly given that only a single 2 × 250 bp PCR-free Illumina library was sequenced. A phylogenomic hypothesis is presented based on thousands of putative orthologs across the 16 transcriptomes. Phylogenetic relationships support the sister group relationship of Apioceridae + Mydidae to Asilidae. A time-calibrated phylogeny is also presented, with seven fossil calibration points, which suggests an older age of the split among Apioceridae, Asilidae, and Mydidae (158 mya) and Apioceridae and Mydidae (135 mya) than proposed in the AToL FlyTree project. Future studies will be able to take advantage of the resources presented here in order to produce large scale phylogenomic and evolutionary studies of assassin fly phylogeny, life histories, or venom. The bioinformatics tools and workflow presented here will be useful to others wishing to generate de novo genomic resources in species-rich taxa without a closely-related reference genome. creator: Rebecca B. Dikow creator: Paul B. Frandsen creator: Mauren Turcatel creator: Torsten Dikow uri: https://doi.org/10.7717/peerj.2951 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Dikow et al. title: The prognostic value of tumor length to resectable esophageal squamous cell carcinoma: a retrospective study link: https://peerj.com/articles/2943 last-modified: 2017-01-31 description: BackgroundThe current TNM classification system does not consider tumor length for patients with esophageal carcinoma (EC). This study explored the effect of tumor length, in addition to tumor depth and lymph node involvement, on survival in patients with esophageal squamous cell carcinoma (ESCC).MethodsA total of 498 ESCC patients who underwent surgical resection as the primary treatment were selected in the retrospective study. Pathological details were collected, which included tumor type, TNM stage, differentiation. Other collected information were: the types of esophageal resection, ABO blood group, family history and demographic and lifestyle factors. A time-dependent receiver operating characteristic (ROC) curve and a regression tree for survival were used to identify the cut-off point of tumor length, which was 3 cm. Univariate and multivariate Cox proportional hazard regression models were used to identify the prognostic factors to ESCC.Results & DiscussionThe 1-, 3-, 5-year overall survival rates were found to be 82.5%, 55.6%, and 35.1%, respectively. Patients who had larger tumor length (>3 cm) had a higher risk for death than the rest patients. From the univariate Cox proportional hazards regression model, the overall survival rate was significantly influenced by the depth of the tumor and lymph node involvement (either as dummy or continuous variables), Sex, and tumor length. Using these four variables in the multivariate Cox proportional hazard regression model, we found that the overall survival was significantly influenced by all variables except Sex. Therefore, in addition to the depth of the tumor and lymph node involvement (as either dummy or continuous variables), the tumor length is also an independent prognostic factor for ESCC. The overall survival rate was higher in a group with smaller tumor length (≤3 cm) than those patients with larger tumor length (>3 cm), no matter what the tumor stage was.ConclusionThe tumor length was found to be an important prognostic factor for ESCC patients without receiving neoadjuvant therapy. The modification of EC staging system may consider tumor length to better predict ESCC survival and identify higher risk patients for postoperative therapy. creator: Xiangwei Zhang creator: Yang Wang creator: Cheng Li creator: Jing Helmersson creator: Yuanzhu Jiang creator: Guoyuan Ma creator: Guanghui Wang creator: Wei Dong creator: Shaowei Sang creator: Jiajun Du uri: https://doi.org/10.7717/peerj.2943 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Zhang et al. title: Nest-site selection, reproductive ecology and shifts within core-use areas of Black-necked Cranes at the northern limit of the Tibetan Plateau link: https://peerj.com/articles/2939 last-modified: 2017-01-31 description: We investigated population dynamics, breeding pairs, breeding habitat selection, nest density, distance between neighboring nests, nest survival, reproductive success, and recruitment rate for Black-necked Cranes (BNC, Grus nigricollis) during 2013–2015 in Yanchiwan National Nature Reserve (YCW), Gansu, China. Numbers of BNC and breeding pairs remained relatively stable at around 140 individuals and 40 pairs. Recruitment rates ranged from 15.7% to 25.8%. The average nest distance was 718.66 ± 430.50 m (2013), 1064.51 ± 323.99 m (2014) and 534.99 ± 195.45 m (2015). Average nest survival rate, hatching success, and breeding success of all 29 nests were 65.56 ± 5.09%, 57.04 ± 6.12% and 32.78% ± 2.55. Water depth, water body area, and distance to land were positively related to nest survival, while disturbance level showed a negative relationship. However, nest site selection of BNC was determined by habitat type, disturbance and water depth. BNC often foraged in mudflats and freshwater marsh but seldom foraged in saline-alkali wet meadows due to food density and quantity in April, the month when BNC choose nest sites. Conservation strategies based on habitats should consider ecological factors that may not be well predicted by nest site selection. Shifts within core-use areas from satellite tracking of BNC demonstrated that maintaining populations demands that conservation areas are large enough to permit breeding BNC changes in space use. Our results are important for conservation management and provide quantitative reproductive data for this species. creator: Lixun Zhang creator: Bei An creator: Meilin Shu creator: Xiaojun Yang uri: https://doi.org/10.7717/peerj.2939 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Zhang et al. title: Investigation of Helicobacter pylori infection among symptomatic children in Hangzhou from 2007 to 2014: a retrospective study with 12,796 cases link: https://peerj.com/articles/2937 last-modified: 2017-01-31 description: Background and AimThe infection of Helicobacter pylori (H. pylori) is acquired in childhood and the prevalence vary greatly in different countries and regions. The study aimed to investigate the characteristics of H. pylori infection among children with gastrointestinal symptoms in Hangzhou, a representative city of eastern China.MethodsA systematic surveillance of H. pylori infection according to the 13C-urea breath test was conducted from January 2007 to December 2014 in the Children’s hospital, Zhejiang University School of Medicine. The demographic information and main symptoms of every subject were recorded.ResultsA total of 12,796 subjects were recruited and 18.6% children evaluated as H. pylori positive. The annual positive rates decreased from 2007 to 2014 (χ2 = 20.461, p < 0.01). The positive rates were 14.8%, 20.2% and 25.8% in 3–6, 7–11 and 12–17 years age group respectively, which increased with age (χ2 = 116.002, p < 0.01). And it was significantly higher in boys than girls (χ2 = 15.090, p < 0.01). Multivariate logistic regression identified possible risk factors for H. pylori infection. Age, gender, gastrointestinal symptoms and history of H. pylori infected family member were all significantly associated with H. pylori infection (all p < 0.05).ConclusionsH. pylori infection rates in children with gastrointestinal symptoms were lower than most of those reported in mainland China. Further studies are required to determine the prevalence in the general population. Comprehensively understanding of the characteristics and the possible risk factors of H. pylori infection will be helpful to its management strategies in children in China. creator: Xiaoli Shu creator: Mingfang Ping creator: Guofeng Yin creator: Mizu Jiang uri: https://doi.org/10.7717/peerj.2937 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Shu et al. title: An early geikiid dicynodont from the Tropidostoma Assemblage Zone (late Permian) of South Africa link: https://peerj.com/articles/2913 last-modified: 2017-01-31 description: Based on specimens previously identified as Tropidostoma, a new taxon of dicynodont (Bulbasaurus phylloxyron gen. et sp. nov.) from the Karoo Basin of South Africa is described. Bulbasaurus is a medium-sized dicynodont (maximum dorsal skull length 16.0 cm) restricted to the Tropidostoma Assemblage Zone (early Lopingian) of the Beaufort Group. Bulbasaurus can be distinguished from Tropidostoma by an array of characters including the presence of a tall, sharp premaxillary ridge, large, rugose, nearly-confluent nasal bosses, a nasofrontal ridge, massive tusks, robust pterygoids, prominently twisted subtemporal bar, and absence of a distinct postfrontal. Inclusion of Bulbasaurus in a phylogenetic analysis of anomodont therapsids recovers it as a member of Geikiidae, a clade of otherwise later Permian dicynodonts such as Aulacephalodon and Pelanomodon. Bulbasaurus exhibits many of the characters typical of adult Aulacephalodon, but at substantially smaller skull size (these characters are absent in comparably-sized Aulacephalodon juveniles), suggesting that the evolution of typical geikiid morphology preceded gigantism in the clade. Bulbasaurus is the earliest known geikiid and the only member of the group known from the Tropidostoma Assemblage Zone; discovery of this taxon shortens a perplexing ghost lineage and indicates that abundant clades from the later Permian of South Africa (e.g., Geikiidae, Dicynodontoidea) may have originated as rare components of earlier Karoo assemblage zones. creator: Christian F. Kammerer creator: Roger M.H. Smith uri: https://doi.org/10.7717/peerj.2913 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Kammerer et al. title: In situ morphometric survey elucidates the evolutionary systematics of the Eurasian Himantoglossum clade (Orchidaceae: Orchidinae) link: https://peerj.com/articles/2893 last-modified: 2017-01-31 description: Background and AimsThe charismatic Himantoglossum s.l. clade of Eurasian orchids contains an unusually large proportion of taxa that are of controversial circumscriptions and considerable conservation concern. Whereas our previously published study addressed the molecular phylogenetics and phylogeography of every named taxon within the clade, here we use detailed morphometric data obtained from the same populations to compare genotypes with associated phenotypes, in order to better explore taxonomic circumscription and character evolution within the clade.MethodsBetween one and 12 plants found in 25 populations that encompassed the entire distribution of the Himantoglossum s.l. clade were measured in situ for 51 morphological characters. Results for 45 of those characters were subjected to detailed multivariate and univariate analyses.Key ResultsMultivariate analyses readily separate subgenus Barlia and subgenus Comperia from subgenus Himantoglossum, and also the early-divergent H. formosum from the less divergent remainder of subgenus Himantoglossum. The sequence of divergence of these four lineages is confidently resolved. Our experimental approach to morphometric character analysis demonstrates clearly that phenotypic evolution within Himantoglossum is unusually multi-dimensional.ConclusionsDegrees of divergence between taxa shown by morphological analyses approximate those previously shown using molecular analyses. Himantoglossum s.l. is readily divisible into three subgenera. The three sections of subgenus Himantoglossum—hircinum, caprinum and formosum—are arrayed from west to east with only limited geographical overlap. At this taxonomic level, their juxtaposition combines with conflict between contrasting datasets to complicate attempts to distinguish between clinal variation and the discontinuities that by definition separate bona fide species. All taxa achieve allogamy via food deceit and have only weak pollinator specificity. Artificial crossing demonstrates that intrinsic sterility barriers are weak. Although we have found evidence of gene flow among and within the three sections of subgenus Himantoglossum, reports of natural hybrids are surprisingly rare, probably because putative parents are sufficiently similar to questionably warrant the status of species. Phenological separation and increased xeromorphy characterise the origin of subgenus Barlia. Several individual morphological characters show evidence of parallel acquisition, and loss of features is especially frequent in floral markings among members of section caprinum. Detailed patterns of gain and loss demonstrate that several different categories of flower markings are inherited independently. Along with the dimensions of labellar lobes, these pigmentation characters have been over-emphasised in previous taxonomic treatments. Increased plant vigour was a crucial element of the origin of the genus, but vegetative characters underwent remarkably little subsequent evolution. Attempts to reconstruct hypothetical ancestors at internal nodes of the phylogeny are weakened by (a) uncertain placement of Steveniella as sister to Himantoglossum s.l. and (b) uncertain relationships among subtly different putative species within section caprinum. Nonetheless, heterochronic/allometric trends, ultimately limited by functional constraints, clearly dictate transitions between contrasting flower sizes and complex labellum shapes. creator: Richard M. Bateman creator: Attila Molnár V. creator: Gábor Sramkó uri: https://doi.org/10.7717/peerj.2893 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Bateman et al. title: Identification of four potential predicting miRNA biomarkers for multiple myeloma from published datasets link: https://peerj.com/articles/2831 last-modified: 2017-01-31 description: BackgroundMultiple myeloma is a cancer which has a high occurrence rate and causes great injury to people worldwide. In recent years, many studies reported the effects of miRNA on the appearance of multiple myeloma. However, due to the differences of samples and sequencing platforms, a large number of inconsistent results have been generated among these studies, which limited the cure of multiple myeloma at the miRNA level.MethodsWe performed meta-analyses to identify the key miRNA biomarkers which could be applied on the treatment of multiple myeloma. The key miRNAs were determined by overlap comparisons of seven datasets in multiple myeloma. Then, the target genes for key miRNAs were predicted by the software TargetScan. Additionally, functional enrichments and binding TFs were investigated by DAVID database and Tfacts database, respectively.ResultsFirstly, comparing the normal tissues, 13 miRNAs were differently expressed miRNAs (DEMs) for at least three datasets. They were considered as key miRNAs, with 12 up-regulated (hsa-miR-106b, hsa-miR-125b, hsa-miR-130b, hsa-miR-138, hsa-miR-15b, hsa-miR-181a, hsa-miR-183, hsa-miR-191, hsa-miR-19a, hsa-miR-20a, hsa-miR-221 and hsa-miR-25) and one down-regulated (hsa-miR-223). Secondly, functional enrichment analyses indicated that target genes of the upregulated miRNAs were mainly transcript factors and enriched in transcription regulation. Besides, these genes were enriched in multiple pathways: the cancer signal pathway, insulin signal metabolic pathway, cell binding molecules, melanin generation, long-term regression and P53 signaling pathway. However, no significant enrichment was found for target genes of the down-regulated genes. Due to the distinct regulation function, four miRNAs (hsa-miR-19a has-miR-221 has-miR25 and has-miR223) were ascertained as the potential prognostic and diagnostic markers in MM. Thirdly, transcript factors analysis unveiled that there were 148 TFs and 60 TFs which bind target genes of the up-regulated miRNAs and target genes of the down-regulated miRNAs, respectively. They respectively generated 652 and 139 reactions of TFs and target genes. Additionally, 50 (31.6%) TFs were shared, while higher specificity was found in TFs of target genes for the upregulated miRNAs.DiscussionsTogether, our findings provided the key miRNAs which affected occurrence of multiple myeloma and regulation function of these miRNAs. It is valuable for the prognosis and diagnosis of multiple myeloma. creator: Tian Xiang creator: Ai-Xin Hu creator: Peng Sun creator: Gao Liu creator: Gang Liu creator: Yan Xiao uri: https://doi.org/10.7717/peerj.2831 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Xiang et al. title: Expanding our understanding of the trade in marine aquarium animals link: https://peerj.com/articles/2949 last-modified: 2017-01-26 description: The trade of live marine animals for home and public aquaria has grown into a major global industry. Millions of marine fishes and invertebrates are removed from coral reefs and associated habitats each year. The majority are imported into the United States, with the remainder sent to Europe, Japan, and a handful of other countries. Despite the recent growth and diversification of the aquarium trade, to date, data collection is not mandatory, and hence comprehensive information on species volume and diversity is lacking. This lack of information makes it impossible to study trade pathways. Without species-specific volume and diversity data, it is unclear how importing and exporting governments can oversee this industry effectively or how sustainability should be encouraged. To expand our knowledge and understanding of the trade, and to effectively communicate this new understanding, we introduce the publically-available Marine Aquarium Biodiversity and Trade Flow online database (https://www.aquariumtradedata.org/). This tool was created to communicate the volume and diversity of marine fishes and/or invertebrates imported into the US over three complete years (2008, 2009, and 2011) and three partial years (2000, 2004, 2005). To create this tool, invoices pertaining to shipments of live marine fishes and invertebrates were scanned and analyzed for species name, species quantities, country of origin, port of entry, and city of import destination. Here we focus on the analysis of the later three years of data and also produce an estimate for the entirety of 2000, 2004, and 2005. The three-year aggregate totals (2008, 2009, 2011) indicate that just under 2,300 fish and 725 invertebrate species were imported into the US cumulatively, although just under 1,800 fish and 550 invertebrate species were traded annually. Overall, the total number of live marine animals decreased between 2008 and 2011. In 2008, 2009, and 2011, the total number of individual fish (8.2, 7.3, and 6.9 million individuals) and invertebrates (4.2, 3.7, and 3.6 million individuals) assessed by analyzing the invoice data are roughly 60% of the total volumes recorded through the Law Enforcement Management Information System (LEMIS) dataset. Using these complete years, we back-calculated the number of individuals of both fishes and invertebrates imported in 2000, 2004, and 2005. These estimates (9.3, 10.8, and 11.2 million individual fish per year) were consistent with the three years of complete data. We also use these data to understand the global trade in two species (Banggai cardinalfish, Pterapogon kauderni, and orange clownfish, Amphiprion ocellaris / percula) recently considered for Endangered Species Act listing. Aquariumtradedata.org can help create more effective management plans for the traded species, and ideally could be implemented at key trade ports to better assess the global trade of aquatic wildlife. creator: Andrew L. Rhyne creator: Michael F. Tlusty creator: Joseph T. Szczebak creator: Robert J. Holmberg uri: https://doi.org/10.7717/peerj.2949 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Rhyne et al. title: Metagenomic identification of active methanogens and methanotrophs in serpentinite springs of the Voltri Massif, Italy link: https://peerj.com/articles/2945 last-modified: 2017-01-26 description: The production of hydrogen and methane by geochemical reactions associated with the serpentinization of ultramafic rocks can potentially support subsurface microbial ecosystems independent of the photosynthetic biosphere. Methanogenic and methanotrophic microorganisms are abundant in marine hydrothermal systems heavily influenced by serpentinization, but evidence for methane-cycling archaea and bacteria in continental serpentinite springs has been limited. This report provides metagenomic and experimental evidence for active methanogenesis and methanotrophy by microbial communities in serpentinite springs of the Voltri Massif, Italy. Methanogens belonging to family Methanobacteriaceae and methanotrophic bacteria belonging to family Methylococcaceae were heavily enriched in three ultrabasic springs (pH 12). Metagenomic data also suggest the potential for hydrogen oxidation, hydrogen production, carbon fixation, fermentation, and organic acid metabolism in the ultrabasic springs. The predicted metabolic capabilities are consistent with an active subsurface ecosystem supported by energy and carbon liberated by geochemical reactions within the serpentinite rocks of the Voltri Massif. creator: William J. Brazelton creator: Christopher N. Thornton creator: Alex Hyer creator: Katrina I. Twing creator: August A. Longino creator: Susan Q. Lang creator: Marvin D. Lilley creator: Gretchen L. Früh-Green creator: Matthew O. Schrenk uri: https://doi.org/10.7717/peerj.2945 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Brazelton et al. title: RelocaTE2: a high resolution transposable element insertion site mapping tool for population resequencing link: https://peerj.com/articles/2942 last-modified: 2017-01-26 description: BackgroundTransposable element (TE) polymorphisms are important components of population genetic variation. The functional impacts of TEs in gene regulation and generating genetic diversity have been observed in multiple species, but the frequency and magnitude of TE variation is under appreciated. Inexpensive and deep sequencing technology has made it affordable to apply population genetic methods to whole genomes with methods that identify single nucleotide and insertion/deletion polymorphisms. However, identifying TE polymorphisms, particularly transposition events or non-reference insertion sites can be challenging due to the repetitive nature of these sequences, which hamper both the sensitivity and specificity of analysis tools.MethodsWe have developed the tool RelocaTE2 for identification of TE insertion sites at high sensitivity and specificity. RelocaTE2 searches for known TE sequences in whole genome sequencing reads from second generation sequencing platforms such as Illumina. These sequence reads are used as seeds to pinpoint chromosome locations where TEs have transposed. RelocaTE2 detects target site duplication (TSD) of TE insertions allowing it to report TE polymorphism loci with single base pair precision.Results and DiscussionThe performance of RelocaTE2 is evaluated using both simulated and real sequence data. RelocaTE2 demonstrate high level of sensitivity and specificity, particularly when the sequence coverage is not shallow. In comparison to other tools tested, RelocaTE2 achieves the best balance between sensitivity and specificity. In particular, RelocaTE2 performs best in prediction of TSDs for TE insertions. Even in highly repetitive regions, such as those tested on rice chromosome 4, RelocaTE2 is able to report up to 95% of simulated TE insertions with less than 0.1% false positive rate using 10-fold genome coverage resequencing data. RelocaTE2 provides a robust solution to identify TE insertion sites and can be incorporated into analysis workflows in support of describing the complete genotype from light coverage genome sequencing. creator: Jinfeng Chen creator: Travis R. Wrightsman creator: Susan R. Wessler creator: Jason E. Stajich uri: https://doi.org/10.7717/peerj.2942 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Chen et al. title: Sequence variability of Chrysanthemum stunt viroid in different chrysanthemum cultivars link: https://peerj.com/articles/2933 last-modified: 2017-01-26 description: Viroids are the smallest infectious agents, and their genomes consist of a short single strand of RNA that does not encode any protein. Chrysanthemum stunt viroid (CSVd), a member of the family Pospiviroidae, causes chrysanthemum stunt disease. Here, we report the genomic variations of CSVd to understand the sequence variability of CSVd in different chrysanthemum cultivars. We randomly sampled 36 different chrysanthemum cultivars and examined the infection of CSVd in each cultivar by reverse transcription polymerase chain reaction (RT-PCR). Eleven cultivars were infected by CSVd. Cloning followed by Sanger sequencing successfully identified a total of 271 CSVd genomes derived from 12 plants from 11 cultivars. They were further classified into 105 CSVd variants. Each single chrysanthemum plant had a different set of CSVd variants. Moreover, different single plants from the same cultivar had different sets of CSVd variants but identical consensus genome sequences. A phylogenetic tree using 12 consensus genome sequences revealed three groups of CSVd genomes, while six different groups were defined by the phylogenetic analysis using 105 variants. Based on the consensus CSVd genome, by combining all variant sequences, we identified 99 single-nucleotide variations (SNVs) as well as three nucleotide positions showing high mutation rates. Although 99 SNVs were identified, most CSVd genomes in this study were derived from variant 1, which is identical to known CSVd SK1 showing pathogenicity. creator: Hoseong Choi creator: Yeonhwa Jo creator: Ju-Yeon Yoon creator: Seung-Kook Choi creator: Won Kyong Cho uri: https://doi.org/10.7717/peerj.2933 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Choi et al. title: Causal events enter awareness faster than non-causal events link: https://peerj.com/articles/2932 last-modified: 2017-01-26 description: Philosophers have long argued that causality cannot be directly observed but requires a conscious inference (Hume, 1967). Albert Michotte however developed numerous visual phenomena in which people seemed to perceive causality akin to primary visual properties like colour or motion (Michotte, 1946). Michotte claimed that the perception of causality did not require a conscious, deliberate inference but, working over 70 years ago, he did not have access to the experimental methods to test this claim. Here we employ Continuous Flash Suppression (CFS)—an interocular suppression technique to render stimuli invisible (Tsuchiya & Koch, 2005)—to test whether causal events enter awareness faster than non-causal events. We presented observers with ‘causal’ and ‘non-causal’ events, and found consistent evidence that participants become aware of causal events more rapidly than non-causal events. Our results suggest that, whilst causality must be inferred from sensory evidence, this inference might be computed at low levels of perceptual processing, and does not depend on a deliberative conscious evaluation of the stimulus. This work therefore supports Michotte’s contention that, like colour or motion, causality is an immediate property of our perception of the world. creator: Pieter Moors creator: Johan Wagemans creator: Lee de-Wit uri: https://doi.org/10.7717/peerj.2932 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Moors et al. title: Floral vasculature and its variation for carpellary supply in Anthurium (Araceae, Alismatales) link: https://peerj.com/articles/2929 last-modified: 2017-01-26 description: Introduction and AimsAnthurium is the largest genus of Araceae, with 950 species distributed in the neotropics. Despite the great diversity of the genus, the knowledge of its floral vasculature is based on observations in only two species, viz. A. denudatum and A. lhotzkyanum, with remarkable variation in vascular carpellary supply: carpels are either vascularized by ventral bundles alone or by reduced dorsal bundles in addition to the ventral ones. Our main objective is to test this peculiar variation through a detailed anatomical study of the floral vasculature in taxa belonging to some sections of Anthurium designated as monophyletic groups in recent phylogenies.MethodsWe compare the floral vasculature of 20 neotropical species belonging to distinct sections of Anthurium, using both light and confocal laser scanning microscopies.ResultsThe number and position of vascular bundles are constant within the tepals and stamens, regardless of the species and sections studied. However, the gynoecium vasculature exhibits variation between species belonging to the same or different sections. Our results reveal two patterns of vasculature: carpels vascularized by synlateral bundles alone (Pattern A) and carpels vascularized by both dorsal and synlateral bundles (Pattern B). Pattern A is shared by the majority of species studied here and corroborates the previous data in the literature. Pattern B occurs in three species: A. affine (Anthurium sect. Pachyneurium series Pachyneurium), A. obtusum and A. scandens (Anthurium sect. Tetraspermium), described here for the first time for the genus.ConclusionsThe variation in the supply to the carpels in Anthurium is corroborated here. However, our results in addition to those from the available literature suggest the existence of three patterns (A, B and C) of carpellary vasculature. Based on the recent phylogeny of Anthurium it is possible to notice that the three patterns of carpellary vasculature occur in representatives of Clade B and deserve to be investigated in a larger number of species. Pattern A could be a plesiomorphy for the genus and the occurrence of dorsal bundles could be a derived character. Our data contributes to the taxonomy and to the understanding of the floral evolution of the largest neotropical genus of Araceae. creator: Letícia P. Poli creator: Lívia G. Temponi creator: Alessandra I. Coan uri: https://doi.org/10.7717/peerj.2929 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Poli et al. title: Evolution of gremlin 2 in cetartiodactyl mammals: gene loss coincides with lack of upper jaw incisors in ruminants link: https://peerj.com/articles/2901 last-modified: 2017-01-26 description: Understanding the processes that give rise to genomic variability in extant species is an active area of research within evolutionary biology. With the availability of whole genome sequences, it is possible to quantify different forms of variability such as variation in gene copy number, which has been described as an important source of genetic variability and in consequence of phenotypic variability. Most of the research on this topic has been focused on understanding the biological significance of gene duplication, and less attention has been given to the evolutionary role of gene loss. Gremlin 2 is a member of the DAN gene family and plays a significant role in tooth development by blocking the ligand-signaling pathway of BMP2 and BMP4. The goal of this study was to investigate the evolutionary history of gremlin 2 in cetartiodactyl mammals, a group that possesses highly divergent teeth morphology. Results from our analyses indicate that gremlin 2 has experienced a mixture of gene loss, gene duplication, and rate acceleration. Although the last common ancestor of cetartiodactyls possessed a single gene copy, pigs and camels are the only cetartiodactyl groups that have retained gremlin 2. According to the phyletic distribution of this gene and synteny analyses, we propose that gremlin 2 was lost in the common ancestor of ruminants and cetaceans between 56.3 and 63.5 million years ago as a product of a chromosomal rearrangement. Our analyses also indicate that the rate of evolution of gremlin 2 has been accelerated in the two groups that have retained this gene. Additionally, the lack of this gene could explain the high diversity of teeth among cetartiodactyl mammals; specifically, the presence of this gene could act as a biological constraint. Thus, our results support the notions that gene loss is a way to increase phenotypic diversity and that gremlin 2 is a dispensable gene, at least in cetartiodactyl mammals. creator: Juan C. Opazo creator: Kattina Zavala creator: Paola Krall creator: Rodrigo A. Arias uri: https://doi.org/10.7717/peerj.2901 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Opazo et al. title: Genomic insight into pathogenicity of dematiaceous fungus Corynespora cassiicola link: https://peerj.com/articles/2841 last-modified: 2017-01-26 description: Corynespora cassiicola is a common plant pathogen that causes leaf spot disease in a broad range of crop, and it heavily affect rubber trees in Malaysia (Hsueh, 2011; Nghia et al., 2008). The isolation of UM 591 from a patient’s contact lens indicates the pathogenic potential of this dematiaceous fungus in human. However, the underlying factors that contribute to the opportunistic cross-infection have not been fully studied. We employed genome sequencing and gene homology annotations in attempt to identify these factors in UM 591 using data obtained from publicly available bioinformatics databases. The assembly size of UM 591 genome is 41.8 Mbp, and a total of 13,531 (≥99 bp) genes have been predicted. UM 591 is enriched with genes that encode for glycoside hydrolases, carbohydrate esterases, auxiliary activity enzymes and cell wall degrading enzymes. Virulent genes comprising of CAZymes, peptidases, and hypervirulence-associated cutinases were found to be present in the fungal genome. Comparative analysis result shows that UM 591 possesses higher number of carbohydrate esterases family 10 (CE10) CAZymes compared to other species of fungi in this study, and these enzymes hydrolyses wide range of carbohydrate and non-carbohydrate substrates. Putative melanin, siderophore, ent-kaurene, and lycopene biosynthesis gene clusters are predicted, and these gene clusters denote that UM 591 are capable of protecting itself from the UV and chemical stresses, allowing it to adapt to different environment. Putative sterigmatocystin, HC-toxin, cercosporin, and gliotoxin biosynthesis gene cluster are predicted. This finding have highlighted the necrotrophic and invasive nature of UM 591. creator: Hong Keat Looi creator: Yue Fen Toh creator: Su Mei Yew creator: Shiang Ling Na creator: Yung-Chie Tan creator: Pei-Sin Chong creator: Jia-Shiun Khoo creator: Wai-Yan Yee creator: Kee Peng Ng creator: Chee Sian Kuan uri: https://doi.org/10.7717/peerj.2841 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Looi et al. title: Computer simulation of Cerebral Arteriovenous Malformation—Validation analysis of hemodynamics parameters link: https://peerj.com/articles/2724 last-modified: 2017-01-26 description: ProblemThe purpose of this work is to provide some validation methods for evaluating the hemodynamic assessment of Cerebral Arteriovenous Malformation (CAVM). This article emphasizes the importance of validating noninvasive measurements for CAVM patients, which are designed using lumped models for complex vessel structure.MethodsThe validation of the hemodynamics assessment is based on invasive clinical measurements and cross-validation techniques with the Philips proprietary validated software’s Qflow and 2D Perfursion.ResultsThe modeling results are validated for 30 CAVM patients for 150 vessel locations. Mean flow, diameter, and pressure were compared between modeling results and with clinical/cross validation measurements, using an independent two-tailed Student t test. Exponential regression analysis was used to assess the relationship between blood flow, vessel diameter, and pressure between them. Univariate analysis is used to assess the relationship between vessel diameter, vessel cross-sectional area, AVM volume, AVM pressure, and AVM flow results were performed with linear or exponential regression.DiscussionModeling results were compared with clinical measurements from vessel locations of cerebral regions. Also, the model is cross validated with Philips proprietary validated software’s Qflow and 2D Perfursion. Our results shows that modeling results and clinical results are nearly matching with a small deviation.ConclusionIn this article, we have validated our modeling results with clinical measurements. The new approach for cross-validation is proposed by demonstrating the accuracy of our results with a validated product in a clinical environment. creator: Y. Kiran Kumar creator: Shashi Bhushan Mehta creator: Manjunath Ramachandra uri: https://doi.org/10.7717/peerj.2724 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Kumar et al. title: The use of gene interaction networks to improve the identification of cancer driver genes link: https://peerj.com/articles/2568 last-modified: 2017-01-26 description: Bioinformaticians have implemented different strategies to distinguish cancer driver genes from passenger genes. One of the more recent advances uses a pathway-oriented approach. Methods that employ this strategy are highly dependent on the quality and size of the pathway interaction network employed, and require a powerful statistical environment for analyses. A number of genomic libraries are available in R. DriverNet and DawnRank employ pathway-based methods that use gene interaction graphs in matrix form. We investigated the benefit of combining data from 3 different sources on the prediction outcome of cancer driver genes by DriverNet and DawnRank. An enriched dataset was derived comprising 13,862 genes with 372,250 interactions, which increased its accuracy by 17% and 28%, respectively, compared to their original networks. The study identified 33 new candidate driver genes. Our study highlights the potential of combining networks and weighting edges to provide greater accuracy in the identification of cancer driver genes. creator: Emilie Ramsahai creator: Kheston Walkins creator: Vrijesh Tripathi creator: Melford John uri: https://doi.org/10.7717/peerj.2568 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Ramsahai et al. title: Retrospective study of prognostic factors in pediatric invasive pneumococcal disease link: https://peerj.com/articles/2941 last-modified: 2017-01-25 description: Streptococcus pneumoniae remains the leading causative pathogen in pediatric pneumonia and bacteremia throughout the world. The invasive pneumococcal disease (IPD) is known as isolation of S. pneumoniae from a normally sterile site (e.g., blood, cerebrospinal fluid, synovial fluid, pericardial fluid, pleural fluid, or peritoneal fluid). The aim of this study is to survey the clinical manifestations and laboratory results of IPD and identify the prognostic factors of mortality. From January 2001 to December 2006, a retrospective review of chart was performed in a teaching hospital in Taipei. The hospitalized pediatric patients with the diagnosis of pneumonia, arthritis, infectious endocarditis, meningitis or sepsis were recruited. Among them, 50 patients were pneumococcal infections proved by positive culture results or antigen tests. Clinical manifestations, laboratory data and hospitalization courses were analyzed. The median age was 3.5-year-old and there were 30 male patients (60%). Eight patients (16%) had underlying disease such as leukemia or congenital heart disease. Hemolytic uremic syndrome (HUS) was observed in ten patients and extracorporeal membrane oxygenation (ECMO) was performed in three patients. Leukocytosis, elevated C-reactive protein and AST level were noted in most of the patients. The overall mortality rate was 10%. We found that leukopenia, thrombocytopenia and high CRP level were significant predictors for mortality. In conclusion, S. pneumoniae remains an important health threat worldwide and IPD is life-threatening with high mortality rate. We found leukopenia, thrombocytopenia, and high CRP levels to be associated with mortality in pediatric IPD, and these factors are worthy of special attention at admission. Although we failed to identify a statistically significant prognostic factor in multivariate analysis due to relatively small sample size, we suggest an aggressive antibiotic treatment in patients with these factors at admission. Further large-scale studies are warranted. creator: Nan-Chang Chiu creator: Hsin Chi creator: Chun-Chih Peng creator: Hung-Yang Chang creator: Daniel Tsung-Ning Huang creator: Lung Chang creator: Wei-Te Lei creator: Chien-Yu Lin uri: https://doi.org/10.7717/peerj.2941 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Chiu et al. title: Antibiotic resistance potential of the healthy preterm infant gut microbiome link: https://peerj.com/articles/2928 last-modified: 2017-01-25 description: BackgroundFew studies have investigated the gut microbiome of infants, fewer still preterm infants. In this study we sought to quantify and interrogate the resistome within a cohort of premature infants using shotgun metagenomic sequencing. We describe the gut microbiomes from preterm but healthy infants, characterising the taxonomic diversity identified and frequency of antibiotic resistance genes detected.ResultsDominant clinically important species identified within the microbiomes included C. perfringens, K. pneumoniae and members of the Staphylococci and Enterobacter genera. Screening at the gene level we identified an average of 13 antimicrobial resistance genes per preterm infant, ranging across eight different antibiotic classes, including aminoglycosides and fluoroquinolones. Some antibiotic resistance genes were associated with clinically relevant bacteria, including the identification of mecA and high levels of Staphylococci within some infants. We were able to demonstrate that in a third of the infants the S. aureus identified was unrelated using MLST or metagenome assembly, but low abundance prevented such analysis within the remaining samples.ConclusionsWe found that the healthy preterm infant gut microbiomes in this study harboured a significant diversity of antibiotic resistance genes. This broad picture of resistances and the wider taxonomic diversity identified raises further caution to the use of antibiotics without consideration of the resident microbial communities. creator: Graham Rose creator: Alexander G. Shaw creator: Kathleen Sim creator: David J. Wooldridge creator: Ming-Shi Li creator: Saheer Gharbia creator: Raju Misra creator: John Simon Kroll uri: https://doi.org/10.7717/peerj.2928 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Rose et al. title: Validation of ultrasonographic muscle thickness measurements as compared to the gold standard of computed tomography in dogs link: https://peerj.com/articles/2926 last-modified: 2017-01-25 description: ObjectiveThe objective was to quantitatively evaluate the validity of ultrasonographic (US) muscle measurements as compared to the gold standard of computed tomography (CT) in the canine.DesignThis was a prospective study.PopulationTwenty-five, client-owned dogs scheduled for CT as part of a diagnostic work-up for the management of their primary disease process were included.Materials and MethodsSpecific appendicular (cubital flexors and extensors, coxofemoral flexors and extensors) and axial (temporalis, supraspinatus, infraspinatus, lumbar epaxials) muscle groups were selected for quantitative measure based on CT planning and patient position. Prior to CT scan, the skin over the muscle sites was shaved and marked with a permanent marker. Patient body position was determined based on the patient’s CT plan; positioning was consistent between CT and US imaging. To ensure identical imaging position for both CT and US measurements, radio-opaque fiducial markers were placed directly over the skin marks once the dog was positioned. Quantitative measurements (cm) for both lean muscle mass (LMM) and subcutaneous adipose (SQA) were recorded. Statistical comparisons between CT and US values were done separately for each site and type.ResultsMuscle groups and associated SQA measured by US and CT were not statistically different based on an adjusted p-value using Bonferroni’s correction (p < 0.0031). In addition, all LMM and SQA sites had good reliability and agreement (Cronbach’s α = 0.8 − 1.0) between the two metrics, excluding the coxofemoral extensor muscle group (Cronbach’s α = 0.73232). Linear regression analysis of muscle measures indicated close agreement (slope range 0.93–1.09) and minimal bias of variation (intercept range 0.05–0.11) between CT versus US modalities, with the exception of the coxofemoral extensor muscle. Similarly, SQA CT and US measures indicated close agreement with the slope range of 0.88–1.02 and minimal bias of variation with an intercept range of 0.021–0.098, excluding the cubital flexor and extensor groups. Additionally, the R2 values for these remaining LMM and SQA sites are reported as >0.897 for LLM and >0.8289 for SQA.ConclusionsUltrasound imaging of selected appendicular and axial muscle groups in dogs can provide comparable assessment of muscle thickness to the current gold standard, CT. In consideration of both statistical reliability to CT and cage-side accessibility, the temporalis, supraspinatus, infraspinatus, and lumbar epaxial LMM sites are considered the most useful targets for US LMM assessment in the canine. Our findings support the potential utility of US as a clinical tool in veterinary medicine to assess LMM status in patients. Additional studies are indicated to develop standardized protocols of its use in a cage-side setting and to elucidate the benefit of this modality, in conjunction with nutritional interventions, to manage body LLM stores in compromised patients. creator: Lindsey E. Bullen creator: Maria G. Evola creator: Emily H. Griffith creator: Gabriela S. Seiler creator: Korinn E. Saker uri: https://doi.org/10.7717/peerj.2926 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Bullen et al. title: Effects of groundwater abstraction on two keystone tree species in an arid savanna national park link: https://peerj.com/articles/2923 last-modified: 2017-01-25 description: BackgroundIn arid systems with no surface water, deep boreholes in ephemeral river beds provide for humans and animals. With continually increasing infrastructure development for tourism in arid wildlife parks such as the Kgalagadi Transfrontier Park in southern Africa, we ask what effects increased abstraction may have on large trees. Large trees in arid savannas perform essential ecosystem services by providing food, shade, nesting sites and increased nutrients for many other plant and animal species and for this are regarded as keystone species.MethodsWe determine seasonal fluctuations in the water table while also determining the water source for the dominant large tree species in the Auob and Nossob rivers in the Park. We also determine the extent to which these trees are physiologically stressed using leaf δ13C, xylem pressure potentials, specific leaf area and an estimate of canopy death. We do this both upstream and downstream of a low water use borehole in the Auob River and a high water use borehole in the Nossob River.ResultsOur results show that the trees are indeed using deep groundwater in the wet season and that this is the same water used by people. In the dry season, trees in the Auob downstream of the active borehole become detached from the aquifer and use more isotopically enriched soil water. In the Nossob in the dry season, all trees use isotopically enriched soil water, and downstream of the active borehole use stomatal regulation to maintain leaf water potentials. These results suggest that trees in the more heavily utilised Nossob are under more water stress than those trees in the Auob but that trees in both rivers demonstrate physiological adaptation to the changes in available water with smaller heavier leaves, no significant canopy dieback and in the dry season in the Nossob stomatal regulation of leaf water potentials.DiscussionAn increase in abstraction of groundwater particularly at the Nossob borehole may cause an additional draw down of the water table adding to the physiological stress demonstrated in our study. The managers of the Kgalagadi Transfrontier Park have a mandate that includes biodiversity conservation. To fulfil this mandate, upper and lower thresholds for groundwater abstraction that allow for an adequate ecological reserve have to be determined. creator: Eleanor Shadwell creator: Edmund February uri: https://doi.org/10.7717/peerj.2923 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Shadwell and February title: Halocarbon emissions by selected tropical seaweeds: species-specific and compound-specific responses under changing pH link: https://peerj.com/articles/2918 last-modified: 2017-01-25 description: Five tropical seaweeds, Kappaphycus alvarezii (Doty) Doty ex P.C. Silva, Padina australis Hauck, Sargassum binderi Sonder ex J. Agardh (syn. S. aquifolium (Turner) C. Agardh), Sargassum siliquosum J. Agardh and Turbinaria conoides (J. Agardh) Kützing, were incubated in seawater of pH 8.0, 7.8 (ambient), 7.6, 7.4 and 7.2, to study the effects of changing seawater pH on halocarbon emissions. Eight halocarbon species known to be emitted by seaweeds were investigated: bromoform (CHBr3), dibro­momethane (CH2Br2), iodomethane (CH3I), diiodomethane (CH2I2), bromoiodomethane (CH2BrI), bromochlorometh­ane (CH2BrCl), bromodichloromethane (CHBrCl2), and dibro­mochloromethane (CHBr2Cl). These very short-lived halocarbon gases are believed to contribute to stratospheric halogen concentrations if released in the tropics. It was observed that the seaweeds emit all eight halocarbons assayed, with the exception of K. alvarezii and S. binderi for CH2I2 and CH3I respectively, which were not measurable at the achievable limit of detection. The effect of pH on halocarbon emission by the seaweeds was shown to be species-specific and compound specific. The highest percentage changes in emissions for the halocarbons of interest were observed at the lower pH levels of 7.2 and 7.4 especially in Padina australis and Sargassum spp., showing that lower seawater pH causes elevated emissions of some halocarbon compounds. In general the seaweed least affected by pH change in terms of types of halocarbon emission, was P. australis. The commercially farmed seaweed K. alvarezii was very sensitive to pH change as shown by the high increases in most of the compounds in all pH levels relative to ambient. In terms of percentage decrease in maximum quantum yield of photosynthesis (Fv∕Fm) prior to and after incubation, there were no significant correlations with the various pH levels tested for all seaweeds. The correlation between percentage decrease in the maximum quantum yield of photosynthesis (Fv∕Fm) and halocarbon emission rates, was significant only for CH2BrCl emission by P. australis (r = 0.47; p ≤ 0.04), implying that photosynthesis may not be closely linked to halocarbon emissions by the seaweeds studied. Bromine was the largest contributor to the total mass of halogen emitted for all the seaweeds at all pH. The highest total amount of bromine emitted by K. alvarezii (an average of 98% of total mass of halogens) and the increase in the total amount of chlorine with decreasing seawater pH fuels concern for the expanding seaweed farming activities in the ASEAN region. creator: Paramjeet Kaur Mithoo-Singh creator: Fiona S.-L. Keng creator: Siew-Moi Phang creator: Emma C. Leedham Elvidge creator: William T. Sturges creator: Gill Malin creator: Noorsaadah Abd Rahman uri: https://doi.org/10.7717/peerj.2918 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Mithoo-Singh et al. title: Polymorphism in the major histocompatibility complex (MHC class II B) genes of the Rufous-backed Bunting (Emberiza jankowskii) link: https://peerj.com/articles/2917 last-modified: 2017-01-25 description: Genetic diversity is one of the pillars of conservation biology research. High genetic diversity and abundant genetic variation in an organism may be suggestive of capacity to adapt to various environmental changes. The major histocompatibility complex (MHC) is known to be highly polymorphic and plays an important role in immune function. It is also considered an ideal model system to investigate genetic diversity in wildlife populations. The Rufous-backed Bunting (Emberiza jankowskii) is an endangered species that has experienced a sharp decline in both population and habitat size. Many historically significant populations are no longer present in previously populated regions, with only three breeding populations present in Inner Mongolia (i.e., the Aolunhua, Gahaitu and Lubei557 populations). Efforts focused on facilitating the conservation of the Rufous-backed Bunting (Emberiza jankowskii) are becoming increasingly important. However, the genetic diversity of E. jankowskii has not been investigated. In the present study, polymorphism in exon 2 of the MHCIIB of E. jankowskii was investigated. This polymorphism was subsequently compared with a related species, the Meadow Bunting (Emberiza cioides). A total of 1.59 alleles/individual were detected in E. jankowskii and 1.73 alleles/individual were identified in E. cioides. The maximum number of alleles per individual from the three E. jankowskii populations suggest the existence of at least three functional loci, while the maximum number of alleles per individual from the three E. cioides populations suggest the presence of at least four functional loci. Two of the alleles were shared between the E. jankowskii and E. cioides. Among the 12 unique alleles identified in E. jankowskii, 10.17 segregating sites per allele were detected, and the nucleotide diversity was 0.1865. Among the 17 unique alleles identified in E. cioides, eight segregating sites per allele were detected, and the nucleotide diversity was 0.1667. Overall, compared to other passerine birds, a relatively low level of MHC polymorphism was revealed in E. jankowskii, which was similar to that in E. cioides. Positive selection was detected by PAML/SLAC/FEL analyses in the region encoding the peptide-binding region in both species, and no recombination was detected. Phylogenetic analysis showed that the alleles from E. jankowskii and E. cioides belong to the same clade and the two species shared similar alleles, suggesting the occurrence of a trans-species polymorphism between the two Emberiza species. creator: Dan Li creator: Keping Sun creator: Yunjiao Zhao creator: Aiqing Lin creator: Shi Li creator: Yunlei Jiang creator: Jiang Feng uri: https://doi.org/10.7717/peerj.2917 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Li et al. title: Size matters: relationships between body size and body mass of common coastal, aquatic invertebrates in the Baltic Sea link: https://peerj.com/articles/2906 last-modified: 2017-01-25 description: BackgroundOrganism biomass is one of the most important variables in ecological studies, making biomass estimations one of the most common laboratory tasks. Biomass of small macroinvertebrates is usually estimated as dry mass or ash-free dry mass (hereafter ‘DM’ vs. ‘AFDM’) per sample; a laborious and time consuming process, that often can be speeded up using easily measured and reliable proxy variables like body size or wet (fresh) mass. Another common way of estimating AFDM (one of the most accurate but also time-consuming estimates of biologically active tissue mass) is the use of AFDM/DM ratios as conversion factors. So far, however, these ratios typically ignore the possibility that the relative mass of biologically active vs. non-active support tissue (e.g., protective exoskeleton or shell)—and therefore, also AFDM/DM ratios—may change with body size, as previously shown for taxa like spiders, vertebrates and trees.MethodsWe collected aquatic, epibenthic macroinvertebrates (>1 mm) in 32 shallow bays along a 360 km stretch of the Swedish coast along the Baltic Sea; one of the largest brackish water bodies on Earth. We then estimated statistical relationships between the body size (length or height in mm), body dry mass and ash-free dry mass for 14 of the most common taxa; five gastropods, three bivalves, three crustaceans and three insect larvae. Finally, we statistically estimated the potential influence of body size on the AFDM/DM ratio per taxon.ResultsFor most taxa, non-linear regression models describing the power relationship between body size and (i) DM and (ii) AFDM fit the data well (as indicated by low SE and high R2). Moreover, for more than half of the taxa studied (including the vast majority of the shelled molluscs), body size had a negative influence on organism AFDM/DM ratios.DiscussionThe good fit of the modelled power relationships suggests that the constants reported here can be used to quickly estimate organism dry- and ash-free dry mass based on body size, thereby freeing up considerable work resources. However, the considerable differences in constants between taxa emphasize the need for taxon-specific relationships, and the potential dangers associated with ignoring body size. The negative influence of body size on the AFDM/DM ratio found in a majority of the molluscs could be caused by increasingly thicker shells with organism age, and/or spawning-induced loss of biologically active tissue in adults. Consequently, future studies utilizing AFDM/DM (and presumably also AFDM/wet mass) ratios should carefully assess the potential influence of body size to ensure more reliable estimates of organism body mass. creator: Johan Eklöf creator: Åsa Austin creator: Ulf Bergström creator: Serena Donadi creator: Britas D.H.K. Eriksson creator: Joakim Hansen creator: Göran Sundblad uri: https://doi.org/10.7717/peerj.2906 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Eklöf et al. title: Exploration of immunoglobulin transcriptomes from mice immunized with three-finger toxins and phospholipases A2 from the Central American coral snake, Micrurus nigrocinctus link: https://peerj.com/articles/2924 last-modified: 2017-01-24 description: Snakebite envenomings represent a neglected public health issue in many parts of the rural tropical world. Animal-derived antivenoms have existed for more than a hundred years and are effective in neutralizing snake venom toxins when timely administered. However, the low immunogenicity of many small but potent snake venom toxins represents a challenge for obtaining a balanced immune response against the medically relevant components of the venom. Here, we employ high-throughput sequencing of the immunoglobulin (Ig) transcriptome of mice immunized with a three-finger toxin and a phospholipase A2 from the venom of the Central American coral snake, Micrurus nigrocinctus. Although exploratory in nature, our indicate results showed that only low frequencies of mRNA encoding IgG isotypes, the most relevant isotype for therapeutic purposes, were present in splenocytes of five mice immunized with 6 doses of the two types of toxins over 90 days. Furthermore, analysis of Ig heavy chain transcripts showed that no particular combination of variable (V) and joining (J) gene segments had been selected in the immunization process, as would be expected after a strong humoral immune response to a single antigen. Combined with the titration of toxin-specific antibodies in the sera of immunized mice, these data support the low immunogenicity of three-finger toxins and phospholipases A2found in M. nigrocinctusvenoms, and highlight the need for future studies analyzing the complexity of antibody responses to toxins at the molecular level. creator: Andreas H. Laustsen creator: Mikael Engmark creator: Christopher Clouser creator: Sonia Timberlake creator: Francois Vigneault creator: José María Gutiérrez creator: Bruno Lomonte uri: https://doi.org/10.7717/peerj.2924 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Laustsen et al. title: No apparent influence of psychometrically-defined schizotypy on orientation-dependent contextual modulation of visual contrast detection link: https://peerj.com/articles/2921 last-modified: 2017-01-24 description: We investigated the relationship between psychometrically-defined schizotypy and the ability to detect a visual target pattern. Target detection is typically impaired by a surrounding pattern (context) with an orientation that is parallel to the target, relative to a surrounding pattern with an orientation that is orthogonal to the target (orientation-dependent contextual modulation). Based on reports that this effect is reduced in those with schizophrenia, we hypothesised that there would be a negative relationship between the relative score on psychometrically-defined schizotypy and the relative effect of orientation-dependent contextual modulation. We measured visual contrast detection thresholds and scores on the Oxford-Liverpool Inventory of Feelings and Experiences (O-LIFE) from a non-clinical sample (N = 100). Contrary to our hypothesis, we find an absence of a monotonic relationship between the relative magnitude of orientation-dependent contextual modulation of visual contrast detection and the relative score on any of the subscales of the O-LIFE. The apparent difference of this result with previous reports on those with schizophrenia suggests that orientation-dependent contextual modulation may be an informative condition in which schizophrenia and psychometrically-defined schizotypy are dissociated. However, further research is also required to clarify the strength of orientation-dependent contextual modulation in those with schizophrenia. creator: Damien J. Mannion creator: Chris Donkin creator: Thomas J. Whitford uri: https://doi.org/10.7717/peerj.2921 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Mannion et al. title: Brood size and sex ratio in response to host quality and wasp traits in the gregarious parasitoid Oomyzus sokolowskii (Hymenoptera: Eulophidae) link: https://peerj.com/articles/2919 last-modified: 2017-01-24 description: This laboratory study investigated whether the larval-pupal parasitoid Oomyzus sokolowskii females adjust their brood size and sex ratio in response to body size and stage of Plutella xylostella larval hosts, as well as to their own body size and the order of oviposition. These factors were analyzed using multiple regression with simultaneous entry of them and their two-way interactions. Parasitoids brood size tended to increase with host body size at parasitism when the 4th instar larval host was attacked, but did not change when the 2nd and 3rd instar larvae were attacked. Parasitoids did not vary in brood size according to their body size, but decreased with their bouts of oviposition on a linear trend from 10 offspring adults emerged per host in the first bout of oviposition down to eight in the third. Parasitoid offspring sex ratio did not change with host instar, host body weight, wasp body size, and oviposition bout. Proportions of male offspring per brood were from 11% to 13% from attacking the 2nd to 4th instar larvae and from 13% to 16% across three successive bouts of oviposition, with a large variation for smaller host larvae and wasps. When fewer than 12 offspring were emerged from a host, one male was most frequently produced; when more than 12 offspring were emerged, two or more males were produced. Our study suggests that O. sokolowskii females may optimize their clutch size in response to body size of mature P. xylostella larvae, and their sex allocation in response to clutch size. creator: Xianwei Li creator: Liangting Zhu creator: Ling Meng creator: Baoping Li uri: https://doi.org/10.7717/peerj.2919 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Li et al. title: Evolutionary origins of the emergent ST796 clone of vancomycin resistant Enterococcus faecium link: https://peerj.com/articles/2916 last-modified: 2017-01-24 description: From early 2012, a novel clone of vancomycin resistant Enterococcus faecium (assigned the multi locus sequence type ST796) was simultaneously isolated from geographically separate hospitals in south eastern Australia and New Zealand. Here we describe the complete genome sequence of Ef_aus0233, a representative ST796 E. faecium isolate. We used PacBio single molecule real-time sequencing to establish a high quality, fully assembled genome comprising a circular chromosome of 2,888,087 bp and five plasmids. Comparison of Ef_aus0233 to other E. faecium genomes shows Ef_aus0233 is a member of the epidemic hospital-adapted lineage and has evolved from an ST555-like ancestral progenitor by the accumulation or modification of five mosaic plasmids and five putative prophage, acquisition of two cryptic genomic islands, accrued chromosomal single nucleotide polymorphisms and a 80 kb region of recombination, also gaining Tn1549 and Tn916, transposons conferring resistance to vancomycin and tetracycline respectively. The genomic dissection of this new clone presented here underscores the propensity of the hospital E. faecium lineage to change, presumably in response to the specific conditions of hospital and healthcare environments. creator: Andrew H. Buultjens creator: Margaret M.C. Lam creator: Susan Ballard creator: Ian R. Monk creator: Andrew A. Mahony creator: Elizabeth A. Grabsch creator: M. Lindsay Grayson creator: Stanley Pang creator: Geoffrey W. Coombs creator: J. Owen Robinson creator: Torsten Seemann creator: Paul D.R. Johnson creator: Benjamin P. Howden creator: Timothy P. Stinear uri: https://doi.org/10.7717/peerj.2916 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Buultjens et al. title: Allometry of sexual size dimorphism in turtles: a comparison of mass and length data link: https://peerj.com/articles/2914 last-modified: 2017-01-24 description: BackgroundThe macroevolutionary pattern of Rensch’s Rule (positive allometry of sexual size dimorphism) has had mixed support in turtles. Using the largest carapace length dataset and only large-scale body mass dataset assembled for this group, we determine (a) whether turtles conform to Rensch’s Rule at the order, suborder, and family levels, and (b) whether inferences regarding allometry of sexual size dimorphism differ based on choice of body size metric used for analyses.MethodsWe compiled databases of mean body mass and carapace length for males and females for as many populations and species of turtles as possible. We then determined scaling relationships between males and females for average body mass and straight carapace length using traditional and phylogenetic comparative methods. We also used regression analyses to evalutate sex-specific differences in the variance explained by carapace length on body mass.ResultsUsing traditional (non-phylogenetic) analyses, body mass supports Rensch’s Rule, whereas straight carapace length supports isometry. Using phylogenetic independent contrasts, both body mass and straight carapace length support Rensch’s Rule with strong congruence between metrics. At the family level, support for Rensch’s Rule is more frequent when mass is used and in phylogenetic comparative analyses. Turtles do not differ in slopes of sex-specific mass-to-length regressions and more variance in body size within each sex is explained by mass than by carapace length.DiscussionTurtles display Rensch’s Rule overall and within families of Cryptodires, but not within Pleurodire families. Mass and length are strongly congruent with respect to Rensch’s Rule across turtles, and discrepancies are observed mostly at the family level (the level where Rensch’s Rule is most often evaluated). At macroevolutionary scales, the purported advantages of length measurements over weight are not supported in turtles. creator: Koy W. Regis creator: Jesse M. Meik uri: https://doi.org/10.7717/peerj.2914 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Regis and Meik title: History of kidney stones and risk of chronic kidney disease: a meta-analysis link: https://peerj.com/articles/2907 last-modified: 2017-01-24 description: BackgroundAlthough the relationship between a history of kidney stones and chronic kidney disease (CKD) has been explored in many studies, it is still far from being well understood. Thus, we conducted a meta-analysis of studies comparing rates of CKD in patients with a history of kidney stones.MethodsPubMed, EMBASE, and the reference lists of relevant articles were searched to identify observational studies related to the topic. A random-effects model was used to combine the study-specific risk estimates. We explored the potential heterogeneity by subgroup analyses and meta-regression analyses.ResultsSeven studies were included in this meta-analysis. Pooled results suggested that a history of kidney stones was associated with an increased adjusted risk estimate for CKD [risk ratio (RR), 1.47 95% confidence interval (CI) [1.23–1.76])], with significant heterogeneity among these studies (I2 = 93.6%, P < 0.001). The observed positive association was observed in most of the subgroup analyses, whereas the association was not significant among studies from Asian countries, the mean age ≥50 years and male patients.ConclusionA history of kidney stones is associated with increased risk of CKD. Future investigations are encouraged to reveal the underlying mechanisms in the connection between kidney stones and CKD, which may point the way to more effective preventive and therapeutic measures. creator: Weifeng Shang creator: Lixi Li creator: Yali Ren creator: Qiangqiang Ge creator: Ming Ku creator: Shuwang Ge creator: Gang Xu uri: https://doi.org/10.7717/peerj.2907 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Shang et al. title: Epidemiological research on parent–child conflict in the United States: subgroup variations by place of birth and ethnicity, 2002–2013 link: https://peerj.com/articles/2905 last-modified: 2017-01-24 description: BackgroundChronically escalated parent–child conflict has been observed to elicit maladaptive behavior and reduced psychological well-being in children and youth. In this epidemiological study, we sought to estimate the occurrence of escalated parent–child conflict for United States (US) adolescent subgroups defined by (a) ethnic self-identification, and (b) nativity (US-born versus foreign-born).MethodsUS study populations of 12-to-17-year-olds were sampled, recruited, and assessed for the National Surveys on Drug Use and Health (NSDUH), 2002–2013 (n = 111, 129). Analysis-weighted contingency table analyses contrasted US-born versus foreign-born who self-identified as: (a) Hispanic, (b) non-Hispanic African-American, (c) non-Hispanic Asian, and (c) non-Hispanic White.ResultsFrequently escalated parent–child conflict was most prevalent among US-born non-Hispanic White adolescents, from 18% at age 12 (95% CI [17.6%, 18.9%]) to 29% at age 17 (95% CI [28.3%, 29.7%]), followed by US-born Hispanic and non-Hispanic Asian children. Estimated prevalence proportions were markedly lower for African-American children, from 8% at age 12 (95% CI [6.8, 8.5]) to 16% at age 17 (95% CI [14.3, 16.7]). Broad and sometimes overlapping CI indicate that larger sample sizes are needed for complete evaluation of an apparent excess occurrence of frequent parent-child conflict among US-born versus foreign-born. Nonetheless, in the larger subgroups, the US-born show a clear excess occurrence of frequent parent-child conflict. For example, US-born Mexican children have 1.7 times higher odds of experiencing frequent parent-child conflict than foreign-born Mexican children (OR = 1.7, 95% CI [1.5, 2.0], p-value < 0.001).DiscussionThe main discovery from this multi-ethnic sample investigation is a rank-ordering of parent-child conflict prevalence estimates from high (non-Hispanic White) to low (non-Hispanic African-American). The pattern also suggests a possibly generalizable excess associated with US-born sub-groups. The epidemiological estimates presented here merit attention in future cross-cultural research focused on parent-child conflict. creator: Jose Ruben Parra-Cardona creator: Hsueh-Han Yeh creator: James C. Anthony uri: https://doi.org/10.7717/peerj.2905 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Parra-Cardona et al. title: Bells, bomas and beefsteak: complex patterns of human-predator conflict at the wildlife-agropastoral interface in Zimbabwe link: https://peerj.com/articles/2898 last-modified: 2017-01-24 description: Reports of livestock depredation by large predators were systematically collected at three study sites in northwestern Zimbabwe from 2008–2013. We recorded 1,527 incidents (2,039 animals killed and 306 injured). Lions (Panthera leo) and spotted hyaenas (Crocuta crocuta) were mostly responsible, and cattle and donkeys most frequently attacked. Patterns of predation were variable among study sites. Nevertheless, some overall patterns were apparent. Predators selected livestock close to the size of their preferred wild prey, suggesting behaviours evolved to optimise foraging success may determine the domestic species primarily preyed upon. Most attacks occurred when livestock were roaming outside and away from their ‘home’ protective enclosures at night. Hyaena attacks were largely nocturnal; lions and leopards (Panthera pardus) were more flexible, with attacks occurring by day and at night. Livestock fitted with bells suffered a disproportionate number of attacks; the sound of bells appears to have conditioned predators to associate the sound with foraging opportunities. Lion and hyaena attacks on cattle were more frequent in the wet season suggesting that seasonal herding practices may result in cattle vulnerability. Only a small proportion of conflict incidents were reported to wildlife management officials with a bias towards lion predation events, potentially prejudicing conflict management policies. Predation on domestic stock involves an intricate interplay between predator behaviour and ecology on the one hand and human behaviour and husbandry practices on the other. Our data suggest that improved livestock husbandry (supervision of grazing animals, protection at night in strong enclosures) would greatly reduce livestock depredation. creator: Andrew J. Loveridge creator: Timothy Kuiper creator: Roger H. Parry creator: Lovemore Sibanda creator: Jane Hunt Hunt creator: Brent Stapelkamp creator: Lovelater Sebele creator: David W. Macdonald uri: https://doi.org/10.7717/peerj.2898 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Loveridge et al. title: An examination of factors potentially influencing birth distributions in golden snub-nosed monkeys (Rhinopithecus roxellana) link: https://peerj.com/articles/2892 last-modified: 2017-01-24 description: Many species of primates are considered seasonal breeders, but the set of factors, such as food availability, day length and temperature, that influence the timing of reproductive events for both wild and captive individuals remains unclear. Here, we examine the role of factors in shaping breeding patterns in Rhinopithecus roxellana, a temperate colobine primate. We used circular statistics to describe and compare the patterns of reproductive seasonality among individuals in 13 captive groups and two free ranging but provisioned groups at various locations throughout China. Almost 90% of births occurred in March, April and May in adult females residing in both free ranging (n = 131) and captive groups (n = 407). Births occurred principally in 2–4 months prior to the peak of food availability, while conceptions occurred in 1–2 months after the peak of food availability in free ranging but provisioned groups. Day length (latitude) had a significant effect on the timing of reproduction. However, females that experienced a wide variation of temperature between the lowest and highest monthly average temperature had a later conception date. These results support that day length and temperature might be factor influencing the timing of reproductive activity. creator: Zuofu Xiang creator: Wanji Yang creator: Xiaoguang Qi creator: Hui Yao creator: Cyril C. Grueter creator: Paul A. Garber creator: Baoguo Li creator: Ming Li uri: https://doi.org/10.7717/peerj.2892 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Xiang et al. title: Increased root hair density by loss of WRKY6 in Arabidopsis thaliana link: https://peerj.com/articles/2891 last-modified: 2017-01-24 description: Root hairs are unicellular elongations of certain rhizodermal cells that improve the uptake of sparingly soluble and immobile soil nutrients. Among different Arabidopsis thaliana genotypes, root hair density, length and the local acclimation to low inorganic phosphate (Pi) differs considerably, when analyzed on split agar plates. Here, genome-wide association fine mapping identified significant single nucleotide polymorphisms associated with the increased root hair density in the absence of local phosphate on chromosome 1. A loss-of-functionmutant of the candidate transcription factor gene WRKY6, which is involved in the acclimation of plants to low phosphorus, had increased root hair density. This is partially explained by a reduced cortical cell diameter in wrky6-3, reducing the rhizodermal cell numbers adjacent to the cortical cells. As a consequence, rhizodermal cells in positions that are in contact with two cortical cells are found more often, leading to higher hair density. Distinct cortical cell diameters and epidermal cell lengths distinguish other Arabidopsis accessions with distinct root hair density and −Pi response from diploid Col-0, while tetraploid Col-0 had generally larger root cell sizes, which explain longer hairs. A distinct radial root morphology within Arabidopsis accessions and wrky6-3explains some, but not all, differences in the root hair acclimation to –Pi. creator: Markus G. Stetter creator: Martin Benz creator: Uwe Ludewig uri: https://doi.org/10.7717/peerj.2891 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Stetter et al. title: The DNA methylation level against the background of the genome size and t-heterochromatin content in some species of the genus Secale L link: https://peerj.com/articles/2889 last-modified: 2017-01-24 description: Methylation of cytosine in DNA is one of the most important epigenetic modifications in eukaryotes and plays a crucial role in the regulation of gene activity and the maintenance of genomic integrity. DNA methylation and other epigenetic mechanisms affect the development, differentiation or the response of plants to biotic and abiotic stress. This study compared the level of methylation of cytosines on a global (ELISA) and genomic scale (MSAP) between the species of the genus Secale. We analyzed whether the interspecific variation of cytosine methylation was associated with the size of the genome (C-value) and the content of telomeric heterochromatin. MSAP analysis showed that S. sylvestre was the most distinct species among the studied rye taxa; however, the results clearly indicated that these differences were not statistically significant. The total methylation level of the studied loci was very similar in all taxa and ranged from 60% in S. strictum ssp. africanum to 66% in S. cereale ssp. segetale, which confirmed the lack of significant differences in the sequence methylation pattern between the pairs of rye taxa. The level of global cytosine methylation in the DNA was not significantly associated with the content of t-heterochromatin and did not overlap with the existing taxonomic rye relationships. The highest content of 5-methylcytosine was found in S. cereale ssp. segetale (83%), while very low in S. strictum ssp. strictum (53%), which was significantly different from the methylation state of all taxa, except for S. sylvestre. The other studied taxa of rye had a similar level of methylated cytosine ranging from 66.42% (S. vavilovii) to 74.41% in (S. cereale ssp. afghanicum). The results obtained in this study are evidence that the percentage of methylated cytosine cannot be inferred solely based on the genome size or t-heterochromatin. This is a significantly more complex issue. creator: Anna Kalinka creator: Magdalena Achrem creator: Paulina Poter uri: https://doi.org/10.7717/peerj.2889 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Kalinka et al. title: Three new karst-dwelling Cnemaspis Strauch, 1887 (Squamata; Gekkoniade) from Peninsular Thailand and the phylogenetic placement of C. punctatonuchalis and C. vandeventeri link: https://peerj.com/articles/2884 last-modified: 2017-01-24 description: Three new species of Rock Geckos Cnemaspis lineogularis sp. nov., C. phangngaensis sp. nov., and C. thachanaensis sp. nov. of the chanthaburiensis and siamensis groups are described from the Thai portion of the Thai-Malay Peninsula. These new species are distinguished from all other species in their two respective groups based on a unique combination of morphological characteristics, which is further supported by mitochondrial DNA (mtDNA) from the NADH dehydrogenase subunit 2 gene (ND2). Cnemaspis lineogularis sp. nov. is differentiated from all other species in the chanthaburiensis group by having a smaller maximum SVL 38 mm, 13 paravertebral tubercles, enlarged femoral scales, no caudal bands, and a 19.5–23.0% pairwise sequence divergence (ND2). Cnemaspis phangngaensis sp. nov. is differentiated from all other species in the siamensis group by having the unique combination of 10 infralabial scales, four continuous pore-bearing precloacal scales, paravertebral tubercles linearly arranged, lacking tubercles on the lower flanks, having ventrolateral caudal tubercles anteriorly present, caudal tubercles restricted to a single paraveterbral row on each side, a single median row of keeled subcaudals, and a 8.8–25.2% pairwise sequence divergence (ND2). Cnemaspis thachanaensis sp. nov. is distinguished from all other species in the siamensis group by having 10 or 11 supralabial scales 9–11 infralabial scales, paravertebral tubercles linearly arranged, ventrolateral caudal tubercles anteriorly, caudal tubercles restricted to a single paravertebral row on each side, a single median row of keeled subcaudal scales, lacking a single enlarged subcaudal scale row, lacking postcloaclal tubercles in males, the presence of an enlarged submetatarsal scale at the base if the 1st toe, and a 13.4–28.8% pairwise sequence divergence (ND2). The new phylogenetic analyses place C. punctatonuchalis and C. vandeventeri in the siamensis group with C. punctatonuchalis as the sister species to C. huaseesom and C. vandeventeri as the sister species to C. siamensis, corroborating previous hypotheses based on morphology. The discovery of three new karst-dwelling endemics brings the total number of nominal Thai Cnemaspis species to 15 and underscores the need for continued field research in poorly known areas of the Thai-Malay Peninsula, especially those that are threatened and often overlooked as biodiversity hot spots. creator: Perry Lee Wood Jr creator: L. Lee Grismer creator: Anchalee Aowphol creator: César A. Aguilar creator: Micheal Cota creator: Marta S. Grismer creator: Matthew L. Murdoch creator: Jack W. Sites Jr uri: https://doi.org/10.7717/peerj.2884 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Wood Jr et al. title: Developing the Leuven Embedded Figures Test (L-EFT): testing the stimulus features that influence embedding link: https://peerj.com/articles/2862 last-modified: 2017-01-24 description: BackgroundThe Embedded Figures Test (EFT, developed by Witkin and colleagues (1971)) has been used extensively in research on individual differences, particularly in the study of autism spectrum disorder. The EFT was originally conceptualized as a measure of field (in)dependence, but in recent years performance on the EFT has been interpreted as a measure of local versus global perceptual style. Although many have used the EFT to measure perceptual style, relatively few have focused on understanding the stimulus features that cause a shape to become embedded. The primary aim of this work was to investigate the relation between the strength of embedding and perceptual grouping on a group level.MethodNew embedded figure stimuli (both targets and contexts) were developed in which stimulus features that may influence perceptual grouping were explicitly manipulated. The symmetry, closure and complexity of the target shape were manipulated as well as its good continuation by varying the number of lines from the target that continued into the context. We evaluated the effect of these four stimulus features on target detection in a new embedded figures task (Leuven Embedded Figures Test, L-EFT) in a group of undergraduate psychology students. The results were then replicated in a second experiment using a slightly different version of the task.ResultsStimulus features that influence perceptual grouping, especially good continuation and symmetry, clearly affected performance (lower accuracy, slower response times) on the L-EFT. Closure did not yield results in line with our predictions.DiscussionThese results show that some stimulus features, which are known to affect perceptual grouping, also influence how effectively a stimulus becomes embedded in different contexts. Whether these results imply that the EFT measures individual differences in perceptual grouping ability must be further investigated. creator: Lee de-Wit creator: Hanne Huygelier creator: Ruth Van der Hallen creator: Rebecca Chamberlain creator: Johan Wagemans uri: https://doi.org/10.7717/peerj.2862 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 De-Wit et al. title: Selection and evaluation of reference genes for analysis of mouse (Mus musculus) sex-dimorphic brain development link: https://peerj.com/articles/2909 last-modified: 2017-01-19 description: The development of the brain is sex-dimorphic, and as a result so are many neurological disorders. One approach for studying sex-dimorphic brain development is to measure gene expression in biological samples using RT-qPCR. However, the accuracy and consistency of this technique relies on the reference gene(s) selected. We analyzed the expression of ten reference genes in male and female samples over three stages of brain development, using popular algorithms NormFinder, GeNorm and Bestkeeper. The top ranked reference genes at each time point were further used to quantify gene expression of three sex-dimorphic genes (Wnt10b, Xist and CYP7B1). When comparing gene expression between the sexes expression at specific time points the best reference gene combinations are: Sdha/Pgk1 at E11.5, RpL38/Sdha E12.5, and Actb/RpL37 at E15.5. When studying expression across time, the ideal reference gene(s) differs with sex. For XY samples a combination of Actb/Sdha. In contrast, when studying gene expression across developmental stage with XX samples, Sdha/Gapdh were the top reference genes. Our results identify the best combination of two reference genes when studying male and female brain development, and emphasize the importance of selecting the correct reference genes for comparisons between developmental stages. creator: Tanya T. Cheung creator: Mitchell K. Weston creator: Megan J. Wilson uri: https://doi.org/10.7717/peerj.2909 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Cheung et al. title: The impact of parental investment on lifetime reproductive success in Iceland link: https://peerj.com/articles/2904 last-modified: 2017-01-19 description: BackgroundDemonstrating the impact that parents have on the fitness of their children is a crucial step towards understanding how parental investment has affected human evolution. Parents not only transfer genes to their children, they also influence their environments. By analyzing reproductive patterns within and between different categories of close relatives, this study provides insight into the genetic and environmental effects that parents have on the fitness of their offspring.MethodsWe use data spanning over two centuries from an exceptionally accurate Icelandic genealogy, Íslendingabók, to analyze the relationship between the fertility rates of close relatives. Also, using genetic data, we determine narrow sense heritability estimates (h2) to further explore the genetic impact on lifetime reproductive success. Finally, we construct four simulations to model the expected contribution of genes and resources on reproductive success.ResultsThe relationship between the reproduction of all full sibling pairs was significant and positive across all birth decades (r = 0.19) while the reproductive relationship between parents and offspring was often negative across many decades and undetectable overall (r = 0.00) (Fig. 1 and Table 1). Meanwhile, genetic data among 8,456 pairs of full siblings revealed a narrow sense heritability estimate (h2) of 0.00 for lifetime reproductive success. A resources model (following the rule that resources are transmitted from parents to children, distributed equally among siblings, and are the only factor affecting reproductive success) revealed a similar trend: a negative relationship between parent and offspring reproduction (r =  − 0.35) but a positive relationship among full siblings (r = 0.28). The relationship between parent and offspring lifetime reproductive success (LRS) and full sibling LRS was strongly and positively correlated across time (r = 0.799, p < 0.001). Similarly, the LRS among full siblings was positively correlated with both the LRS among half siblings (r = 0.532, p = 0.011) and the relationship between the LRS of aunts and uncles with their nieces and nephews (r = 0.438, p = 0.042).DiscussionWe show that an individual’s lifetime reproductive success is best predicted by the reproduction of their full and half siblings, but not their parents, grandparents or aunts and uncles. Because all siblings share at least one parent, we believe parental investment has had an important impact on fitness. Overall, these results indicate that direct parental investment, but not genes, is likely to have had an important and persistent impact on lifetime reproductive success across more than two centuries of Icelandic history. creator: Robert F. Lynch creator: Emily C. Lynch uri: https://doi.org/10.7717/peerj.2904 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Lynch and Lynch title: Accelerometer-measured physical activity is not associated with two-year weight change in African-origin adults from five diverse populations link: https://peerj.com/articles/2902 last-modified: 2017-01-19 description: BackgroundIncreasing population-levels of physical activity (PA) is a controversial strategy for managing the obesity epidemic, given the conflicting evidence for weight loss from PA alone per se. We measured PA and weight change in a three-year prospective cohort study in young adults from five countries (Ghana, South Africa, Jamaica, Seychelles and USA).MethodsA total of 1,944 men and women had baseline data, and at least 1 follow-up examination including measures of anthropometry (weight/BMI), and objective PA (accelerometer, 7-day) following the three-year study period. PA was explored as 1-minute bouts of moderate and vigorous PA (MVPA) as well as daily sedentary time.ResultsAt baseline; Ghanaian and South African men had the lowest body weights (63.4 ± 9.5, 64.9 ± 11.8 kg, respectively) and men and women from the USA the highest (93.6 ± 25.9, 91.7 ± 23.4 kg, respectively). Prevalence of normal weight ranged from 85% in Ghanaian men to 29% in USA men and 52% in Ghanaian women to 15% in USA women. Over the two-year follow-up period, USA men and Jamaican women experienced the smallest yearly weight change rate (0.1 ± 3.3 kg/yr; −0.03 ± 3.0 kg/yr, respectively), compared to South African men and Ghanaian women greatest yearly change (0.6.0 ± 3.0 kg/yr; 1.22 ± 2.6 kg/yr, respectively). Mean yearly weight gain tended to be larger among normal weight participants at baseline than overweight/obese at baseline. Neither baseline MVPA nor sedentary time were associated with weight gain. Using multiple linear regression, only baseline weight, age and gender were significantly associated with weight gain.DiscussionFrom our study it is not evident that higher volumes of PA alone are protective against future weight gain, and by deduction our data suggest that other environmental factors such as the food environment may have a more critical role. creator: Lara R. Dugas creator: Stephanie Kliethermes creator: Jacob Plange-Rhule creator: Liping Tong creator: Pascal Bovet creator: Terrence E. Forrester creator: Estelle V. Lambert creator: Dale A. Schoeller creator: Ramon A. Durazo-Arvizu creator: David A. Shoham creator: Guichan Cao creator: Soren Brage creator: Ulf Ekelund creator: Richard S. Cooper creator: Amy Luke uri: https://doi.org/10.7717/peerj.2902 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Dugas et al. title: Panel strain of Klebsiella pneumoniae for beta-lactam antibiotic evaluation: their phenotypic and genotypic characterization link: https://peerj.com/articles/2896 last-modified: 2017-01-19 description: Klebsiella pneumoniae is responsible for numerous infections caused in hospitals, leading to mortality and morbidity. It has been evolving as a multi-drug resistant pathogen, acquiring multiple resistances such as such as horizontal gene transfer, transposon-mediated insertions or change in outer membrane permeability. Therefore, constant efforts are being carried out to control the infections using various antibiotic therapies. Considering the severity of the acquired resistance, we developed a panel of strains of K. pneumoniae expressing different resistance profiles such as high-level penicillinase and AmpC production, extended spectrum beta-lactamases and carbapenemases. Bacterial strains expressing different resistance phenotypes were collected and examined for resistance genes, mutations and porin alterations contributing to the detected phenotypes. Using the Massive parallel sequencing (MPS) technology we have constructed and genotypically characterized the panel strains to elucidate the multidrug resistance. These panel strains can be used in the clinical laboratory as standard reference strains. In addition, these strains could be significant in the field of pharmaceuticals for the antibiotic drug testing to verify its efficiency on pathogens expressing various  resistances. creator: Roshan Dsouza creator: Naina Adren Pinto creator: InSik Hwang creator: YoungLag Cho creator: Dongeun Yong creator: Jongrak Choi creator: Kyungwon Lee creator: Yunsop Chong uri: https://doi.org/10.7717/peerj.2896 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Dsouza et al. title: Jellyfish extract induces apoptotic cell death through the p38 pathway and cell cycle arrest in chronic myelogenous leukemia K562 cells link: https://peerj.com/articles/2895 last-modified: 2017-01-19 description: Jellyfish species are widely distributed in the world’s oceans, and their population is rapidly increasing. Jellyfish extracts have several biological functions, such as cytotoxic, anti-microbial, and antioxidant activities in cells and organisms. However, the anti-cancer effect of Jellyfish extract has not yet been examined. We used chronic myelogenous leukemia K562 cells to evaluate the mechanisms of anti-cancer activity of hexane extracts from Nomura’s jellyfish in vitro. In this study, jellyfish are subjected to hexane extraction, and the extract is shown to have an anticancer effect on chronic myelogenous leukemia K562 cells. Interestingly, the present results show that jellyfish hexane extract (Jellyfish-HE) induces apoptosis in a dose- and time-dependent manner. To identify the mechanism(s) underlying Jellyfish-HE-induced apoptosis in K562 cells, we examined the effects of Jellyfish-HE on activation of caspase and mitogen-activated protein kinases (MAPKs), which are responsible for cell cycle progression. Induction of apoptosis by Jellyfish-HE occurred through the activation of caspases-3,-8 and -9 and phosphorylation of p38. Jellyfish-HE-induced apoptosis was blocked by a caspase inhibitor, Z-VAD. Moreover, during apoptosis in K562 cells, p38 MAPK was inhibited by pretreatment with SB203580, an inhibitor of p38. SB203580 blocked jellyfish-HE-induced apoptosis. Additionally, Jellyfish-HE markedly arrests the cell cycle in the G0/G1 phase. Therefore, taken together, the results imply that the anti-cancer activity of Jellyfish-HE may be mediated apoptosis by induction of caspases and activation of MAPK, especially phosphorylation of p38, and cell cycle arrest at the Go/G1 phase in K562 cells. creator: Sun-Hyung Ha creator: Fansi Jin creator: Choong-Hwan Kwak creator: Fukushi Abekura creator: Jun-Young Park creator: Nam Gyu Park creator: Young-Chae Chang creator: Young-Choon Lee creator: Tae-Wook Chung creator: Ki-Tae Ha creator: Jong-Keun Son creator: Hyeun Wook Chang creator: Cheorl-Ho Kim uri: https://doi.org/10.7717/peerj.2895 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Ha et al. title: Ca2+ dynamics in zebrafish morphogenesis link: https://peerj.com/articles/2894 last-modified: 2017-01-19 description: Intracellular calcium ion (Ca2+) signaling is heavily involved in development, as illustrated by the use of a number of Ca2+ indicators. However, continuous Ca2+ patterns during morphogenesis have not yet been studied using fluorescence resonance energy transfer to track the Ca2+ sensor. In the present study, we monitored Ca2+ levels during zebrafish morphogenesis and differentiation with yellow cameleon, YC2.12. Our results show not only clear changes in Ca2+ levels but also continuous Ca2+ patterns at 24 hpf and later periods for the first time. Serial Ca2+dynamics during early pharyngula period (Prim-5-20; 24–33 hpf) was successfully observed with cameleon, which have not reported anywhere yet. In fact, high Ca2+ level occurred concurrently with hindbrain development in segmentation and pharyngula periods. Ca2+ patterns in the late gastrula through segmentation periods which were obtained with cameleon, were similar to those obtained previously with other Ca2+sensor. Our results suggested that the use of various Ca2+ sensors may lead to novel findings in studies of Ca2+ dynamics. We hope that these results will prove valuable for further research in Ca2+ signaling. creator: Yusuke Tsuruwaka creator: Eriko Shimada creator: Kenta Tsutsui creator: Tomohisa Ogawa uri: https://doi.org/10.7717/peerj.2894 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Tsuruwaka et al. title: Lepidopteran HMG-CoA reductase is a potential selective target for pest control link: https://peerj.com/articles/2881 last-modified: 2017-01-19 description: As a consequence of the negative impacts on the environment of some insecticides, discovery of eco-friendly insecticides and target has received global attention in recent years. Sequence alignment and structural comparison of the rate-limiting enzyme HMG-CoA reductase (HMGR) revealed differences between lepidopteran pests and other organisms, which suggested insect HMGR could be a selective insecticide target candidate. Inhibition of JH biosynthesis in vitro confirmed that HMGR inhibitors showed a potent lethal effect on the lepidopteran pest Manduca sexta, whereas there was little effect on JH biosynthesis in Apis mellifera and Diploptera punctata. The pest control application of these inhibitors demonstrated that they can be insecticide candidates with potent ovicidal activity, larvicidal activity and insect growth regulatory effects. The present study has validated that Lepidopteran HMGR can be a potent selective insecticide target, and the HMGR inhibitors (especially type II statins) could be selective insecticide candidates and lead compounds. Furthermore, we demonstrated that sequence alignment, homology modeling and structural comparison may be useful for determining potential enzymes or receptors which can be eco-friendly pesticide  targets. creator: Yuan-mei Li creator: Zhen-peng Kai creator: Juan Huang creator: Stephen S. Tobe uri: https://doi.org/10.7717/peerj.2881 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Li et al. title: Detecting heterogeneity in single-cell RNA-Seq data by non-negative matrix factorization link: https://peerj.com/articles/2888 last-modified: 2017-01-19 description: Single-cell RNA-Sequencing (scRNA-Seq) is a fast-evolving technology that enables the understanding of biological processes at an unprecedentedly high resolution. However, well-suited bioinformatics tools to analyze the data generated from this new technology are still lacking. Here we investigate the performance of non-negative matrix factorization (NMF) method to analyze a wide variety of scRNA-Seq datasets, ranging from mouse hematopoietic stem cells to human glioblastoma data. In comparison to other unsupervised clustering methods including K-means and hierarchical clustering, NMF has higher accuracy in separating similar groups in various datasets. We ranked genes by their importance scores (D-scores) in separating these groups, and discovered that NMF uniquely identifies genes expressed at intermediate levels as top-ranked genes. Finally, we show that in conjugation with the modularity detection method FEM, NMF reveals meaningful protein-protein interaction modules. In summary, we propose that NMF is a desirable method to analyze heterogeneous single-cell RNA-Seq data. The NMF based subpopulation detection package is available at: https://github.com/lanagarmire/NMFEM. creator: Xun Zhu creator: Travers Ching creator: Xinghua Pan creator: Sherman M. Weissman creator: Lana Garmire uri: https://doi.org/10.7717/peerj.2888 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Zhu et al. title: Detailed anatomy of the braincase of Macelognathus vagans Marsh, 1884 (Archosauria, Crocodylomorpha) using high resolution tomography and new insights on basal crocodylomorph phylogeny link: https://peerj.com/articles/2801 last-modified: 2017-01-19 description: BackgroundMacelognathus vagansMarsh, 1884 from the Late Jurassic Morrison Fm. of Wyoming was originally described as a dinosaur by Marsh and in 1971 Ostrom suggested crocodilian affinities. In 2005, Göhlich and collaborators identified new material of this species from Colorado as a basal crocodylomorph. However, a partial skull found in association with mandibular and postcranial remains was not described.MethodsDue to the small size and delicate structures within the braincase, micro CT studies were performed on this specimen. The new anatomical information was incorporated in a phylogenetic dataset, expanding both character and taxon sampling.ResultsThis new material reinforces the non-crocodyliform crocodylomorph affinities of Macelognathusas it bears a large otic aperture, unfused frontals and lacks ornamentation on the dorsal cranial bones. The internal structures also support these affinities as this specimen bears traits (i.e., heavily pneumatized and expanded basisphenoid; the presence of additional pneumatic features on the braincase; and the otoccipital-quadrate contact) not present in most basal crocodylomorphs. Furthermore, the presence of a wide supraoccipital and a cranioquadrate passage are traits shared with Almadasuchus from the early Late Jurassic of Argentina. Macelognathus was recovered as one of the closest relatives of crocodyliforms, forming a clade (Hallopodidae) with two other Late Jurassic taxa (Almadasuchus and Hallopus).DiscussionThe clade formed by Almadasuchus + Hallopus + Macelognathus, the Hallopodidae, is characterized by a higher degree of suturing of the braincase, posteriorly closed otic aperture (paralleled in mesoeucrocodylians) and cursorial adaptations. Also, the phylogenetic position of this lineage of derived crocodylomorphs as the sister group of Crocodyliformes implies a large amount of unsampled record (ghost lineage), at least 50 million years. creator: Juan Martin Leardi creator: Diego Pol creator: James Matthew Clark uri: https://doi.org/10.7717/peerj.2801 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Leardi et al. title: Osmolyte regulation by TonEBP/NFAT5 during anoxia-recovery and dehydration–rehydration stresses in the freeze-tolerant wood frog (Rana sylvatica) link: https://peerj.com/articles/2797 last-modified: 2017-01-19 description: BackgroundThe wood frog, Rana sylvatica, tolerates freezing as a means of winter survival. Freezing is considered to be an ischemic/anoxic event in which oxygen delivery is significantly impaired. In addition, cellular dehydration occurs during freezing because water is lost to extracellular compartments in order to promote freezing. In order to prevent severe cell shrinkage and cell death, it is important for the wood frog to have adaptive mechanisms for osmoregulation. One important mechanism of cellular osmoregulation occurs through the cellular uptake/production of organic osmolytes like sorbitol, betaine, and myo-inositol. Betaine and myo-inositol are transported by the proteins BGT-1 and SMIT, respectively. Sorbitol on the other hand, is synthesized inside the cell by the enzyme aldose reductase. These three proteins are regulated at the transcriptional level by the transcription factor, NFAT5/TonEBP. Therefore, the objective of this study was to elucidate the role of NFAT5/TonEBP in regulating BGT-1, SMIT, and aldose reductase, during dehydration and anoxia in the wood frog muscle, liver, and kidney tissues.MethodsWood frogs were subjected to 24 h anoxia-4 h recovery and 40% dehydration-full rehydration experiments. Protein levels of NFAT5, BGT-1, SMIT, and aldose reductase were studied using immunoblotting in muscle, liver, and kidney tissues.ResultsImmunoblotting results demonstrated downregulations in NFAT5 protein levels in both liver and kidney tissues during anoxia (decreases by 41% and 44% relative to control for liver and kidney, respectively). Aldose reductase protein levels also decreased in both muscle and kidney tissues during anoxia (by 37% and 30% for muscle and kidney, respectively). On the other hand, BGT-1 levels increased during anoxia in muscle (0.9-fold compared to control) and kidney (1.1-fold). Under 40% dehydration, NFAT5 levels decreased in liver by 53%. Aldose reductase levels also decreased by 42% in dehydrated muscle, and by 35% in dehydrated liver. In contrast, BGT-1 levels increased by 1.4-fold in dehydrated liver. SMIT levels also increased in both dehydrated muscle and liver (both by 0.8-fold).DiscussionOverall, we observed that osmoregulation through an NFAT5-mediated pathway is both tissue- and stress-specific. In both anoxia and dehydration, there appears to be a general reduction in NFAT5 levels resulting in decreased aldose reductase levels, however BGT-1 and SMIT levels still increase in certain tissues. Therefore, the regulation of osmoregulatory genes during dehydration and anoxia occurs beyond the transcriptional level, and it possibly involves RNA processing as well. These novel findings on the osmoregulatory mechanisms utilized by the wood frog advances our knowledge of osmoregulation during anoxia and dehydration. In addition, these findings highlight the importance of using this model to study molecular adaptations during stress. creator: Rasha Al-attar creator: Yichi Zhang creator: Kenneth B. Storey uri: https://doi.org/10.7717/peerj.2797 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Al-attar et al. title: Species-specific pharmacology of maximakinin, an amphibian homologue of bradykinin: putative prodrug activity at the human B2 receptor and peptidase resistance in rats link: https://peerj.com/articles/2911 last-modified: 2017-01-18 description: Maximakinin (MK), an amphibian peptide possessing the C-terminal sequence of bradykinin (BK), is a BK B2 receptor (B2R) agonist eliciting prolonged signaling. We reinvestigated this 19-mer for species-specific pharmacologic profile, in vivo confirmation of resistance to inactivation by angiotensin converting enzyme (ACE), value as a module for the design of fusion proteins that bind to the B2R in mammalian species and potential activity as a histamine releaser. Competition of the binding of [3H]BK to recombinant human myc-B2Rs in cells that express these receptors revealed that MK possessed a tenuous fraction (<0.1%) of the affinity of BK, despite being only ∼20-fold less potent than BK in a contractility assay based on the human isolated umbilical vein. These findings are reconciled by the generation of C-terminal fragments, like Lys-Gly-Pro-BK and Gly-Pro-BK, when the latent MK is incubated with human venous tissue (LC-MS), supporting activation via hydrolysis upstream of the BK sequence. At the rat recombinant myc-B2R, MK had a lesser affinity than that of BK, but with a narrower margin (6.2-fold, radioligand binding competition). Accordingly, MK (10 nM) stimulated calcium transients in cells that expressed the rat receptors, but not the human B2R. Recombinant MRGPRX2, a receptor that mediates cationic peptide-induced mast cell secretion, minimally responded by increased [Ca+2]i to MK at 10 µM. Enhanced green fluorescent protein fused to MK (EGFP-MK) labeled cells that expressed rat, but not human B2Rs. Intravenous MK induced dose-dependent hypotensive, vasodilator and tachycardic responses in anesthetized rats and the effects were antagonized by pretreatment with icatibant but not modified by pyrilamine or enalaprilat. Strong species-specific responses to the toxin-derived peptide MK and its prodrug status in the isolated human vein were evidenced. Accordingly, MK in the EGFP-MK fusion protein is a pharmacophore module that confers affinity for the rat B2R, but not for the human form of the B2R. MK is unlikely to be an efficient mast cell activator, but its resistance to inactivation by ACE was confirmed in vivo. creator: Xavier Charest-Morin creator: Hélène Bachelard creator: Melissa Jean creator: Francois Marceau uri: https://doi.org/10.7717/peerj.2911 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Charest-Morin et al. title: The wild tomato species Solanum chilense shows variation in pathogen resistance between geographically distinct populations link: https://peerj.com/articles/2910 last-modified: 2017-01-18 description: Wild tomatoes are a valuable source of disease resistance germplasm for tomato (Solanum lycopersicum) breeders. Many species are known to possess a certain degree of resistance against certain pathogens; however, evolution of resistance traits is yet poorly understood. For some species, like Solanum chilense, both differences in habitat and within species genetic diversity are very large. Here we aim to investigate the occurrence of spatially heterogeneous coevolutionary pressures between populations of S. chilense. We investigate the phenotypic differences in disease resistance within S. chilense against three common tomato pathogens (Alternaria solani, Phytophthora infestans and a Fusarium sp.) and confirm high degrees of variability in resistance properties between selected populations. Using generalised linear mixed models, we show that disease resistance does not follow the known demographic patterns of the species. Models with up to five available climatic and geographic variables are required to best describe resistance differences, confirming the complexity of factors involved in local resistance variation. We confirm that within S. chilense, resistance properties against various pathogens show a mosaic pattern and do not follow environmental patterns, indicating the strength of local pathogen pressures. Our study can form the basis for further investigations of the genetic traits involved. creator: Remco Stam creator: Daniela Scheikl creator: Aurélien Tellier uri: https://doi.org/10.7717/peerj.2910 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Stam et al. title: Neck biomechanics indicate that giant Transylvanian azhdarchid pterosaurs were short-necked arch predators link: https://peerj.com/articles/2908 last-modified: 2017-01-18 description: Azhdarchid pterosaurs include the largest animals to ever take to the skies with some species exceeding 10 metres in wingspan and 220 kg in mass. Associated skeletons show that azhdarchids were long-necked, long-jawed predators that combined a wing planform suited for soaring with limb adaptations indicative of quadrupedal terrestrial foraging. The postcranial proportions of the group have been regarded as uniform overall, irrespective of their overall size, notwithstanding suggestions that minor variation may have been present. Here, we discuss a recently discovered giant azhdarchid neck vertebra referable to Hatzegopteryx from the Maastrichtian Sebeş Formation of the Transylvanian Basin, Romania, which shows how some azhdarchids departed markedly from conventional views on their proportions. This vertebra, which we consider a cervical VII, is 240 mm long as preserved and almost as wide. Among azhdarchid cervicals, it is remarkable for the thickness of its cortex (4–6 mm along its ventral wall) and robust proportions. By comparing its dimensions to other giant azhdarchid cervicals and to the more completely known necks of smaller taxa, we argue that Hatzegopteryx had a proportionally short, stocky neck highly resistant to torsion and compression. This specimen is one of several hinting at greater disparity within Azhdarchidae than previously considered, but is the first to demonstrate such proportional differences within giant taxa. On the assumption that other aspects of Hatzegopteryx functional anatomy were similar to those of other azhdarchids, and with reference to the absence of large terrestrial predators in the Maastrichtian of Transylvania, we suggest that this pterosaur played a dominant predatory role among the unusual palaeofauna of ancient Haţeg. creator: Darren Naish creator: Mark P. Witton uri: https://doi.org/10.7717/peerj.2908 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Naish and Witton title: The helical domain of the EcoR124I motor subunit participates in ATPase activity and dsDNA translocation link: https://peerj.com/articles/2887 last-modified: 2017-01-18 description: Type I restriction-modification enzymes are multisubunit, multifunctional molecular machines that recognize specific DNA target sequences, and their multisubunit organization underlies their multifunctionality. EcoR124I is the archetype of Type I restriction-modification family IC and is composed of three subunit types: HsdS, HsdM, and HsdR. DNA cleavage and ATP-dependent DNA translocation activities are housed in the distinct domains of the endonuclease/motor subunit HsdR. Because the multiple functions are integrated in this large subunit of 1,038 residues, a large number of interdomain contacts might be expected. The crystal structure of EcoR124I HsdR reveals a surprisingly sparse number of contacts between helicase domain 2 and the C-terminal helical domain that is thought to be involved in assembly with HsdM. Only two potential hydrogen-bonding contacts are found in a very small contact region. In the present work, the relevance of these two potential hydrogen-bonding interactions for the multiple activities of EcoR124I is evaluated by analysing mutant enzymes using in vivo and in vitro experiments. Molecular dynamics simulations are employed to provide structural interpretation of the functional data. The results indicate that the helical C-terminal domain is involved in the DNA translocation, cleavage, and ATPase activities of HsdR, and a role in controlling those activities is suggested. creator: Vitali Bialevich creator: Dhiraj Sinha creator: Katsiaryna Shamayeva creator: Alena Guzanova creator: David Řeha creator: Eva Csefalvay creator: Jannette Carey creator: Marie Weiserova creator: Rüdiger H. Ettrich uri: https://doi.org/10.7717/peerj.2887 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Bialevich et al. title: Jaguar interactions with pumas and prey at the northern edge of jaguars’ range link: https://peerj.com/articles/2886 last-modified: 2017-01-18 description: We present the first study that evaluates jaguar-puma interactions in the arid lands of northern Mexico, where jaguars have their northernmost breeding population and both predators are persecuted for livestock depredation. We tested whether jaguars are the dominant species in this unique ecosystem, where: (1) pumas outnumber jaguars, (2) pumas are better adapted to arid environments, and (3) jaguars and pumas are of similar size. We analyzed four years of data with two approaches; a two species conditional occupancy model and an activity patterns analysis. We used camera location and prey presence as covariates for jaguar and puma detection and presence probabilities. We also explored overlap in activities of predators and prey. Where both species were detected, peccary presence was positively correlated with both jaguar and puma presence, whereas in areas where jaguars were detected but pumas were not, deer presence explained the probability of jaguar presence. We found that both predators were more likely to co-occur together than to be found independently, and so we rejected the hypothesis that jaguars were the dominant species in our study area. Predators were mainly nocturnal and their activity patterns overlapped by 60%. Jaguar, as compared with puma, overlapped more with deer and calves; puma overlapped with calves more than with other prey, suggesting a preference. We believe exploring predator relationships at different scales may help elucidate mechanisms that regulate their coexistence. creator: Carmina E. Gutiérrez-González creator: Carlos A. López-González uri: https://doi.org/10.7717/peerj.2886 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Gutiérrez-González and López-González title: A large-scale perspective for managing prairie avifauna assemblages across the western US: influences of habitat, land ownership and latitude link: https://peerj.com/articles/2879 last-modified: 2017-01-18 description: Future demands for increased food production are expected to have severe impacts on prairie biodiversity and ecosystem integrity. Prairie avifauna of North America have experienced drastic population declines, prompting numerous conservation efforts, which have been informed primarily by small-scale studies. We applied a large-scale perspective that integrates scale dependency in avian responses by analyzing observations of 20 prairie bird species (17 grassland obligates and three sagebrush obligate species) from 2009–2012 in the western prairie region of the United States. We employed a multi-species model approach to examine the relationship of land ownership, habitat, and latitude to landscape-scale species richness. Our findings suggest that patterns and processes influencing avian assemblages at the focal-scale (e.g., inference at the sampling unit) may not function at the landscape-scale (e.g., inference amongst sampling units). Individual species responses to land ownership, habitat and latitude were highly variable. The broad spatial extent of our study demonstrates the need to include lands in private ownership to assess biodiversity and the importance of maintaining habitat diversity to support avian assemblages. Lastly, focal-scale information can document species presence within a study area, but landscape-scale information provides an essential complement to inform conservation actions and policies by placing local biodiversity in the context of an entire region, landscape or ecosystem. creator: Victoria J. Dreitz creator: Lani T. Stinson creator: Beth A. Hahn creator: Jason D. Tack creator: Paul M. Lukacs uri: https://doi.org/10.7717/peerj.2879 license: http://creativecommons.org/publicdomain/zero/1.0/ rights: title: Skeletal pathology and variable anatomy in elephant feet assessed using computed tomography link: https://peerj.com/articles/2877 last-modified: 2017-01-18 description: Foot problems are a major cause of morbidity and mortality in elephants, but are underreported due to difficulties in diagnosis, particularly of conditions affecting the bones and internal structures. Here we evaluate post-mortem computer tomographic (CT) scans of 52 feet from 21 elephants (seven African Loxodonta africana and 14 Asian Elephas maximus), describing both pathology and variant anatomy (including the appearance of phalangeal and sesamoid bones) that could be mistaken for disease. We found all the elephants in our study to have pathology of some type in at least one foot. The most common pathological changes observed were bone remodelling, enthesopathy, osseous cyst-like lesions, and osteoarthritis, with soft tissue mineralisation, osteitis, infectious osteoarthriti, subluxation, fracture and enostoses observed less frequently. Most feet had multiple categories of pathological change (81% with two or more diagnoses, versus 10% with a single diagnosis, and 9% without significant pathology). Much of the pathological change was focused over the middle/lateral digits, which bear most weight and experience high peak pressures during walking. We found remodelling and osteoarthritis to be correlated with increasing age, more enthesopathy in Asian elephants, and more cyst-like lesions in females. We also observed multipartite, missing and misshapen phalanges as common and apparently incidental findings. The proximal (paired) sesamoids can appear fused or absent, and the predigits (radial/tibial sesamoids) can be variably ossified, though are significantly more ossified in Asian elephants. Our study reinforces the need for regular examination and radiography of elephant feet to monitor for pathology and as a tool for improving welfare. creator: Sophie Regnault creator: Jonathon J.I. Dixon creator: Chris Warren-Smith creator: John R. Hutchinson creator: Renate Weller uri: https://doi.org/10.7717/peerj.2877 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Regnault et al. title: Comparison of different sample preparation methods for platinum determination in cultured cells by graphite furnace atomic absorption spectrometry link: https://peerj.com/articles/2873 last-modified: 2017-01-18 description: BackgroundPlatinum-based agents are widely used in chemotherapy against solid tumors and insufficient intracellular drug accumulation is one of the leading causes of platinum resistance which is associated with poor survival of tumor patients. Thus, the detection of intracellular platinum is pivotal for studies aiming to overcome platinum resistance. In the present study, we aimed to establish a reliable graphite furnace atomic absorption spectrometry (GFAAS)-based assay to quantify the intracellular platinum content for cultured cells.MethodsSeveral most commonly applied cell preparation methods, including 0.2% HNO3, 0.2% Triton X-100, concentrated nitric acid, RIPA combined with concentrated nitric acid and hydroxide, followed by GFAAS for platinum detection were compared in ovarian, cervical and liver cancer cell lines to obtain the optimal one, and parameters regarding linearity, accuracy, precision and sensitivity were evaluated. Influence of other metals on platinum detection and the storage conditions of samples were also determined.ResultsThe treatment of cells with 0.2% HNO3 was superior to other approaches with fewer platinum loss and better repeatability. The recovery rate and precision of this method were 97.3%–103.0% and 1.4%–3.8%, respectively. The average recoveries in the presence of other metals were 95.1%–103.1%. The detection limit was 13.23 ug/L. The recovery rate of platinum remained acceptable even in cell samples stored in −20 °C or −80 °C for two months.DiscussionAfter comparison, we found that 0.2% HNO3 was optimal for intracellular platinum quantification based on GFAAS, which presented values compatible with that of inductively-coupled plasma mass-spectrometry (ICP-MS), and this is partially attributed to the simplicity of this method. Moreover, the assay was proved to be accurate, sensitive, cost-effective and suitable for the research of platinum-based antitumor therapy. creator: Man Xiao creator: Zaiju Huang creator: Jing Cai creator: Jinghui Jia creator: Yuzeng Zhang creator: Weihong Dong creator: Zehua Wang uri: https://doi.org/10.7717/peerj.2873 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Xiao et al. title: Symbiodinium spp. associated with scleractinian corals from Dongsha Atoll (Pratas), Taiwan, in the South China Sea link: https://peerj.com/articles/2871 last-modified: 2017-01-18 description: Dongsha Atoll (also known as Pratas) in Taiwan is the northernmost atoll in the South China Sea and a designated marine national park since 2007. The marine park’s scope of protection covers the bio-resources of its waters in addition to uplands, so it is important to have data logging information and analyses of marine flora and fauna, including their physiology, ecology, and genetics. As part of this effort, we investigated Symbiodinium associations in scleractinian corals from Dongsha Atoll through surveys carried out at two depth ranges (shallow, 1–5 m; and deep, 10–15 m) in 2009 and during a bleaching event in 2010. Symbiodinium composition was assessed using restriction fragment length polymorphism (RFLP) of 28S nuclear large subunit ribosomal DNA (nlsrDNA). Our results showed that the 796 coral samples from seven families and 20 genera collected in 2009 and 132 coral samples from seven families and 12 genera collected in 2010 were associated with Symbiodinium C, D and C+D. Occurrence of clade D in shallow water (24.5%) was higher compared to deep (14.9%). Due to a bleaching event in 2010, up to 80% of coral species associated with Symbiodinium C underwent moderate to severe bleaching. Using the fine resolution technique of denaturing gradient gel electrophoresis (DGGE) of internal transcribed spacer 2 (ITS2) in 175 randomly selected coral samples, from 2009 and 2010, eight Symbiodinium C types and two Symbiodinium D types were detected. This study is the first baseline survey on Symbiodinium associations in the corals of Dongsha Atoll in the South China Sea, and it shows the dominance of Symbiodinium clade C in the population. creator: Shashank Keshavmurthy creator: Kuo-Hsun Tang creator: Chia-Min Hsu creator: Chai-Hsia Gan creator: Chao-Yang Kuo creator: Keryea Soong creator: Hong-Nong Chou creator: Chaolun Allen Chen uri: https://doi.org/10.7717/peerj.2871 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Keshavmurthy et al. title: A systematic review of animal predation creating pierced shells: implications for the archaeological record of the Old World link: https://peerj.com/articles/2903 last-modified: 2017-01-17 description: BackgroundThe shells of molluscs survive well in many sedimentary contexts and yield information about the diet of prehistoric humans. They also yield evidence of symbolic behaviours through their use as beads for body adornments. Researchers often analyse the location of perforations in shells to make judgements about their use as symbolic objects (e.g., beads), the assumption being that holes attributable to deliberate human behaviour are more likely to exhibit low variability in their anatomical locations, while holes attributable to natural processes yield more random perforations. However, there are non-anthropogenic factors that can cause perforations in shells and these may not be random. The aim of the study is compare the variation in holes in shells from archaeological sites from the Old World with the variation of holes in shells pierced by mollusc predators.MethodsThree hundred and sixteen scientific papers were retrieved from online databases by using keywords, (e.g., ‘shell beads’; ‘pierced shells’; ‘drilling predators’); 79 of these publications enabled us to conduct a systematic review to qualitatively assess the location of the holes in the shells described in the published articles. In turn, 54 publications were used to assess the location of the holes in the shells made by non-human predators.ResultsAlmost all archaeological sites described shells with holes in a variety of anatomical locations. High variation of hole-placement was found within the same species from the same site, as well as among sites. These results contrast with research on predatory molluscs, which tend to be more specific in where they attacked their prey. Gastropod and bivalve predators choose similar hole locations to humans.DiscussionBased on figures in the analysed articles, variation in hole-location on pierced shells from archaeological sites was similar to variation in the placement of holes created by non-human animals. Importantly, we found that some predators choose similar hole locations to humans. We discuss these findings and identify factors researchers might want to consider when interpreting shells recovered from archaeological contexts. creator: Anna Maria Kubicka creator: Zuzanna M. Rosin creator: Piotr Tryjanowski creator: Emma Nelson uri: https://doi.org/10.7717/peerj.2903 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Kubicka et al. title: Detecting communicative intent in a computerised test of joint attention link: https://peerj.com/articles/2899 last-modified: 2017-01-17 description: The successful navigation of social interactions depends on a range of cognitive faculties—including the ability to achieve joint attention with others to share information and experiences. We investigated the influence that intention monitoring processes have on gaze-following response times during joint attention. We employed a virtual reality task in which 16 healthy adults engaged in a collaborative game with a virtual partner to locate a target in a visual array. In the Search task, the virtual partner was programmed to engage in non-communicative gaze shifts in search of the target, establish eye contact, and then display a communicative gaze shift to guide the participant to the target. In the NoSearch task, the virtual partner simply established eye contact and then made a single communicative gaze shift towards the target (i.e., there were no non-communicative gaze shifts in search of the target). Thus, only the Search task required participants to monitor their partner’s communicative intent before responding to joint attention bids. We found that gaze following was significantly slower in the Search task than the NoSearch task. However, the same effect on response times was not observed when participants completed non-social control versions of the Search and NoSearch tasks, in which the avatar’s gaze was replaced by arrow cues. These data demonstrate that the intention monitoring processes involved in differentiating communicative and non-communicative gaze shifts during the Search task had a measurable influence on subsequent joint attention behaviour. The empirical and methodological implications of these findings for the fields of autism and social neuroscience will be discussed. creator: Nathan Caruana creator: Genevieve McArthur creator: Alexandra Woolgar creator: Jon Brock uri: https://doi.org/10.7717/peerj.2899 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Caruana et al. title: Effects of adults aging on word encoding in reading Chinese: evidence from disappearing text link: https://peerj.com/articles/2897 last-modified: 2017-01-17 description: The effect of aging on the process of word encoding for fixated words and words presented to the right of the fixation point during the reading of sentences in Chinese was investigated with two disappearing text experiments. The results of Experiment 1 showed that only the 40-ms onset disappearance of word n disrupted young adults’ reading performance. However, for old readers, the disappearance of word n caused disruptions until the onset time was 120 ms. The results of Experiment 2 showed that the disappearance of word n + 1 did not cause disruptions to young adults, but these conditions made old readers spend more time reading a sentence compared to the normal display condition. These results indicated a reliable aging effect on the process of word encoding when reading Chinese, and that the encoding process in the preview frame was more susceptible to normal aging compared to that in the fixation frame. We propose that sensory, cognitive, and specific factors related to the Chinese language are important contributors to these age-related differences. creator: Zhifang Liu creator: Yun Pan creator: Wen Tong creator: Nina Liu uri: https://doi.org/10.7717/peerj.2897 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Liu et al. title: A comparative in-silico analysis of autophagy proteins in ciliates link: https://peerj.com/articles/2878 last-modified: 2017-01-17 description: Autophagy serves as a turnover mechanism for the recycling of redundant and/or damaged macromolecules present in eukaryotic cells to re-use them under starvation conditions via a double-membrane structure known as autophagosome. A set of eukaryotic genes called autophagy-related genes (ATGs) orchestrate this highly elaborative process. The existence of these genes and the role they play in different eukaryotes are well-characterized. However, little is known of their role in some eukaryotes such as ciliates. Here, we report the computational analyses of ATG genes in five ciliate genomes to understand their diversity. Our results show that Oxytricha trifallax is the sole ciliate which has a conserved Atg12 conjugation system (Atg5-Atg12-Atg16). Interestingly, Oxytricha Atg16 protein includes WD repeats in addition to its N-terminal Atg16 domain as is the case in multicellular organisms. Additionally, phylogenetic analyses revealed that E2-like conjugating protein Atg10 is only present in Tetrahymena thermophila. We fail to find critical autophagy components Atg5, Atg7 and Atg8 in the parasitic ciliate Ichthyophthirius multifiliis. Contrary to previous reports, we also find that ciliate genomes do not encode typical Atg1 since all the candidate sequences lack an Atg1-specific C-terminal domain which is essential for Atg1 complex formation. Consistent with the absence of Atg1, ciliates also lack other members of the Atg1 complex. However, the presence of Atg6 in all ciliates examined here may rise the possibility that autophagosome formation could be operated through Atg6 in ciliates, since Atg6 has been shown as an alternative autophagy inducer. In conclusion, our results highlight that Atg proteins are partially conserved in ciliates. This may provide a better understanding for the autophagic destruction of the parental macronucleus, a developmental process also known as programmed nuclear death in ciliates. creator: Erhan Aslan creator: Nurçin Küçükoğlu creator: Muhittin Arslanyolu uri: https://doi.org/10.7717/peerj.2878 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Aslan et al. title: Prevalence and correlates of psychological distress among diabetes mellitus adults in the Jilin province in China: a cross-sectional study link: https://peerj.com/articles/2869 last-modified: 2017-01-17 description: BackgroundPsychological disorders are common in diabetes mellitus (DM) patients, and the aim of this study was to estimate the prevalence of psychological distress and to determine the influence factors associated with psychological distress among DM patients in the Jilin province of China.Methods and MaterialsMultistage, stratified cluster sampling was used in this cross-sectional study. The 12-item General Health Questionnaire (GHQ-12) was used to assess psychological status with the total score of ≥4 as the threshold for psychological distress.ResultsA total of 1,956 subjects with DM were included in the study. Out of this total diabetic participants, 524 (26.8%) had psychological distress. Multiple logistic regression analysis showed that low educational level, divorce or separation from one’s spouse, low family average monthly income, short sleep duration, being aware of DM status, and multiple co-morbidities are positively associated with psychological distress (all P < 0.05).ConclusionsThis study revealed a high rate of psychological distress among DM population in Jilin province. Low educational level, divorce or separation from one’s spouse, low family average monthly income, short sleep duration, awareness of DM status, and multiple co-morbidities are all associated with psychological distress among our study subjects. Interventions to control these factors are needed to address the psychological problems among diabetics in Jilin Province. creator: Shuang Qiu creator: Hongxuan Sun creator: Yawen Liu creator: Joseph Sam Kanu creator: Ri Li creator: Yaqin Yu creator: Xufeng Huang creator: Bo Li creator: Xiangyang Zhang uri: https://doi.org/10.7717/peerj.2869 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Qiu et al. title: Cry1Ac and Vip3Aa proteins from Bacillus thuringiensis targeting Cry toxin resistance in Diatraea flavipennella and Elasmopalpus lignosellus from sugarcane link: https://peerj.com/articles/2866 last-modified: 2017-01-17 description: The biological potential of Vip and Cry proteins from Bacillus is well known and widely established. Thus, it is important to look for new genes showing different modes of action, selecting those with differentiated entomotoxic activity against Diatraea flavipennella and Elasmopalpus lignosellus, which are secondary pests of sugarcane. Therefore, Cry1 and Vip3 proteins were expressed in Escherichia coli, and their toxicities were evaluated based on bioassays using neonate larvae. Of those, the most toxic were Cry1Ac and Vip3Aa considering the LC50 values. Toxins from E. coli were purified, solubilized, trypsinized, and biotinylated. Brush Border Membrane Vesicles (BBMVs) were prepared from intestines of the two species to perform homologous and heterologous competition assays. The binding assays demonstrated interactions between Cry1Aa, Cry1Ac, and Vip3Aa toxins and proteins from the BBMV of D. flavipennella and E. lignosellus. Homologous competition assays demonstrated that binding to one of the BBMV proteins was specific for each toxin. Heterologous competition assays indicated that Vip3Aa was unable to compete for Cry1Ac toxin binding. Our results suggest that Cry1Ac and Vip3Aa may have potential in future production of transgenic sugarcane for control of D. flavipennella and E. lignosellus, but more research is needed on the potential antagonism or synergism of the toxins in these pests. creator: Ana Rita Nunes Lemes creator: Camila Soares Figueiredo creator: Isis Sebastião creator: Liliane Marques da Silva creator: Rebeka da Costa Alves creator: Herbert Álvaro Abreu de Siqueira creator: Manoel Victor Franco Lemos creator: Odair Aparecido Fernandes creator: Janete Apparecida Desidério uri: https://doi.org/10.7717/peerj.2866 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Lemes et al. title: From tameness to wariness: chemical recognition of snake predators by lizards in a Mediterranean island link: https://peerj.com/articles/2828 last-modified: 2017-01-17 description: Antipredatory defenses are maintained when benefit exceeds cost. A weak predation pressure may lead insular lizards to tameness. Podarcis lilfordi exhibits a high degree of insular tameness, which may explain its extinction from the main island of Menorca when humans introduced predators. There are three species of lizards in Menorca: the native P. lilfordi, only on the surrounding islets, and two introduced lizards in the main island, Scelarcis perspicillata and Podarcis siculus. In addition, there are three species of snakes, all introduced: one non-saurophagous (Natrix maura), one potentially non-saurophagous (Rhinechis scalaris) and one saurophagous (Macroprotodon mauritanicus). We studied the reaction to snake chemical cues in five populations: (1) P. lilfordi of Colom, (2) P. lilfordi of Aire, (3) P. lilfordi of Binicodrell, (4) S. perspicillata, and (5) P. siculus, ordered by increasing level of predation pressure. The three snakes are present in the main island, while only R. scalaris is present in Colom islet, Aire and Binicodrell being snake-free islets. We aimed to assess the relationship between predation pressure and the degree of insular tameness regarding scent recognition. We hypothesized that P. lilfordi should show the highest degree of tameness, S. perspicillata should show intermediate responses, and P. siculus should show the highest wariness. Results are clear: neither P. lilfordi nor S. perspicillata recognize any of the snakes, while P. siculus recognizes the scent of M. mauritanicus and reacts to it with typical well-defined antipredatory behaviours as tail waving and slow motion. These results rise questions about the loss of chemical recognition of predators during island tameness and its related costs and benefits for lizards of insular habitats. In addition, this highlights the necessity for strong conservation measures to avoid the introduction of alien predators. creator: Abraham Mencía creator: Zaida Ortega creator: Valentín Pérez-Mellado uri: https://doi.org/10.7717/peerj.2828 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Mencía et al. title: Cryoelectrolysis—electrolytic processes in a frozen physiological saline medium link: https://peerj.com/articles/2810 last-modified: 2017-01-17 description: BackgroundCryoelectrolysis is a new minimally invasive tissue ablation surgical technique that combines the ablation techniques of electrolytic ablation with cryosurgery. The goal of this study is to examine the hypothesis that electrolysis can take place in a frozen aqueous saline solution.MethodTo examine the hypothesis we performed a cryoelectrolytic ablation protocol in which electrolysis and cryosurgery are delivered simultaneously in a tissue simulant made of physiological saline gel with a pH dye. We measured current flow, voltage and extents of freezing and pH dye staining.ResultsUsing optical measurements and measurements of currents, we have shown that electrolysis can occur in frozen physiological saline, at high subzero freezing temperatures, above the eutectic temperature of the frozen salt solution. It was observed that electrolysis occurs when the tissue resides at high subzero temperatures during the freezing stage and essentially throughout the entire thawing stage. We also found that during thawing, the frozen lesion temperature raises rapidly to high subfreezing values and remains at those values throughout the thawing stage. Substantial electrolysis occurs during the thawing stage. Another interesting finding is that electro-osmotic flows affect the process of cryoelectrolysis at the anode and cathode, in different ways.DiscussionThe results showing that electrical current flow and electrolysis occur in frozen saline solutions imply a mechanism involving ionic movement in the fluid concentrated saline solution channels between ice crystals, at high subfreezing temperatures. Temperatures higher than the eutectic are required for the brine to be fluid. The particular pattern of temperature and electrical currents during the thawing stage of frozen tissue, can be explained by the large amounts of energy that must be removed at the outer edge of the frozen lesion because of the solid/liquid phase transformation on that interface.ConclusionElectrolysis can occur in a frozen domain at high subfreezing temperature, probably above the eutectic. It appears that the most effective period for delivering electrolytic currents in cryoelectrolysis is during the high subzero temperatures stage while freezing and immediately after cooling has stopped, throughout the thawing stage. creator: Franco Lugnani creator: Matteo Macchioro creator: Boris Rubinsky uri: https://doi.org/10.7717/peerj.2810 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Lugnani et al. title: Genetic and morphological analyses indicate that the Australian endemic scorpion Urodacus yaschenkoi (Scorpiones: Urodacidae) is a species complex link: https://peerj.com/articles/2759 last-modified: 2017-01-17 description: BackgroundAustralian scorpions have received far less attention from researchers than their overseas counterparts. Here we provide the first insight into the molecular variation and evolutionary history of the endemic Australian scorpion Urodacus yaschenkoi. Also known as the inland robust scorpion, it is widely distributed throughout arid zones of the continent and is emerging as a model organism in biomedical research due to the chemical nature of its venom.MethodsWe employed Bayesian Inference (BI) methods for the phylogenetic reconstructions and divergence dating among lineages, using unique haplotype sequences from two mitochondrial loci (COXI, 16S) and one nuclear locus (28S). We also implemented two DNA taxonomy approaches (GMYC and PTP/dPTP) to evaluate the presence of cryptic species. Linear Discriminant Analysis was used to test whether the linear combination of 21 variables (ratios of morphological measurements) can predict individual’s membership to a putative species.ResultsGenetic and morphological data suggest that U. yaschenkoi is a species complex. High statistical support for the monophyly of several divergent lineages was found both at the mitochondrial loci and at a nuclear locus. The extent of mitochondrial divergence between these lineages exceeds estimates of interspecific divergence reported for other scorpion groups. The GMYC model and the PTP/bPTP approach identified major lineages and several sub-lineages as putative species. Ratios of several traits that approximate body shape had a strong predictive power (83–100%) in discriminating two major molecular lineages. A time-calibrated phylogeny dates the early divergence at the onset of continental-wide aridification in late Miocene and Pliocene, with finer-scale phylogeographic patterns emerging during the Pleistocene. This structuring dynamics is congruent with the diversification history of other fauna of the Australian arid zones.DiscussionOur results indicate that the taxonomic status of U. yaschenkoi requires revision, and we provide recommendations for such future efforts. A complex evolutionary history and extensive diversity highlights the importance of conserving U. yaschenkoi populations from different Australian arid zones in order to preserve patterns of endemism and evolutionary potential. creator: Karen Luna-Ramirez creator: Adam D. Miller creator: Gordana Rašić uri: https://doi.org/10.7717/peerj.2759 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Luna-Ramirez et al. title: A critical issue in model-based inference for studying trait-based community assembly and a solution link: https://peerj.com/articles/2885 last-modified: 2017-01-12 description: Statistical testing of trait-environment association from data is a challenge as there is no common unit of observation: the trait is observed on species, the environment on sites and the mediating abundance on species-site combinations. A number of correlation-based methods, such as the community weighted trait means method (CWM), the fourth-corner correlation method and the multivariate method RLQ, have been proposed to estimate such trait-environment associations. In these methods, valid statistical testing proceeds by performing two separate resampling tests, one site-based and the other species-based and by assessing significance by the largest of the two p-values (the pmax test). Recently, regression-based methods using generalized linear models (GLM) have been proposed as a promising alternative with statistical inference via site-based resampling. We investigated the performance of this new approach along with approaches that mimicked the pmax test using GLM instead of fourth-corner. By simulation using models with additional random variation in the species response to the environment, the site-based resampling tests using GLM are shown to have severely inflated type I error, of up to 90%, when the nominal level is set as 5%. In addition, predictive modelling of such data using site-based cross-validation very often identified trait-environment interactions that had no predictive value. The problem that we identify is not an “omitted variable bias” problem as it occurs even when the additional random variation is independent of the observed trait and environment data. Instead, it is a problem of ignoring a random effect. In the same simulations, the GLM-based pmax test controlled the type I error in all models proposed so far in this context, but still gave slightly inflated error in more complex models that included both missing (but important) traits and missing (but important) environmental variables. For screening the importance of single trait-environment combinations, the fourth-corner test is shown to give almost the same results as the GLM-based tests in far less computing time. creator: Cajo J.F. ter Braak creator: Pedro Peres-Neto creator: Stéphane Dray uri: https://doi.org/10.7717/peerj.2885 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Ter Braak et al. title: Effects of an 8-week yoga program on sustained attention and discrimination function in children with attention deficit hyperactivity disorder link: https://peerj.com/articles/2883 last-modified: 2017-01-12 description: This study investigated whether a yoga exercise intervention influenced the sustained attention and discrimination function in children with ADHD. Forty-nine participants (mean age = 10.50 years) were assigned to either a yoga exercise or a control group. Participants were given the Visual Pursuit Test and Determination Test prior to and after an eight-week exercise intervention (twice per week, 40 min per session) or a control intervention. Significant improvements in accuracy rate and reaction time of the two tests were observed over time in the exercise group compared with the control group. These findings suggest that alternative therapies such as yoga exercises can be complementary to behavioral interventions for children with attention and inhibition problems. Schools and parents of children with ADHD should consider alternatives for maximizing the opportunities that children with ADHD can engage in structured yoga  exercises. creator: Chien-Chih Chou creator: Chung-Ju Huang uri: https://doi.org/10.7717/peerj.2883 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Chou and Huang title: Resources available for autism research in the big data era: a systematic review link: https://peerj.com/articles/2880 last-modified: 2017-01-12 description: Recently, there has been a move encouraged by many stakeholders towards generating big, open data in many areas of research. One area where big, open data is particularly valuable is in research relating to complex heterogeneous disorders such as Autism Spectrum Disorder (ASD). The inconsistencies of findings and the great heterogeneity of ASD necessitate the use of big and open data to tackle important challenges such as understanding and defining the heterogeneity and potential subtypes of ASD. To this end, a number of initiatives have been established that aim to develop big and/or open data resources for autism research. In order to provide a useful data reference for autism researchers, a systematic search for ASD data resources was conducted using the Scopus database, the Google search engine, and the pages on ‘recommended repositories’ by key journals, and the findings were translated into a comprehensive list focused on ASD data. The aim of this review is to systematically search for all available ASD data resources providing the following data types: phenotypic, neuroimaging, human brain connectivity matrices, human brain statistical maps, biospecimens, and ASD participant recruitment. A total of 33 resources were found containing different types of data from varying numbers of participants. Description of the data available from each data resource, and links to each resource is provided. Moreover, key implications are addressed and underrepresented areas of data are identified. creator: Reem Al-jawahiri creator: Elizabeth Milne uri: https://doi.org/10.7717/peerj.2880 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Al-jawahiri and Milne title: Italian and Swedish adolescents: differences and associations in subjective well-being and psychological well-being link: https://peerj.com/articles/2868 last-modified: 2017-01-12 description: BackgroundOne important aspect of subjective judgments about one’s well-being (i.e., subjective well-being: life satisfaction, positive affect, and negative affect) is that cultural features, such as, nationality seem to shape cognitive judgments about the “the ideal life.” In this comparative study we examined differences in subjective well-being and psychological well-being between Italian and Swedish adolescents and tested if the relationship between the three constructs of subjective well-being (i.e., satisfaction with life, positive affect, and negative affect) and psychological well-being was moderated by the adolescents’ nationality.MethodItalian (n = 255) and Swedish (n = 277) adolescents answered to the Satisfaction with Life Scale, the Positive Affect Negative Affect Schedule, and Ryff’s Scales of Psychological Well-Being. Differences between samples were tested using a Multiple Analysis of Variance. We also conducted a multiple group analysis (Italy and Sweden) using Structural Equation Modelling to investigate the relationship between all three subjective well-being constructs and psychological well-being.ResultsItalian adolescents scored significantly higher in satisfaction with life than Swedish adolescents. Additionally, across countries, girls scored significantly higher in negative affect than boys. In both countries, all three constructs of subjective well-being were significantly associated to adolescents’ psychological well-being. Nevertheless, while the effect of the relationship between affect and psychological well-being was almost the same across countries, life satisfaction was more strongly related to psychological well-being among Swedish adolescents.ConclusionsThe present study shows that there are larger variations between these two cultures in the cognitive construct of subjective well-being than in the affective construct. Accordingly, associations between the cognitive component, not the affective component, of subjective well-being and psychological well-being differ between countries as well. creator: Danilo Garcia creator: Elisabetta Sagone creator: Maria Elvira De Caroli creator: Ali Al Nima uri: https://doi.org/10.7717/peerj.2868 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Garcia et al. title: Upregulation of NOXA by 10-Hydroxycamptothecin plays a key role in inducing fibroblasts apoptosis and reducing epidural fibrosis link: https://peerj.com/articles/2858 last-modified: 2017-01-12 description: The fibrosis that develops following laminectomy or discectomy often causes serious complications, and the proliferation of fibroblasts is thought to be the major cause of epidural fibrosis. 10-Hydroxycamptothecin (HCPT) has been proven to be efficient in preventing epidural fibrosis, but the exact mechanism is still unclear. NOXA is a significant regulator of cell apoptosis, which has been reported to be beneficial in the treatment of fibrosis. We performed a series of experiments, both in vitro and in vivo, to explore the intrinsic mechanism of HCPT that underlies the induction of apoptosis in fibroblasts, and also to investigate whether HCPT has positive effects on epidural fibrosis following laminectomy in rats. Fibroblasts were cultured in vitro and stimulated by varying concentrations of HCPT (0, 1, 2, 4 µg/ml) for various durations (0, 24, 48, 72 h); the effect of HCPT in inducing the apoptosis of fibroblasts was investigated via Western blots and TUNEL assay. Our results showed that HCPT could induce apoptosis in fibroblasts and up-regulate the expression of NOXA. Following the knockdown of NOXA in fibroblasts, the results of Western blot analysis showed that the level of apoptotic markers, such as cleaved-PARP and Bax, was decreased. The results from the TUNEL assay also showed a decreased rate of apoptosis in NOXA-knocked down fibroblasts. For the in vivo studies, we performed a laminectomy at the L1-L2 levels in rats and applied HCPT of different concentrations (0.2, 0.1, 0.05 mg/ml and saline) locally; the macroscopic histological assessment, hydroxyproline content analysis and histological staining were performed to evaluate the effect of HCPT on reducing epidural fibrosis. The TUNEL assay in epidural tissues showed that HCPT could obviously induce apoptosis in fibroblasts in a dose-dependent manner. Also, immunohistochemical staining showed that the expression of NOXA increased as the concentrations of HCPT increased. Our findings are the first to demonstrate that upregulation of NOXA by HCPT plays a key role in inducing fibroblast apoptosis and in reducing epidural fibrosis. These findings might provide a potential therapeutic target for preventing epidural fibrosis following laminectomy. creator: Jihang Dai creator: Yu Sun creator: Lianqi Yan creator: Jingcheng Wang creator: Xiaolei Li creator: Jun He uri: https://doi.org/10.7717/peerj.2858 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Dai et al. title: Phalangeal joints kinematics during ostrich (Struthio camelus) locomotion link: https://peerj.com/articles/2857 last-modified: 2017-01-12 description: The ostrich is a highly cursorial bipedal land animal with a permanently elevated metatarsophalangeal joint supported by only two toes. Although locomotor kinematics in walking and running ostriches have been examined, these studies have been largely limited to above the metatarsophalangeal joint. In this study, kinematic data of all major toe joints were collected from gaits with double support (slow walking) to running during stance period in a semi-natural setup with two selected cooperative ostriches. Statistical analyses were conducted to investigate the effect of locomotor gait on toe joint kinematics. The MTP3 and MTP4 joints exhibit the largest range of motion whereas the first phalangeal joint of the 4th toe shows the largest motion variability. The interphalangeal joints of the 3rd and 4th toes present very similar motion patterns over stance phases of slow walking and running. However, the motion patterns of the MTP3 and MTP4 joints and the vertical displacement of the metatarsophalangeal joint are significantly different during running and slow walking. Because of the biomechanical requirements, osctriches are likely to select the inverted pendulum gait at low speeds and the bouncing gait at high speeds to improve movement performance and energy economy. Interestingly, the motions of the MTP3 and MTP4 joints are highly synchronized from slow to fast locomotion. This strongly suggests that the 3rd and 4th toes really work as an “integrated system” with the 3rd toe as the main load bearing element whilst the 4th toe as the complementary load sharing element with a primary role to ensure the lateral stability of the permanently elevated metatarsophalangeal joint. creator: Rui Zhang creator: Qiaoli Ji creator: Gang Luo creator: Shuliang Xue creator: Songsong Ma creator: Jianqiao Li creator: Lei Ren uri: https://doi.org/10.7717/peerj.2857 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Zhang et al. title: Diversity and distribution of the Caddisflies (Insecta: Trichoptera) of Ecuador link: https://peerj.com/articles/2851 last-modified: 2017-01-12 description: BackgroundAquatic insects and other freshwater animals are some of the most threatened forms of life on Earth. Caddisflies (Trichoptera) are highly biodiverse in the Neotropics and occupy a wide variety of freshwater habitats. In Andean countries, including Ecuador, knowledge of the aquatic biota is limited, and there is a great need for baseline data on the species found in these countries. Here we present the first list of Trichoptera known from Ecuador, a country that harbors two global biodiversity “hotspots.”MethodsWe conducted a literature review of species previously reported from Ecuador and supplemented these data with material we collected during five recent field inventories from about 40 localities spanning both hotspots. Using species presence data for each Ecuadorian province, we calculated the CHAO 2 species estimator to obtain the minimum species richness for the country.ResultsWe recorded 310 species, including 48 new records from our own field inventories for the country. CHAO 2 calculations showed that only 54% of the species have been found. Hydroptilidae and Hydropsychidae were the most species rich families. We report the family Xiphocentronidae for the first time from Ecuador as well as several new records of genera from different families.DiscussionAs in the neighboring Andean countries of Colombia and Peru, it is common to find undescribed species of caddisflies. There are vast areas of Ecuador and the northern Andes that are completely unexplored, and we expect that hundreds of new species are yet to be discovered. creator: Blanca Ríos-Touma creator: Ralph W. Holzenthal creator: Jolanda Huisman creator: Robin Thomson creator: Ernesto Rázuri-Gonzales uri: https://doi.org/10.7717/peerj.2851 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Ríos-Touma et al. title: Why choose Random Forest to predict rare species distribution with few samples in large undersampled areas? Three Asian crane species models provide supporting evidence link: https://peerj.com/articles/2849 last-modified: 2017-01-12 description: Species distribution models (SDMs) have become an essential tool in ecology, biogeography, evolution and, more recently, in conservation biology. How to generalize species distributions in large undersampled areas, especially with few samples, is a fundamental issue of SDMs. In order to explore this issue, we used the best available presence records for the Hooded Crane (Grus monacha, n = 33), White-naped Crane (Grus vipio, n = 40), and Black-necked Crane (Grus nigricollis, n = 75) in China as three case studies, employing four powerful and commonly used machine learning algorithms to map the breeding distributions of the three species: TreeNet (Stochastic Gradient Boosting, Boosted Regression Tree Model), Random Forest, CART (Classification and Regression Tree) and Maxent (Maximum Entropy Models). In addition, we developed an ensemble forecast by averaging predicted probability of the above four models results. Commonly used model performance metrics (Area under ROC (AUC) and true skill statistic (TSS)) were employed to evaluate model accuracy. The latest satellite tracking data and compiled literature data were used as two independent testing datasets to confront model predictions. We found Random Forest demonstrated the best performance for the most assessment method, provided a better model fit to the testing data, and achieved better species range maps for each crane species in undersampled areas. Random Forest has been generally available for more than 20 years and has been known to perform extremely well in ecological predictions. However, while increasingly on the rise, its potential is still widely underused in conservation, (spatial) ecological applications and for inference. Our results show that it informs ecological and biogeographical theories as well as being suitable for conservation applications, specifically when the study area is undersampled. This method helps to save model-selection time and effort, and allows robust and rapid assessments and decisions for efficient conservation. creator: Chunrong Mi creator: Falk Huettmann creator: Yumin Guo creator: Xuesong Han creator: Lijia Wen uri: https://doi.org/10.7717/peerj.2849 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Mi et al. title: In vitro effect of direct current electrical stimulation on rat mesenchymal stem cells link: https://peerj.com/articles/2821 last-modified: 2017-01-12 description: BackgroundElectrical stimulation (ES) has been successfully used to treat bone defects clinically. Recently, both cellular and molecular approaches have demonstrated that ES can change cell behavior such as migration, proliferation and differentiation.MethodsIn the present study we exposed rat bone marrow- (BM-) and adipose tissue- (AT-) derived mesenchymal stem cells (MSCs) to direct current electrical stimulation (DC ES) and assessed temporal changes in osteogenic differentiation. We applied 100 mV/mm of DC ES for 1 h per day for three, seven and 14 days to cells cultivated in osteogenic differentiation medium and assessed viability and calcium deposition at the different time points. In addition, expression of osteogenic genes, Runx2, Osteopontin, and Col1A2 was assessed in BM- and AT-derived MSCs at the different time points.ResultsResults showed that ES changed osteogenic gene expression patterns in both BM- and AT-MSCs, and these changes differed between the two groups. In BM-MSCs, ES caused a significant increase in mRNA levels of Runx2, Osteopontin and Col1A2 at day 7, while in AT-MSCs, the increase in Runx2 and Osteopontin expression were observed after 14 days of ES.DiscussionThis study shows that rat bone marrow- and adipose tissue-derived stem cells react differently to electrical stimuli, an observation that could be important for application of electrical stimulation in tissue engineering. creator: Sahba Mobini creator: Liudmila Leppik creator: Vishnu Thottakkattumana Parameswaran creator: John Howard Barker uri: https://doi.org/10.7717/peerj.2821 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Mobini et al. title: Effects of grazing intensity and the use of veterinary medical products on dung beetle biodiversity in the sub-mountainous landscape of Central Italy link: https://peerj.com/articles/2780 last-modified: 2017-01-12 description: Grazing extensification and intensification are among the main problems affecting European grasslands. We analyze the impact of grazing intensity (low and moderate) and the use of veterinary medical products (VMPs) on the dung beetle community in the province of Pesaro-Urbino (Italy). Grazing intensity is a key factor in explaining the diversity of dung beetles. In the case of the alpha diversity component, sites with a low level of grazing activity—related in a previous step to the subsequent abandonment of traditional farming—is characterized by a loss of species richness (q = 0) and a reduction in alpha diversity at the levels q = 1 and q = 2. In the case of beta diversity, sites with a different grazing intensity show remarkable differences in terms of the composition of their species assemblages. The use of VMPs is another important factor in explaining changes in dung beetle diversity. In sites with a traditional use of VMPs, a significant loss of species richness and biomass is observed, as is a notable effect on beta diversity. In addition, the absence of indicator species in sites with a historical use of VMPs corroborates the hypothesis that these substances have a ubiquitous effect on dung beetles. However, the interaction between grazing activity and VMPs when it comes to explaining changes in dung beetle diversity is less significant (or is not significant) than the main effects (each factor separately) for alpha diversity, biomass and species composition. This may be explained if we consider that both factors affect the various species differently. In other words, the reduction in dung availability affects several larger species more than it does very small species, although this does not imply that the former are more susceptible to injury caused by the ingestion of dung contaminated with VMPs. Finally, in order to prevent negative consequences for dung beetle diversity, we propose the maintenance of a moderate grazing intensity and the rational use of VMPs. It is our view that organic management can prevent excessive extensification while providing an economic stimulus to the sector. Simultaneously, it can also prevent the abuse of VMPs. creator: Mattia Tonelli creator: José R. Verdú creator: Mario E. Zunino uri: https://doi.org/10.7717/peerj.2780 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Tonelli et al. title: Optimizing the use of a sensor resource for opponent polarization coding link: https://peerj.com/articles/2772 last-modified: 2017-01-12 description: Flies use specialized photoreceptors R7 and R8 in the dorsal rim area (DRA) to detect skylight polarization. R7 and R8 form a tiered waveguide (central rhabdomere pair, CRP) with R7 on top, filtering light delivered to R8. We examine how the division of a given resource, CRP length, between R7 and R8 affects their ability to code polarization angle. We model optical absorption to show how the length fractions allotted to R7 and R8 determine the rates at which they transduce photons, and correct these rates for transduction unit saturation. The rates give polarization signal and photon noise in R7, and in R8. Their signals are combined in an opponent unit, intrinsic noise added, and the unit’s output analysed to extract two measures of coding ability, number of discriminable polarization angles and mutual information. A very long R7 maximizes opponent signal amplitude, but codes inefficiently due to photon noise in the very short R8. Discriminability and mutual information are optimized by maximizing signal to noise ratio, SNR. At lower light levels approximately equal lengths of R7 and R8 are optimal because photon noise dominates. At higher light levels intrinsic noise comes to dominate and a shorter R8 is optimum. The optimum R8 length fractions falls to one third. This intensity dependent range of optimal length fractions corresponds to the range observed in different fly species and is not affected by transduction unit saturation. We conclude that a limited resource, rhabdom length, can be divided between two polarization sensors, R7 and R8, to optimize opponent coding. We also find that coding ability increases sub-linearly with total rhabdom length, according to the law of diminishing returns. Consequently, the specialized shorter central rhabdom in the DRA codes polarization twice as efficiently with respect to rhabdom length than the longer rhabdom used in the rest of the eye. creator: Francisco J.H. Heras creator: Simon B. Laughlin uri: https://doi.org/10.7717/peerj.2772 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Heras and Laughlin title: Handling stress may confound murine gut microbiota studies link: https://peerj.com/articles/2876 last-modified: 2017-01-11 description: BackgroundAccumulating evidence indicates interactions between human milk composition, particularly sugars (human milk oligosaccharides or HMO), the gut microbiota of human infants, and behavioral effects. Some HMO secreted in human milk are unable to be endogenously digested by the human infant but are able to be metabolized by certain species of gut microbiota, including Bifidobacterium longum subsp. infantis (B. infantis), a species sensitive to host stress (Bailey & Coe, 2004). Exposure to gut bacteria like B. infantisduring critical neurodevelopment windows in early life appears to have behavioral consequences; however, environmental, physical, and social stress during this period can also have behavioral and microbial consequences. While rodent models are a useful method for determining causal relationships between HMO, gut microbiota, and behavior, murine studies of gut microbiota usually employ oral gavage, a technique stressful to the mouse. Our aim was to develop a less-invasive technique for HMO administration to remove the potential confound of gavage stress. Under the hypothesis that stress affects gut microbiota, particularly B. infantis, we predicted the pups receiving a prebiotic solution in a less-invasive manner would have the highest amount of Bifidobacteria in their gut.MethodsThis study was designed to test two methods, active and passive, of solution administration to mice and the effects on their gut microbiome. Neonatal C57BL/6J mice housed in a specific-pathogen free facility received increasing doses of fructooligosaccharide (FOS) solution or deionized, distilled water. Gastrointestinal (GI) tracts were collected from five dams, six sires, and 41 pups over four time points. Seven fecal pellets from unhandled pups and two pellets from unhandled dams were also collected. Qualitative real-time polymerase chain reaction (qRT-PCR) was used to quantify and compare the amount of Bifidobacterium, Bacteroides, Bacteroidetes, and Firmicutes.ResultsOur results demonstrate a significant difference between the amount of Firmicutes in pups receiving water passively and those receiving FOS actively (p-value = 0.009). Additionally, we found significant differences between the fecal microbiota from handled and non-handled mouse pups.DiscussionFrom our results, we conclude even handling pups for experimental purposes, without gavage, may induce enough stress to alter the murine gut microbiota profile. We suggest further studies to examine potential stress effects on gut microbiota caused by experimental techniques. Stress from experimental techniques may need to be accounted for in future gut microbiota studies. creator: Cary R. Allen-Blevins creator: Xiaomeng You creator: Katie Hinde creator: David A. Sela uri: https://doi.org/10.7717/peerj.2876 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Allen-Blevins et al. title: Sponge exhalent seawater contains a unique chemical profile of dissolved organic matter link: https://peerj.com/articles/2870 last-modified: 2017-01-11 description: Sponges are efficient filter feeders, removing significant portions of particulate and dissolved organic matter (POM, DOM) from the water column. While the assimilation and respiration of POM and DOM by sponges and their abundant microbial symbiont communities have received much attention, there is virtually no information on the impact of sponge holobiont metabolism on the composition of DOM at a molecular-level. We applied untargeted and targeted metabolomics techniques to characterize DOM in seawater samples prior to entering the sponge (inhalant reef water), in samples exiting the sponge (exhalent seawater), and in samples collected just outside the reef area (off reef seawater). Samples were collected from two sponge species, Ircinia campana and Spheciospongia vesparium, on a near-shore hard bottom reef in the Florida Keys. Metabolic profiles generated from untargeted metabolomics analysis indicated that many more compounds were enhanced in the exhalent samples than in the inhalant samples. Targeted metabolomics analysis revealed differences in diversity and concentration of metabolites between exhalent and off reef seawater. For example, most of the nucleosides were enriched in the exhalent seawater, while the aromatic amino acids, caffeine and the nucleoside xanthosine were elevated in the off reef water samples. Although the metabolic profile of the exhalent seawater was unique, the impact of sponge metabolism on the overall reef DOM profile was spatially limited in our study. There were also no significant differences in the metabolic profiles of exhalent water between the two sponge species, potentially indicating that there is a characteristic DOM profile in the exhalent seawater of Caribbean sponges. Additional work is needed to determine whether the impact of sponge DOM is greater in habitats with higher sponge cover and diversity. This work provides the first insight into the molecular-level impact of sponge holobiont metabolism on reef DOM and establishes a foundation for future experimental studies addressing the influence of sponge-derived DOM on chemical and ecological processes in coral reef ecosystems. creator: Cara L. Fiore creator: Christopher J. Freeman creator: Elizabeth B. Kujawinski uri: https://doi.org/10.7717/peerj.2870 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Fiore et al. title: Hepatic fibrosis and factors associated with liver stiffness in HIV mono-infected individuals link: https://peerj.com/articles/2867 last-modified: 2017-01-11 description: BackgroundLiver disease has become an important cause of morbidity and mortality even in those HIV-infected individuals who are devoid of hepatitis virus co-infection. The aim of this study was to evaluate the degree of hepatic fibrosis and the role of associated factors using liver stiffness measurement in HIV mono-infected patients without significant alcohol intake.MethodsWe performed a cross-sectional study of 101 HIV mono-infected patients recruited prospectively from March 1, 2014 to October 30, 2014 at the Center for HIV, St István and St László Hospital, Budapest, Hungary. To determine hepatic fibrosis, liver stiffness was measured with transient elastography. Demographic, immunologic and other clinical parameters were collected to establish a multivariate model. Bayesian Model Averaging (BMA) was performed to identify predictors of liver stiffness.ResultsLiver stiffness ranged from 3.0–34.3 kPa, with a median value of 5.1 kPa (IQR 1.7). BMA provided a very high support for age (Posterior Effect Probability-PEP: 84.5%), moderate for BMI (PEP: 49.3%), CD4/8 ratio (PEP: 44.2%) and lipodystrophy (PEP: 44.0%). For all remaining variables, the model rather provides evidence against their effect. These results overall suggest that age and BMI have a positive association with LS, while CD4/8 ratio and lipodystrophy are negatively associated.DiscussionOur findings shed light on the possible importance of ageing, overweight and HIV-induced immune dysregulation in the development of liver fibrosis in the HIV-infected population. Nonetheless, further controlled studies are warranted to clarify causal relations. creator: Mihály Sulyok creator: Tamás Ferenci creator: Mihály Makara creator: Gábor Horváth creator: János Szlávik creator: Zsófia Rupnik creator: Luca Kormos creator: Zsuzsanna Gerlei creator: Zita Sulyok creator: István Vályi-Nagy uri: https://doi.org/10.7717/peerj.2867 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Sulyok et al. title: Rhea: a transparent and modular R pipeline for microbial profiling based on 16S rRNA gene amplicons link: https://peerj.com/articles/2836 last-modified: 2017-01-11 description: The importance of 16S rRNA gene amplicon profiles for understanding the influence of microbes in a variety of environments coupled with the steep reduction in sequencing costs led to a surge of microbial sequencing projects. The expanding crowd of scientists and clinicians wanting to make use of sequencing datasets can choose among a range of multipurpose software platforms, the use of which can be intimidating for non-expert users. Among available pipeline options for high-throughput 16S rRNA gene analysis, the R programming language and software environment for statistical computing stands out for its power and increased flexibility, and the possibility to adhere to most recent best practices and to adjust to individual project needs. Here we present the Rhea pipeline, a set of R scripts that encode a series of well-documented choices for the downstream analysis of Operational Taxonomic Units (OTUs) tables, including normalization steps, alpha- and beta-diversity analysis, taxonomic composition, statistical comparisons, and calculation of correlations. Rhea is primarily a straightforward starting point for beginners, but can also be a framework for advanced users who can modify and expand the tool. As the community standards evolve, Rhea will adapt to always represent the current state-of-the-art in microbial profiles analysis in the clear and comprehensive way allowed by the R language. Rhea scripts and documentation are freely available at https://lagkouvardos.github.io/Rhea. creator: Ilias Lagkouvardos creator: Sandra Fischer creator: Neeraj Kumar creator: Thomas Clavel uri: https://doi.org/10.7717/peerj.2836 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Lagkouvardos et al. title: Hydrogel scaffolds promote neural gene expression and structural reorganization in human astrocyte cultures link: https://peerj.com/articles/2829 last-modified: 2017-01-11 description: Biomaterial scaffolds have the potential to enhance neuronal development and regeneration. Understanding the genetic responses of astrocytes and neurons to biomaterials could facilitate the development of synthetic environments that enable the specification of neural tissue organization with engineered scaffolds. In this study, we used high throughput transcriptomic and imaging methods to determine the impact of a hydrogel, PuraMatrix™, on human glial cells in vitro. Parallel studies were undertaken with cells grown in a monolayer environment on tissue culture polystyrene. When the Normal Human Astrocyte (NHA) cell line is grown in a hydrogel matrix environment, the glial cells adopt a structural organization that resembles that of neuronal-glial cocultures, where neurons form clusters that are distinct from the surrounding glia. Statistical analysis of next generation RNA sequencing data uncovered a set of genes that are differentially expressed in the monolayer and matrix hydrogel environments. Functional analysis demonstrated that hydrogel-upregulated genes can be grouped into three broad categories: neuronal differentiation and/or neural plasticity, response to neural insult, and sensory perception. Our results demonstrate that hydrogel biomaterials have the potential to transform human glial cell identity, and may have applications in the repair of damaged brain tissue. creator: V. Bleu Knight creator: Elba E. Serrano uri: https://doi.org/10.7717/peerj.2829 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Knight and Serrano title: Aggregations and parental care in the Early Triassic basal cynodonts Galesaurus planiceps and Thrinaxodon liorhinus link: https://peerj.com/articles/2875 last-modified: 2017-01-10 description: Non-mammaliaform cynodonts gave rise to mammals but the reproductive biology of this extinct group is still poorly known. Two exceptional fossils of Galesaurus planiceps and Thrinaxodon liorhinus, consisting of juveniles closely associated with an adult, were briefly described more than 50 years ago as examples of parental care in non-mammaliaform cynodonts. However, these two Early Triassic fossils have largely been excluded from recent discussions of parental care in the fossil record. Here we re-analyse these fossils in the context of an extensive survey of other aggregations found in these two basal cynodont taxa. Our analysis revealed six other unequivocal cases of aggregations in Thrinaxodon, with examples of same-age aggregations among immature or adult individuals as well as mixed-age aggregations between subadult and adult individuals. In contrast, only one additional aggregation of Galesauruswas identified. Taking this comprehensive survey into account, the two previously described cases of parental care in Galesaurus and Thrinaxodon are substantiated. The juveniles are the smallest specimens known for each taxon, and the size difference between the adult and the two associated juveniles is the largest found for any of the aggregations. The juveniles of Thrinaxodon are approximately only 37% of the associated adult size; whereas in Galesaurus, the young are at least 60% of the associated adult size. In each case, the two juvenile individuals are similar in size, suggesting they were from the same clutch. Even though parental care was present in both Galesaurus and Thrinaxodon, intraspecific aggregations were much more common in Thrinaxodon, suggesting it regularly lived in aggregations consisting of both similar and different aged individuals. creator: Sandra C. Jasinoski creator: Fernando Abdala uri: https://doi.org/10.7717/peerj.2875 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Jasinoski and Abdala title: The fishing and natural mortality of large, piscivorous Bull Trout and Rainbow Trout in Kootenay Lake, British Columbia (2008–2013) link: https://peerj.com/articles/2874 last-modified: 2017-01-10 description: BackgroundEstimates of fishing and natural mortality are important for understanding, and ultimately managing, commercial and recreational fisheries. High reward tags with fixed station acoustic telemetry provides a promising approach to monitoring mortality rates in large lake recreational fisheries. Kootenay Lake is a large lake which supports an important recreational fishery for large Bull Trout and Rainbow Trout.MethodsBetween 2008 and 2013, 88 large (≥500 mm) Bull Trout and 149 large (≥500 mm) Rainbow Trout were marked with an acoustic transmitter and/or high reward ($100) anchor tags in Kootenay Lake. The subsequent detections and angler recaptures were analysed using a Bayesian individual state-space Cormack–Jolly–Seber (CJS) survival model with indicator variable selection.ResultsThe final CJS survival model estimated that the annual interval probability of being recaptured by an angler was 0.17 (95% CRI [0.11–0.23]) for Bull Trout and 0.14 (95% CRI [0.09–0.19]) for Rainbow Trout. The annual interval survival probability for Bull Trout was estimated to have declined from 0.91 (95% CRI [0.76–0.97]) in 2009 to just 0.46 (95% CRI [0.24–0.76]) in 2013. Rainbow Trout survival was most strongly affected by spawning. The annual interval survival probability was 0.77 (95% CRI [0.68–0.85]) for a non-spawning Rainbow Trout compared to 0.41 (95% CRI [0.30–0.53]) for a spawner. The probability of spawning increased with the fork length for both species and decreased over the course of the study for Rainbow Trout.DiscussionFishing mortality was relatively low and constant while natural mortality was relatively high and variable. The results indicate that angler effort is not the primary driver of short-term population fluctations in the Rainbow Trout abundance. Variation in the probability of Rainbow Trout spawning suggests that the spring escapement at the outflow of Trout Lake may be a less reliable index of abundance than previously assumed. Multi-species stock assessment models need to account for the fact that large Bull Trout are more abundant than large Rainbow Trout in Kootenay Lake. creator: Joseph L. Thorley creator: Greg F. Andrusak uri: https://doi.org/10.7717/peerj.2874 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Thorley and Andrusak title: Anatomical mechanism of spontaneous recovery in regions caudal to thoracic spinal cord injury lesions in rats link: https://peerj.com/articles/2865 last-modified: 2017-01-10 description: BackgroundThe nerve fibre circuits around a lesion play a major role in the spontaneous recovery process after spinal cord hemisection in rats. The aim of the present study was to answer the following question: in the re-control process, do all spinal cord nerves below the lesion site participate, or do the spinal cord nerves of only one vertebral segment have a role in repair?MethodsFirst we made a T7 spinal cord hemisection in 50 rats. Eight weeks later, they were divided into three groups based on distinct second operations at T7: ipsilateral hemisection operation, contralateral hemisection, or transection. We then tested recovery of hindlimbs for another eight weeks. The first step was to confirm the lesion had role or not in the spontaneous recovery process. Secondly, we performed T7 spinal cord hemisections in 125 rats. Eight weeks later, we performed a second single hemisection on the ipsilateral side at T8–T12 and then tested hindlimb recovery for another six weeks.ResultsIn the first part, the Basso, Beattie, Bresnahan (BBB) scores and the electrophysiology tests of both hindlimbs weren’t significantly different after the second hemisection of the ipsilateral side. In the second part, the closer the second hemisection was to T12, the more substantial the resulting impairment in BBB score tests and prolonged latency periods.ConclusionsThe nerve regeneration from the lesion area after hemisection has no effect on spontaneous recovery of the spinal cord. Repair is carried out by all vertebrae caudal and ipsilateral to the lesion, with T12 being most important. creator: Lu-sheng Li creator: Hao Yu creator: Raynald Raynald creator: Xiao-dong Wang creator: Guang-hui Dai creator: Hong-bin Cheng creator: Xue-bin Liu creator: Yi-hua An uri: https://doi.org/10.7717/peerj.2865 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Li et al. title: Henricia djakonovi sp. nov. (Echinodermata, Echinasteridae): a new sea star species from the Sea of Japan link: https://peerj.com/articles/2863 last-modified: 2017-01-10 description: A new sea star species, H. djakonovi sp.n., was discovered in Rudnaya Bay in the Sea of Japan. This is a sympatric species of the well-known and common species Henricia pseudoleviusculaDjakonov, 1958. Both species are similar in body size and proportions, shape of skeletal plates, and life coloration, which distinguishes them from the other Henricia species inhabiting the Sea of Japan. Nevertheless, these species can be distinguished by their abactinal spines: in both species, they are short and barrel-like, but the new species is the only Henricia species in Russian waters of the Pacific that possesses such spines with a massive, smooth, bullet-like tip. The spines in H. pseudoleviuscula are crowned with a variable number of well-developed thorns. About half (<50%) of the abactinal pseudopaxillae in the new species are oval, not crescent-shaped as in H. pseudoleviuscula. creator: Anton Chichvarkhin uri: https://doi.org/10.7717/peerj.2863 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Chichvarkhin title: Interferon-γ responses to Plasmodium falciparum vaccine candidate antigens decrease in the absence of malaria transmission link: https://peerj.com/articles/2855 last-modified: 2017-01-10 description: BackgroundMalaria elimination campaigns are planned or active in many countries. The effects of malaria elimination on immune responses such as antigen-specific IFN- γ responses are not well characterized.MethodsIFN- γ responses to the P. falciparum antigens circumsporozoite protein, liver stage antigen-1, thrombospondin-related adhesive protein, apical membrane antigen-1, MB2, and merozoite surface protein-1 were tested by ELISA in 243 individuals in highland Kenya in April 2008, October 2008, and April 2009, after a one-year period of interrupted malaria transmission from April 2007 to March 2008.ResultsWhile one individual (0.4%) tested positive for P. falciparum by PCR inOctober 2008 and another two (0.9%) tested positive in April 2009, no clinical malaria cases were detected during weekly visits. Levels of IFN-γ to all antigens decreased significantly from April 2008 to April 2009 (all P < 0.001).DiscussionNaturally acquired IFN- γ responses to P. falciparum antigensare short-lived in the absence of repeated P. falciparum infection. Even short periods of malaria interruption may significantly decrease IFN-γ responses to P. falciparum antigens. creator: Cyrus Ayieko creator: Bilha S. Ogola creator: Lyticia Ochola creator: Gideon A.M. Ngwena creator: George Ayodo creator: James S. Hodges creator: Gregory S. Noland creator: Chandy C. John uri: https://doi.org/10.7717/peerj.2855 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Ayieko et al. title: Contrasting molecular and morphological evidence for the identification of an anomalous Buteo: a cautionary tale for hybrid diagnosis link: https://peerj.com/articles/2850 last-modified: 2017-01-10 description: An adult Buteo was found dead as a road-kill south of Sacramento, California, and was thought to represent the first state record of the eastern Red-shouldered Hawk (B. lineatus lineatus;). It is now a specimen in the Museum of Wildlife and Fisheries Biology (WFB 4816) at the University of California, Davis. We examined this specimen and found that many of its plumage characters differed from all other adult Red-shouldered Hawks examined, including nominate adults. Plumage markings and measurements were intermediate between Red-tailed Hawk (Buteo jamaicensis, ssp calurus) and Red-shouldered Hawk (ssp elegans), leading us to hypothesize that the bird was a hybrid. However, mtDNA sequences and nuDNA microsatellites proved definitively that the bird was a Red-shouldered Hawk, most likely of eastern origin. This case illustrates that apparent hybrids or apparent vagrants could be individuals with anomalous phenotypes caused by rare genetic variation or novel epigenetic effects. creator: William S. Clark creator: Spencer C. Galen creator: Joshua M. Hull creator: Megan A. Mayo creator: Christopher C. Witt uri: https://doi.org/10.7717/peerj.2850 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Clark et al. title: The complete chloroplast genome sequence of an endemic monotypic genus Hagenia (Rosaceae): structural comparative analysis, gene content and microsatellite detection link: https://peerj.com/articles/2846 last-modified: 2017-01-10 description: Hagenia is an endangered monotypic genus endemic to the topical mountains of Africa. The only species, Hagenia abyssinica (Bruce) J.F. Gmel, is an important medicinal plant producing bioactive compounds that have been traditionally used by African communities as a remedy for gastrointestinal ailments in both humans and animals. Complete chloroplast genomes have been applied in resolving phylogenetic relationships within plant families. We employed high-throughput sequencing technologies to determine the complete chloroplast genome sequence of H. abyssinica. The genome is a circular molecule of 154,961 base pairs (bp), with a pair of Inverted Repeats (IR) 25,971 bp each, separated by two single copies; a large (LSC, 84,320 bp) and a small single copy (SSC, 18,696). H. abyssinica’s chloroplast genome has a 37.1% GC content and encodes 112 unique genes, 78 of which code for proteins, 30 are tRNA genes and four are rRNA genes. A comparative analysis with twenty other species, sequenced to-date from the family Rosaceae, revealed similarities in structural organization, gene content and arrangement. The observed size differences are attributed to the contraction/expansion of the inverted repeats. The translational initiation factor gene (infA) which had been previously reported in other chloroplast genomes was conspicuously missing in H. abyssinica. A total of 172 microsatellites and 49 large repeat sequences were detected in the chloroplast genome. A Maximum Likelihood analyses of 71 protein-coding genes placed Hagenia in Rosoideae. The availability of a complete chloroplast genome, the first in the Sanguisorbeae tribe, is beneficial for further molecular studies on taxonomic and phylogenomic resolution within the Rosaceae family. creator: Andrew W. Gichira creator: Zhizhong Li creator: Josphat K. Saina creator: Zhicheng Long creator: Guangwan Hu creator: Robert W. Gituru creator: Qingfeng Wang creator: Jinming Chen uri: https://doi.org/10.7717/peerj.2846 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Gichira et al. title: Determining virus-host interactions and glycerol metabolism profiles in geographically diverse solar salterns with metagenomics link: https://peerj.com/articles/2844 last-modified: 2017-01-10 description: Solar salterns are excellent model ecosystems for studying virus-microbial interactions because of their low microbial diversity, environmental stability, and high viral density. By using the power of CRISPR spacers to link viruses to their prokaryotic hosts, we explored virus-host interactions in geographically diverse salterns. Using taxonomic profiling, we identified hosts such as archaeal Haloquadratum, Halorubrum, and Haloarcula and bacterial Salinibacter, and we found that community composition related to not only salinity but also local environmental dynamics. Characterizing glycerol metabolism genes in these metagenomes suggested Halorubrum and Haloquadratum possess most dihydroxyacetone kinase genes while Salinibacter possesses most glycerol-3-phosphate dehydrogenase genes. Using two different methods, we detected fewer CRISPR spacers in Haloquadratum-dominated compared with Halobacteriaceae-dominated saltern metagenomes. After CRISPR detection, spacers were aligned against haloviral genomes to map virus to host. While most alignments for each saltern metagenome linked viruses to Haloquadratum walsbyi, there were also alignments indicating interactions with the low abundance taxa Haloarcula and Haloferax. Further examination of the dinucleotide and trinucleotide usage differences between paired viruses and their hosts confirmed viruses and hosts had similar nucleotide usage signatures. Detection of cas genes in the salterns supported the possibility of CRISPR activity. Taken together, our studies suggest similar virus-host interactions exist in different solar salterns and that the glycerol metabolism gene dihydroxyacetone kinase is associated with Haloquadratum and Halorubrum. creator: Abraham G. Moller creator: Chun Liang uri: https://doi.org/10.7717/peerj.2844 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Moller and Liang title: A gene expression study of dorso-ventrally restricted pigment pattern in adult fins of Neolamprologus meeli, an African cichlid species link: https://peerj.com/articles/2843 last-modified: 2017-01-10 description: Fish color patterns are among the most diverse phenotypic traits found in the animal kingdom. Understanding the molecular and cellular mechanisms that control in chromatophore distribution and pigmentation underlying this diversity is a major goal in developmental and evolutionary biology, which has predominantly been pursued in the zebrafish model system. Here, we apply results from zebrafish work to study a naturally occurring color pattern phenotype in the fins of an African cichlid species from Lake Tanganyika. The cichlid fish Neolamprologus meeli displays a distinct dorsal color pattern, with black and white stripes along the edges of the dorsal fin and of the dorsal half of the caudal fin, corresponding with differences in melanophore density. To elucidate the molecular mechanisms controlling the differences in dorsal and ventral color patterning in the fins, we quantitatively assessed the expression of 15 candidate target genes involved in adult zebrafish pigmentation and stripe formation. For reference gene validation, we screened the expression stability of seven widely expressed genes across the investigated tissue samples and identified tbp as appropriate reference. Relative expression levels of the candidate target genes were compared between the dorsal, striped fin regions and the corresponding uniform, grey-colored regions in the anal and ventral caudal fin. Dorso-ventral expression differences, with elevated levels in both white and black stripes, were observed in two genes, the melanosome protein coding gene pmel and in igsf11, which affects melanophore adhesion, migration and survival. Next, we predicted potential shared upstream regulators of pmel and igsf11. Testing the expression patterns of six predicted transcriptions factors revealed dorso-ventral expression difference of irf1 and significant, negative expression correlation of irf1 with both pmel and igsf11. Based on these results, we propose pmel, igsf11 and irf1 as likely components of the genetic mechanism controlling distinct dorso-ventral color patterns in N. meeli fins. creator: Ehsan Pashay Ahi creator: Kristina M. Sefc uri: https://doi.org/10.7717/peerj.2843 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Ahi and Sefc title: Comprehensive transcriptome analysis provides new insights into nutritional strategies and phylogenetic relationships of chrysophytes link: https://peerj.com/articles/2832 last-modified: 2017-01-10 description: BackgroundChrysophytes are protist model species in ecology and ecophysiology and important grazers of bacteria-sized microorganisms and primary producers. However, they have not yet been investigated in detail at the molecular level, and no genomic and only little transcriptomic information is available. Chrysophytes exhibit different trophic modes: while phototrophic chrysophytes perform only photosynthesis, mixotrophs can gain carbon from bacterial food as well as from photosynthesis, and heterotrophs solely feed on bacteria-sized microorganisms. Recent phylogenies and megasystematics demonstrate an immense complexity of eukaryotic diversity with numerous transitions between phototrophic and heterotrophic organisms. The question we aim to answer is how the diverse nutritional strategies, accompanied or brought about by a reduction of the plasmid and size reduction in heterotrophic strains, affect physiology and molecular processes.ResultsWe sequenced the mRNA of 18 chrysophyte strains on the Illumina HiSeq platform and analysed the transcriptomes to determine relations between the trophic mode (mixotrophic vs. heterotrophic) and gene expression. We observed an enrichment of genes for photosynthesis, porphyrin and chlorophyll metabolism for phototrophic and mixotrophic strains that can perform photosynthesis. Genes involved in nutrient absorption, environmental information processing and various transporters (e.g., monosaccharide, peptide, lipid transporters) were present or highly expressed only in heterotrophic strains that have to sense, digest and absorb bacterial food. We furthermore present a transcriptome-based alignment-free phylogeny construction approach using transcripts assembled from short reads to determine the evolutionary relationships between the strains and the possible influence of nutritional strategies on the reconstructed phylogeny. We discuss the resulting phylogenies in comparison to those from established approaches based on ribosomal RNA and orthologous genes. Finally, we make functionally annotated reference transcriptomes of each strain available to the community, significantly enhancing publicly available data on Chrysophyceae.ConclusionsOur study is the first comprehensive transcriptomic characterisation of a diverse set of Chrysophyceaen strains. In addition, we showcase the possibility of inferring phylogenies from assembled transcriptomes using an alignment-free approach. The raw and functionally annotated data we provide will prove beneficial for further examination of the diversity within this taxon. Our molecular characterisation of different trophic modes presents a first such example. creator: Daniela Beisser creator: Nadine Graupner creator: Christina Bock creator: Sabina Wodniok creator: Lars Grossmann creator: Matthijs Vos creator: Bernd Sures creator: Sven Rahmann creator: Jens Boenigk uri: https://doi.org/10.7717/peerj.2832 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Beisser et al. title: Population dynamics of epidemic and endemic states of drug-resistance emergence in infectious diseases link: https://peerj.com/articles/2817 last-modified: 2017-01-10 description: The emergence and spread of drug-resistance during treatment of many infectious diseases continue to degrade our ability to control and mitigate infection outcomes using therapeutic measures. While the coverage and efficacy of treatment remain key factors in the population dynamics of resistance, the timing for the start of the treatment in infectious individuals can significantly influence such dynamics. We developed a between-host disease transmission model to investigate the short-term (epidemic) and long-term (endemic) states of infections caused by two competing pathogen subtypes, namely the wild-type and resistant-type, when the probability of developing resistance is a function of delay in start of the treatment. We characterize the behaviour of disease equilibria and obtain a condition to minimize the fraction of population infectious at the endemic state in terms of probability of developing resistance and its transmission fitness. For the short-term epidemic dynamics, we illustrate that depending on the likelihood of resistance development at the time of treatment initiation, the same epidemic size may be achieved with different delays in start of the treatment, which may correspond to significantly different treatment coverages. Our results demonstrate that early initiation of treatment may not necessarily be the optimal strategy for curtailing the incidence of resistance or the overall disease burden. The risk of developing drug-resistance in-host remains an important factor in the management of resistance in the population. creator: Diána Knipl creator: Gergely Röst creator: Seyed M. Moghadas uri: https://doi.org/10.7717/peerj.2817 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Knipl et al. title: CTLA-4 polymorphisms associate with breast cancer susceptibility in Asians: a meta-analysis link: https://peerj.com/articles/2815 last-modified: 2017-01-10 description: Previous studies have investigated the association between cytotoxic T-lymphocyte antigen-4 (CTLA-4) polymorphisms and breast cancer susceptibility, but the results remained inconsistent. Therefore, we evaluated the relationship between four common CTLA-4 polymorphisms and breast cancer risk by a meta-analysis, aiming to derive a comprehensive and precise conclusion. We searched EMBASE, Pubmed, Web of Science, CNKI, and Wanfang databases until July 18th, 2016. Finally, ten eligible studies involving 4,544 breast cancer patients and 4,515 cancer-free controls were included; all these studies were from Asia. Odds ratio (OR) and 95% confidence interval (CI) were used to evaluate the breast cancer risk in five genetic models. The results indicated that the CTLA-4 +49A>G (rs231775) polymorphism had a significant association with decreased breast cancer risk in allelic, homozygous, dominant and recessive models. Also, the +6230G>A (rs3087243) polymorphism reduced breast cancer risk especially in the Chinese population under homozygous and recessive models. In contrast, the −1661A>G (rs4553808) polymorphism increased breast cancer risk in allelic, heterozygous and dominant models, whereas −1722 T>C (rs733618) did not relate to breast cancer risk. In conclusion, CTLA-4 polymorphisms significantly associate with breast cancer susceptibility in Asian populations, and different gene loci may have different effects on breast cancer development. Further large-scale studies including multi-racial populations are required to confirm our findings. creator: Zhiming Dai creator: Tian Tian creator: Meng Wang creator: Xinghan Liu creator: Shuai Lin creator: Pengtao Yang creator: Kang Liu creator: Yi Zheng creator: Peng Xu creator: Meng Liu creator: Xuewen Yang creator: Zhijun Dai uri: https://doi.org/10.7717/peerj.2815 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Dai et al. title: Microbial communities mediating algal detritus turnover under anaerobic conditions link: https://peerj.com/articles/2803 last-modified: 2017-01-10 description: BackgroundAlgae encompass a wide array of photosynthetic organisms that are ubiquitously distributed in aquatic and terrestrial habitats. Algal species often bloom in aquatic ecosystems, providing a significant autochthonous carbon input to the deeper anoxic layers in stratified water bodies. In addition, various algal species have been touted as promising candidates for anaerobic biogas production from biomass. Surprisingly, in spite of its ecological and economic relevance, the microbial community involved in algal detritus turnover under anaerobic conditions remains largely unexplored.ResultsHere, we characterized the microbial communities mediating the degradation of Chlorella vulgaris (Chlorophyta), Chara sp. strain IWP1 (Charophyceae), and kelp Ascophyllum nodosum (phylum Phaeophyceae), using sediments from an anaerobic spring (Zodlteone spring, OK; ZDT), sludge from a secondary digester in a local wastewater treatment plant (Stillwater, OK; WWT), and deeper anoxic layers from a seasonally stratified lake (Grand Lake O’ the Cherokees, OK; GL) as inoculum sources. Within all enrichments, the majority of algal biomass was metabolized within 13–16 weeks, and the process was accompanied by an increase in cell numbers and a decrease in community diversity. Community surveys based on the V4 region of the 16S rRNA gene identified different lineages belonging to the phyla Bacteroidetes, Proteobacteria (alpha, delta, gamma, and epsilon classes), Spirochaetes, and Firmicutes that were selectively abundant under various substrate and inoculum conditions. Within all kelp enrichments, the microbial communities structures at the conclusion of the experiment were highly similar regardless of the enrichment source, and were dominated by the genus Clostridium, or family Veillonellaceae within the Firmicutes. In all other enrichments the final microbial community was dependent on the inoculum source, rather than the type of algae utilized as substrate. Lineages enriched included the uncultured groups VadinBC27 and WCHB1-69 within the Bacteroidetes, genus Spirochaeta and the uncultured group SHA-4 within Spirochaetes, Ruminococcaceae, Lachnospiraceae, Yongiibacter, Geosporobacter, and Acidaminobacter within the Firmicutes, and genera Kluyvera, Pantoea, Edwardsiella and Aeromonas, and Buttiauxella within the Gamma-Proteobaceteria order Enterobacteriales.ConclusionsOur results represent the first systematic survey of microbial communities mediating turnover of algal biomass under anaerobic conditions, and highlights the diversity of lineages putatively involved in the degradation process. creator: Jessica M. Morrison creator: Chelsea L. Murphy creator: Kristina Baker creator: Richard M. Zamor creator: Steve J. Nikolai creator: Shawn Wilder creator: Mostafa S. Elshahed creator: Noha H. Youssef uri: https://doi.org/10.7717/peerj.2803 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Morrison et al. title: Sharing refuges on arid islands: ecological and social influence on aggregation behaviour of wall geckos link: https://peerj.com/articles/2802 last-modified: 2017-01-10 description: BackgroundThe extent of social behaviour among reptiles is underappreciated. Two types of aggregations are recognized in lizards: ecological and social, i.e., related to the attraction to a site or to animals of the same species, respectively. As most lizards are territorial, aggregations increase the probability of aggressive interactions among individuals, a density-dependent behaviour.MethodsAfter some spurious observations of aggregation behaviour in the endemic Cabo Verde nocturnal gecko Tarentola substituta, we conducted a field-based study in order to thoroughly characterize it. We sampled 48 transects and 40 10 × 10 m quadrats on São Vicente Island to describe the incidence, size and composition of aggregations and to study the effect of gecko and refuge density, plus refuge quality, on refuge sharing. We hypothesize that when density of animals and scarcity of high-quality refuges is higher, lizards have increased probability of aggregating. We also predict a consistent pattern of size and composition of groups (male–female pairs, only one adult male per group) throughout the year if there is a selected behaviour to avoid agonistic interactions, and low thermal advantage to aggregating individuals.ResultsWe present one of the first evidences of aggregation for Phyllodactylidae geckos. We found that T. substituta forms aggregations around 30–40% of the time, and that refuges are almost always shared by a female-male pair, sometimes with a juvenile, probably a mechanism to avoid aggressive interactions. We also observed that refuge sharing is dependent on refuge quality, as medium–large (thermally more stable and positively selected) rocks are shared much more frequently than small ones, but independent of adult sizes. Refuge sharing is also directly related to the density of geckos and inversely related to the density of high-quality refuges. We found no relation between body temperatures of geckos and refuge sharing when controlling the effect of rock/air temperature, suggesting that huddling does not improve thermoregulation.DiscussionOur results suggest that in this harsh environment (rocks reach 46 °C) aggregation incidence is mainly driven by an ecological factor (scarcity of high-quality refuges) and its intersexual composition by social factors (avoidance of agonistic interactions by males, and possible increased reproductive success of the pair). This study sheds some light on the little explored gecko aggregation behaviour and other studies should follow. creator: Raquel Vasconcelos creator: Sara Rocha creator: Xavier Santos uri: https://doi.org/10.7717/peerj.2802 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Vasconcelos et al. title: Transcriptomic analysis of diplomonad parasites reveals a trans-spliced intron in a helicase gene in Giardia link: https://peerj.com/articles/2861 last-modified: 2017-01-05 description: BackgroundThe mechanisms by which DNA sequences are expressed is the central preoccupation of molecular genetics. Recently, ourselves and others reported that in the diplomonad protist Giardia lamblia, the coding regions of several mRNAs are produced by ligation of independent RNA species expressed from distinct genomic loci. Such trans-splicing of introns was found to affect nearly as many genes in this organism as does classical cis-splicing of introns. These findings raised questions about the incidence of intron trans-splicing both across the G. lambliatranscriptome and across diplomonad diversity in general, however a dearth of transcriptomic data at the time prohibited systematic study of these questions.MethodsI leverage newly available transcriptomic data from G. lamblia and the related diplomonad Spironucleus salmonicidato search for trans-spliced introns. My computational pipeline recovers all four previously reported trans-spliced introns in G. lamblia, suggesting good sensitivity.ResultsScrutiny of thousands of potential cases revealed only a single additional trans-spliced intron in G. lamblia, in the p68 helicase gene, and no cases in S. salmonicida. The p68 intron differs from the previously reported trans-spliced introns in its high degree of streamlining: the core features of G. lamblia trans-spliced introns are closely packed together, revealing striking economy in the implementation of a seemingly inherently uneconomical molecular mechanism.DiscussionThese results serve to circumscribe the role of trans-splicing in diplomonads both in terms of the number of genes effected and taxonomically. Future work should focus on the molecular mechanisms, evolutionary origins and phenotypic implications of this intriguing phenomenon. creator: Scott William Roy uri: https://doi.org/10.7717/peerj.2861 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Roy title: An updated assessment of Symbiodinium spp. that associate with common scleractinian corals from Moorea (French Polynesia) reveals high diversity among background symbionts and a novel finding of clade B link: https://peerj.com/articles/2856 last-modified: 2017-01-05 description: The adaptative bleaching hypothesis (ABH) states that, depending on the symbiotic flexibility of coral hosts (i.e., the ability of corals to “switch” or “shuffle” their algal symbionts), coral bleaching can lead to a change in the composition of their associated Symbiodinium community and, thus, contribute to the coral’s overall survival. In order to determine the flexibility of corals, molecular tools are required to provide accurate species delineations and to detect low levels of coral-associated Symbiodinium. Here, we used highly sensitive quantitative (real-time) PCR (qPCR) technology to analyse five common coral species from Moorea (French Polynesia), previously screened using only traditional molecular methods, to assess the presence of low-abundance (background) Symbiodinium spp. Similar to other studies, each coral species exhibited a strong specificity to a particular clade, irrespective of the environment. In addition, however, each of the five species harboured at least one additional Symbiodinium clade, among clades A–D, at background levels. Unexpectedly, and for the first time in French Polynesia, clade B was detected as a coral symbiont. These results increase the number of known coral-Symbiodinium associations from corals found in French Polynesia, and likely indicate an underestimation of the ability of the corals in this region to associate with and/or “shuffle” different Symbiodinium clades. Altogether our data suggest that corals from French Polynesia may favor a trade-off between optimizing symbioses with a specific Symbiodinium clade(s), maintaining associations with particular background clades that may play a role in the ability of corals to respond to environmental change. creator: Héloïse Rouzé creator: Gaël J. Lecellier creator: Denis Saulnier creator: Serge Planes creator: Yannick Gueguen creator: Herman H. Wirshing creator: Véronique Berteaux-Lecellier uri: https://doi.org/10.7717/peerj.2856 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Rouzé et al. title: miR-21 increases c-kit+ cardiac stem cell proliferation in vitro through PTEN/PI3K/Akt signaling link: https://peerj.com/articles/2859 last-modified: 2017-01-05 description: The low survival rate of cardiac stem cells (CSCs) in the ischemic myocardium is one of the obstacles in ischemic cardiomyopathy cell therapy. The MicroRNA (miR)-21 and one of its target protein, the tensin homolog deleted on chromosome ten (PTEN), contributes to the proliferation of many kinds of tissues and cell types. It is reported that miR-21 promotes proliferation through PTEN/PI3K/Akt pathway, but its effects on c-kit+ CSC remain unclear. The authors hypothesized that miR-21 promotes the proliferation in c-kit+ CSC, and evaluated the involvement of PTEN/PI3K/Akt pathway in vitro. miR-21 up-regulation with miR-21 efficiently mimics accelerated cell viability and proliferation in c-kit+ CSC, which was evidenced by the CCK-8, EdU and cell cycle analyses. In addition, the over-expression of miR-21 in c-kit+ CSCs notably down-regulated the protein expression of PTEN although the mRNA level of PTEN showed little change. Gain-of-function of miR-21 also increased the phosphor-Akt (p-Akt) level. Phen, the selective inhibitor of PTEN, reproduced the pro-proliferation effects of miR-21, while PI3K inhibitor, LY294002, totally attenuated the pro-survival effect of miR-21. These results indicate that miR-21 is efficient in promoting proliferation in c-kit+ CSCs, which is contributed by the PTEN/PI3K/Akt pathway. miR-21 holds the potential to facilitate CSC therapy in ischemic myocardium. creator: Bei Shi creator: Wenwen Deng creator: Xianping Long creator: Ranzun Zhao creator: Yan Wang creator: Wenming Chen creator: Guanxue Xu creator: Jin Sheng creator: Dongmei Wang creator: Song Cao uri: https://doi.org/10.7717/peerj.2859 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Shi et al. title: Endophytic fungal diversity of Fragaria vesca, a crop wild relative of strawberry, along environmental gradients within a small geographical area link: https://peerj.com/articles/2860 last-modified: 2017-01-05 description: BackgroundFungal endophytes are highly diverse ubiquitous asymptomatic microorganisms, some of which appear to be symbiotic. Depending on abiotic conditions and genotype of the plant, the diversity of endophytes may confer fitness benefits to plant communities.MethodsWe studied a crop wild relative (CWR) of strawberry, along environmental gradients with a view to understand the cultivable root-derived endophytic fungi that can be evaluated for promoting growth and tolerating stress in selected plant groups. The main objectives were to understand whether: (a) suboptimal soil types are drivers for fungal distribution and diversity; (b) high pH and poor nutrient availability lead to fungal-plant associations that help deliver fitness benefits; and (c) novel fungi can be identified for their use in improving plant growth, and alleviate stress in diverse crops.ResultsThe study revealed that habitats with high pH and low nutrient availability have higher fungal diversity, with more rare fungi isolated from locations with chalky soil. Plants from location G were the healthiest even though soil from this location was the poorest in nutrients. Study of environmental gradients, especially extreme habitat types, may help understand the root zone fungal diversity of different functional classes. Two small in vitro pilot studies conducted with two isolates showed that endophytic fungi from suboptimal habitats can promote plant growth and fitness benefits in selected plant groups.DiscussionTargeting native plants and crop wild relatives for research offers opportunities to unearth diverse functional groups of root-derived endophytic fungi that are beneficial for crops. creator: Kazutomo Yokoya creator: Sarah Postel creator: Rui Fang creator: Viswambharan Sarasan uri: https://doi.org/10.7717/peerj.2860 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Yokoya et al. title: Gait disorder as a predictor of spatial learning and memory impairment in aged mice link: https://peerj.com/articles/2854 last-modified: 2017-01-05 description: ObjectiveTo investigate whether gait dysfunction is a predictor of severe spatial learning and memory impairment in aged mice.MethodsA total of 100 12-month-old male mice that had no obvious abnormal motor ability and whose Morris water maze performances were not significantly different from those of two-month-old male mice were selected for the study. The selected aged mice were then divided into abnormal or normal gait groups according to the results from the quantitative gait assessment. Gaits of aged mice were defined as abnormal when the values of quantitative gait parameters were two standard deviations (SD) lower or higher than those of 2-month-old male mice. Gait parameters included stride length, variability of stride length, base of support, cadence, and average speed. After nine months, mice exhibiting severe spatial learning and memory impairment were separated from mice with mild or no cognitive dysfunction. The rate of severe spatial learning and memory impairment in the abnormal and normal gait groups was tested by a chi-square test and the correlation between gait dysfunction and decline in cognitive function was tested using a diagnostic test.ResultsThe 12-month-old aged mice were divided into a normal gait group (n = 75) and an abnormal gait group (n = 25). Nine months later, three mice in the normal gait group and two mice in the abnormal gait group had died. The remaining mice were subjected to the Morris water maze again, and 17 out of 23 mice in the abnormal gait group had developed severe spatial learning and memory impairment, including six with stride length deficits, 15 with coefficient of variation (CV) in stride length, two with base of support (BOS) deficits, five with cadence dysfunction, and six with average speed deficits. In contrast, only 15 out of 72 mice in the normal gait group developed severe spatial learning and memory impairment. The rate of severe spatial learning and memory impairment was significantly higher in the abnormal gait group as compared to that in the normal gait group (x = 21.986, P < 0.001). All five parameters used to assess gait predicted severe spatial learning and memory impairment in aged mice (P < 0.01). However, the difference of the area under the ROC (receiver operating characteristic) curve for each quantitative gait parameter was not statistically significant.ConclusionGait disorders are a predictor of severe spatial learning and memory impairment in aged mice, and stride length, variability of stride length, base of support, cadence, and average speed are all sensitive parameters for assessing gait. creator: Xin Wang creator: Qing M. Wang creator: Zhaoxiang Meng creator: Zhenglu Yin creator: Xun Luo creator: Duonan Yu uri: https://doi.org/10.7717/peerj.2854 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Wang et al. title: Identifying zooplankton community changes between shallow and upper-mesophotic reefs on the Mesoamerican Barrier Reef, Caribbean link: https://peerj.com/articles/2853 last-modified: 2017-01-05 description: Mesophotic coral ecosystems (MCEs, reefs 30–150 m) are understudied, yet the limited research conducted has been biased towards large sessile taxa, such as scleractinian corals and sponges, or mobile taxa such as fishes. Here we investigate zooplankton communities on shallow reefs and MCEs around Utila on the southern Mesoamerican Barrier Reef using planktonic light traps. Zooplankton samples were sorted into broad taxonomic groups. Our results indicate similar taxonomic zooplankton richness and overall biomass between shallow reefs and MCEs. However, the abundance of larger bodied (>2 mm) zooplanktonic groups, including decapod crab zoea, mysid shrimps and peracarid crustaceans, was higher on MCEs than shallow reefs. Our findings highlight the importance of considering zooplankton when identifying broader reef community shifts across the shallow reef to MCE depth gradient. creator: Dominic A. Andradi-Brown creator: Catherine E. I. Head creator: Dan A. Exton creator: Christina L. Hunt creator: Alicia Hendrix creator: Erika Gress creator: Alex D. Rogers uri: https://doi.org/10.7717/peerj.2853 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Andradi-Brown et al. title: Taxonomic revision and molecular phylogenetics of the Idarnes incertus species-group (Hymenoptera, Agaonidae, Sycophaginae) link: https://peerj.com/articles/2842 last-modified: 2017-01-05 description: Sycophaginae is a group of non-pollinating fig wasps considered closely related to the fig pollinators (Agaoninae, Tetrapusiinae, and Kradibiinae) in the most recent phylogenetic analyses. They occur in all tropical regions and are associated with Ficus subgenera Urostigma and Sycomorus. There are six described genera of Sycophaginae, and two are native and confined to the Neotropics, namely IdarnesWalker, 1843 and AnidarnesBouček, 1993. Genus Idarnes is divided into three morphologically distinct groups that were proven to be monophyletic by recent molecular phylogenetic analyses. In this paper we reviewed the Idarnes incertus species-group and provide detailed morphological descriptions and illustrations for the species belonging to this group. Three previously described species were redescribed: I. brasiliensis (Mayr, 1906) comb. nov., I. hansoniBouček, 1993, and I. incertus (Ashmead, 1900). Seventeen new species are described by Farache and Rasplus: I. amacayacuensissp. n., I. amazonicussp. n., I. americanaesp. n., I. badiovertexsp. n., I. brevissp. n., I. brunneussp. n., I. comptonisp. n., I. cremersiaesp. n., I. dimorphicussp. n., I. flavicrussp. n., I. flaviventrissp. n., I. gibberosussp. n., I. gordhisp. n., I. maximussp. n., I. nigriventrissp. n., I. pseudoflavussp. n. and I. ramirezisp. n. We provided keys for the identification of the species as well as for recognising the different species-groups of Idarnes and a closely related genus (SycophagaWestwood, 1840). Additionally, phylogenetic relationships among 13 species of the I. incertus species-group were inferred using four molecular markers and discussed in the light of Ficus taxonomy and host specificity. creator: Fernando H.A. Farache creator: Astrid Cruaud creator: Gwenaëlle Genson creator: Jean-Yves Rasplus creator: Rodrigo A.S. Pereira uri: https://doi.org/10.7717/peerj.2842 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Farache et al. title: Transcriptomic response of maize primary roots to low temperatures at seedling emergence link: https://peerj.com/articles/2839 last-modified: 2017-01-05 description: BackgroundMaize (Zea mays) is a C4 tropical cereal and its adaptation to temperate climates can be problematic due to low soil temperatures at early stages of establishment.MethodsIn the current study we have firstly investigated the physiological response of twelve maize varieties, from a chilling condition adapted gene pool, to sub-optimal growth temperature during seedling emergence. To identify transcriptomic markers of cold tolerance in already adapted maize genotypes, temperature conditions were set below the optimal growth range in both control and low temperature groups. The conditions were as follows; control (18 °C for 16 h and 12 °C for 8 h) and low temperature (12 °C for 16 h and 6 °C for 8 h). Four genotypes were identified from the condition adapted gene pool with significant contrasting chilling tolerance.ResultsPicker and PR39B29 were the more cold-tolerant lines and Fergus and Codisco were the less cold-tolerant lines. These four varieties were subjected to microarray analysis to identify differentially expressed genes under chilling conditions. Exposure to low temperature during establishment in the maize varieties Picker, PR39B29, Fergus and Codisco, was reflected at the transcriptomic level in the varieties Picker and PR39B29. No significant changes in expression were observed in Fergus and Codisco following chilling stress. A total number of 64 genes were differentially expressed in the two chilling tolerant varieties. These two varieties exhibited contrasting transcriptomic profiles, in which only four genes overlapped.DiscussionWe observed that maize varieties possessing an enhanced root growth ratio under low temperature were more tolerant, which could be an early and inexpensive measure for germplasm screening under controlled conditions. We have identified novel cold inducible genes in an already adapted maize breeding gene pool. This illustrates that further varietal selection for enhanced chilling tolerance is possible in an already preselected gene pool. creator: Mauro Di Fenza creator: Bridget Hogg creator: Jim Grant creator: Susanne Barth uri: https://doi.org/10.7717/peerj.2839 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Di Fenza et al. title: Simulated mussel mortality thresholds as a function of mussel biomass and nutrient loading link: https://peerj.com/articles/2838 last-modified: 2017-01-04 description: A freshwater “mussel mortality threshold” was explored as a function of porewater ammonium (NH4+) concentration, mussel biomass, and total nitrogen (N) utilizing a numerical model calibrated with data from mesocosms with and without mussels. A mortality threshold of 2 mg-N L−1 porewater NH4+ was selected based on a study that estimated 100% mortality of juvenile Lampsilis mussels exposed to 1.9 mg-N L−1 NH4+ in equilibrium with 0.18 mg-N L−1 NH3. At the highest simulated mussel biomass (560 g m−2) and the lowest simulated influent water “food” concentration (0.1 mg-N L−1), the porewater NH4+ concentration after a 2,160 h timespan without mussels was 0.5 mg-N L−1 compared to 2.25 mg-N L−1 with mussels. Continuing these simulations while varying mussel biomass and N content yielded a mortality threshold contour that was essentially linear which contradicted the non-linear and non-monotonic relationship suggested by Strayer (2014). Our model suggests that mussels spatially focus nutrients from the overlying water to the sediments as evidenced by elevated porewater NH4+ in mesocosms with mussels. However, our previous work and the model utilized here show elevated concentrations of nitrite and nitrate in overlying waters as an indirect consequence of mussel activity. Even when the simulated overlying water food availability was quite low, the mortality threshold was reached at a mussel biomass of about 480 g m−2. At a food concentration of 10 mg-N L−1, the mortality threshold was reached at a biomass of about 250 g m−2. Our model suggests the mortality threshold for juvenile Lampsilis species could be exceeded at low mussel biomass if exposed for even a short time to the highly elevated total N loadings endemic to the agricultural Midwest. creator: Jeremy S. Bril creator: Kathryn Langenfeld creator: Craig L. Just creator: Scott N. Spak creator: Teresa J. Newton uri: https://doi.org/10.7717/peerj.2838 license: http://creativecommons.org/publicdomain/zero/1.0/ rights: title: Resistance to starvation of first-stage juveniles of the Caribbean spiny lobster link: https://peerj.com/articles/2852 last-modified: 2017-01-04 description: The non-feeding postlarva (puerulus) of spiny lobsters actively swims from the open ocean to the coastal habitats where it settles and molts to the first-stage juvenile (JI). Because pueruli use much of their energy reserves swimming and preparing for the post-settlement molt, the survival of JIs presumably depends on resuming feeding as soon as possible. To test this hypothesis, the resistance to starvation of JIs of the Caribbean spiny lobster, Panulirus argus, was evaluated by measuring their point-of-no-return (PNR, minimum time of initial starvation preventing recovery after later feeding) and point-of-reserve-saturation (PRS, minimum time of initial feeding allowing for food-independent development through the rest of the molting cycle) in a warm and a cold season. Each experiment consisted of eight groups: a continuously fed control (FC) group, a continuously starved control (SC) group, and six groups subjected to differential periods of either initial starvation and subsequent feeding (PNR experiments) or initial feeding and subsequent starvation (PSR experiments). No JIs molted under continuous absence of food (SC). In both PNR experiments (temperature in warm season: 29.79 ± 0.07°C, mean ± 95% CI; in cold season: 25.63 ± 0.12°C) mortality increased sharply after 9 d of initial starvation and intermolt periods increased with period of initial starvation, but were longer in the cold season. The PNR50 was longer in the warm season (12.1 ± 1.2 d, mean ± 95% CI) than in the cold season (9.5 ± 2.1 d). In PRS experiments (temperature in warm season: 29.54 ± 0.07 °C; in cold season: 26.20 ± 0.12 °C), JIs that molted did so near the end of the feeding period; all JIs initially fed for up to 6 d succumbed, and no JIs molted after 13 d of starvation despite having fed previously. The PRS50 did not differ between the cold (13.1 ± 0.7 d) and warm seasons (12.1 ± 1.1 d). JIs of P. argus exhibit a remarkable resistance to starvation considering that the previous non-feeding, energy-demanding puerulus phase lasts for ∼3 weeks. However, JIs appear to have a relatively higher degree of dependence on food to complete development to JII during the cold season than during the warm season. Therefore, JIs of P. argus would appear to be more resistant to starvation during the warm season. creator: Alí Espinosa-Magaña creator: Enrique Lozano-Álvarez creator: Patricia Briones-Fourzán uri: https://doi.org/10.7717/peerj.2852 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Espinosa-Magaña et al. title: The outcomes of most aggressive interactions among closely related bird species are asymmetric link: https://peerj.com/articles/2847 last-modified: 2017-01-04 description: Aggressive interactions among closely related species are common, and can play an important role as a selective pressure shaping species traits and assemblages. The nature of this selective pressure depends on whether the outcomes of aggressive contests are asymmetric between species (i.e., one species is consistently dominant), yet few studies have estimated the prevalence of asymmetric versus symmetric outcomes to aggressive contests. Here we use previously published data involving 26,212 interactions between 270 species pairs of birds from 26 taxonomic families to address the question: How often are aggressive interactions among closely related bird species asymmetric? We define asymmetry using (i) the proportion of contests won by one species, and (ii) statistical tests for asymmetric outcomes of aggressive contests. We calculate these asymmetries using data summed across different sites for each species pair, and compare results to asymmetries calculated using data separated by location. We find that 80% of species pairs had aggressive outcomes where one species won 80% or more of aggressive contests. We also find that the majority of aggressive interactions among closely related species show statistically significant asymmetries, and above a sample size of 52 interactions, all outcomes are asymmetric following binomial tests. Species pairs with dominance data from multiple sites showed the same dominance relationship across locations in 93% of the species pairs. Overall, our results suggest that the outcome of aggressive interactions among closely related species are usually consistent and asymmetric, and should thus favor ecological and evolutionary strategies specific to the position of a species within a dominance hierarchy. creator: Paul R. Martin creator: Cameron Freshwater creator: Cameron K. Ghalambor uri: https://doi.org/10.7717/peerj.2847 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Martin et al. title: Transcriptome analysis of the oriental melon (Cucumis melo L. var. makuwa) during fruit development link: https://peerj.com/articles/2834 last-modified: 2017-01-04 description: BackgroundThe oriental melon (Cucumis melo L. var. makuwa) is one of the most important cultivated cucurbits grown widely in Korea, Japan, and northern China. It is cultivated because its fruit has a sweet aromatic flavor and is rich in soluble sugars, organic acids, minerals, and vitamins. In order to elucidate the genetic and molecular basis of the developmental changes that determine size, color, and sugar contents of the fruit, we performed de novo transcriptome sequencing to analyze the genes expressed during fruit development.ResultsWe identified a total of 47,666 of representative loci from 100,875 transcripts and functionally annotated 33,963 of the loci based on orthologs in Arabidopsis thaliana. Among those loci, we identified 5,173 differentially expressed genes, which were classified into 14 clusters base on the modulation of their expression patterns. The expression patterns suggested that the differentially expressed genes were related to fruit development and maturation through diverse metabolic pathways. Analyses based on gene set enrichment and the pathways described in the Kyoto Encyclopedia of Genes and Genomes suggested that the expression of genes involved in starch and sucrose metabolism and carotenoid biosynthesis were regulated dynamically during fruit development and subsequent maturation.ConclusionOur results provide the gene expression patterns related to different stages of fruit development and maturation in the oriental melon. The expression patterns give clues about important regulatory mechanisms, especially those involving starch, sugar, and carotenoid biosynthesis, in the development of the oriental melon fruit. creator: Ah-Young Shin creator: Yong-Min Kim creator: Namjin Koo creator: Su Min Lee creator: Seokhyeon Nahm creator: Suk-Yoon Kwon uri: https://doi.org/10.7717/peerj.2834 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Shin et al. title: Characterization of adipose tissue macrophages and adipose-derived stem cells in critical wounds link: https://peerj.com/articles/2824 last-modified: 2017-01-04 description: BackgroundSubcutaneous adipose tissue is a rich source of adipose tissue macrophages and adipose-derived stem cells which both play a key role in wound repair. While macrophages can be divided into the classically-activated M1 and the alternatively-activated M2 phenotype, ASCs are characterized by the expression of specific stem cell markers.MethodsIn the present study, we have investigated the expression of common macrophage polarization and stem cell markers in acutely inflamed adipose tissue. Subcutaneous adipose tissue adjacent to acutely inflamed wounds of 20 patients and 20 healthy subjects were harvested and underwent qPCR and flow cytometry analysis.ResultsExpression levels of the M1-specific markers CD80, iNOS, and IL-1b were significantly elevated in inflammatory adipose tissue when compared to healthy adipose tissue, whereas the M2-specific markers CD163 and TGF-β were decreased. By flow cytometry, a significant shift of adipose tissue macrophage populations towards the M1 phenotype was confirmed. Furthermore, a decrease in the mesenchymal stem cell markers CD29, CD34, and CD105 was observed whereas CD73 and CD90 remained unchanged.DiscussionThis is the first report describing the predominance of M1 adipose tissue macrophages and the reduction of stem cell marker expression in acutely inflamed, non-healing wounds. creator: Bong-Sung Kim creator: Pathricia V. Tilstam creator: Katrin Springenberg-Jung creator: Arne Hendrick Boecker creator: Corinna Schmitz creator: Daniel Heinrichs creator: Soo Seok Hwang creator: Jan Philipp Stromps creator: Bergita Ganse creator: Ruedger Kopp creator: Matthias Knobe creator: Juergen Bernhagen creator: Norbert Pallua creator: Richard Bucala uri: https://doi.org/10.7717/peerj.2824 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Kim et al. title: Marine turtles are not fussy nesters: a novel test of small-scale nest site selection using structure from motion beach terrain information link: https://peerj.com/articles/2770 last-modified: 2017-01-04 description: BackgroundNest selection is widely regarded as a key process determining the fitness of individuals and viability of animal populations. For marine turtles that nest on beaches, this is particularly pivotal as the nesting environment can significantly control reproductive success.The aim of this study was to identify the environmental attributes of beaches (i.e., morphology, vegetation, urbanisation) that may be associated with successful oviposition in green and loggerhead turtle nests.MethodsWe quantified the proximity of turtle nests (and surrounding beach locations) to urban areas, measured their exposure to artificial light, and used ultra-high resolution (cm-scale) digital surface models derived from Structure-from-Motion (SfM) algorithms, to characterise geomorphic and vegetation features of beaches on the Sunshine Coast, eastern Australia.ResultsAt small spatial scales (i.e., <100 m), we found no evidence that turtles selected nest sites based on a particular suite of environmental attributes (i.e., the attributes of nest sites were not consistently different from those of surrounding beach locations). Nest sites were, however, typically characterised by occurring close to vegetation, on parts of the shore where the beach- and dune-face was concave and not highly rugged, and in areas with moderate exposure to artificial light.ConclusionThis study used a novel empirical approach to identify the attributes of turtle nest sites from a broader ‘envelope’ of environmental nest traits, and is the first step towards optimizing conservation actions to mitigate, at the local scale, present and emerging human impacts on turtle nesting beaches. creator: Ilana Kelly creator: Javier X. Leon creator: Ben L. Gilby creator: Andrew D. Olds creator: Thomas A. Schlacher uri: https://doi.org/10.7717/peerj.2770 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Kelly et al. title: The acceptability of TV-based game platforms as an instrument to support the cognitive evaluation of senior adults at home link: https://peerj.com/articles/2845 last-modified: 2017-01-03 description: IntroductionThe computing capabilities of state-of-the-art television sets and media centres may facilitate the introduction of computer-assisted evaluation at home. This approach would help to overcome the drawbacks of traditional pen-and-paper evaluations administered in clinical facilities, as they could be performed in a more comfortable environment, the subject’s home, and they would be more flexible for designing complex environments for the evaluation of neuropsychological constructs that are difficult to assess through traditional testing. The objective of this work was to obtain some initial evidence about the technical acceptance by senior adults of serious games played at home on the TV set and therefore about the convenience of further investigating such an approach to cognitive assesment.Materials and MethodsWe developed a collection of games to be deployed on a TV environment. These games were tried by a group of senior adults at their homes. The Technology Acceptance Model (TAM) was used to validate this approach. Surveys were performed to study the perceived usefulness and perceived ease of use of such technical setting as an instrument for their cognitive evaluation; that is, its technical acceptance. Subjective information collected from participants was correlated with actual interaction data captured. An additional survey was performed 36 months after pilot testing to have an indication about the long-term perceptions about usefulness and ease of use.ResultsMore than 90% of participating subjects perceived cognitive games on TV as useful or very useful. The majority of participants selected the TV set as their preferred option to interact with serious games at home, when compared to other devices such as smartphones, tablets or PCs. This result correlates with the number of participants perceiving them as easily usable or very easy to use, and also with automatically captured interaction data. Three out of four seniors expressed their interest in keeping the system at home after the pilot. Besides, these perceptions are fairly stable in time as shown by the survey performed 36 months after pilot testing.LimitationsAlthough participating users are a representative sample of the Galician population, which in turn is comparable to the population of most rural areas in Europe, a larger and more diverse user sample would be needed to obtain significant results for a wider population profile.ConclusionThe study confirmed the technical acceptance, that is, the perceived usefulness and perceived ease of use, of the TV-based home technical setting introduced as a means of cognitive evaluation. This study provides initial evidence on the viability of a TV-based serious games approach for cognitive longitudinal screening at home with little intervention of clinical professionals, thus contributing to the early detection of cognitive impairments in the senior population. creator: Carlos Rivas Costa creator: Manuel José Fernández Iglesias creator: Luis Eulogio Anido Rifón creator: Miguel Gómez Carballa creator: Sonia Valladares Rodríguez uri: https://doi.org/10.7717/peerj.2845 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Rivas Costa et al. title: Impact of birth weight and postnatal diet on the gut microbiota of young adult guinea pigs link: https://peerj.com/articles/2840 last-modified: 2017-01-03 description: BackgroundThe gastrointestinal tract (GIT) microbiota is essential to metabolic health, and the prevalence of the Western diet (WD) high in fat and sugar is increasing, with evidence highlighting a negative interaction between the GIT and WD, resulting in liver dysfunction. Additionally, an adverse in utero environment such as placental insufficiency resulting in low birth weight (LBW) offspring, contributes to an increased risk of metabolic diseases such as fatty liver infiltration and liver dysfunction in later life. We sought to understand the potential interactive effects of exposure to a WD upon growing LBW offspring. We postulated that LBW offspring when challenged with a poor postnatal diet, would display an altered microbiota and more severe liver metabolic dysfunction.MethodsThe fecal microbiota of normal birth weight (NBW) and LBW young guinea pig offspring, weaned onto either a control diet (CD) or WD was determined with 16S rRNA gene next generation sequencing at young adulthood following the early rapid growth phase after weaning. A liver blood chemistry profile was also performed.ResultsThe life-long consumption of WD following weaning into young adulthood resulted in increased total cholesterol, triglycerides and alanine aminotransferase levels in association with an altered GIT microbiota when compared to offspring consuming CD. Neither birth weight nor sex were associated with any significant changes in microbiota alpha diversity, by measuring the Shannon’s diversity index. One hundred forty-eight operational taxonomic units were statistically distinct between the diet groups, independent of birth weight. In the WD group, significant decreases were detected in Barnesiella, Methanobrevibacter smithii and relatives of Oscillospira guillermondii, while Butyricimonas and Bacteroides spp. were increased.DiscussionThese results describe the GIT microbiota in a guinea pig model of LBW and WD associated metabolic syndrome and highlight several WD specific GIT alterations associated with human metabolic disease. creator: Kait Al creator: Ousseynou Sarr creator: Kristyn Dunlop creator: Gregory B. Gloor creator: Gregor Reid creator: Jeremy Burton creator: Timothy R.H. Regnault uri: https://doi.org/10.7717/peerj.2840 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Al et al. title: Metabolic phenotype of clinical and environmental Mycobacterium avium subsp. hominissuis isolates link: https://peerj.com/articles/2833 last-modified: 2017-01-03 description: BackgroundMycobacterium avium subsp. hominissuis (MAH) is an emerging opportunistic human pathogen. It can cause pulmonary infections, lymphadenitis and disseminated infections in immuno-compromised patients. In addition, MAH is widespread in the environment, since it has been isolated from water, soil or dust. In recent years, knowledge on MAH at the molecular level has increased substantially. In contrast, knowledge of the MAH metabolic phenotypes remains limited.MethodsIn this study, for the first time we analyzed the metabolic substrate utilization of ten MAH isolates, five from a clinical source and five from an environmental source. We used BIOLOG Phenotype MicroarrayTM technology for the analysis. This technology permits the rapid and global analysis of metabolic phenotypes.ResultsThe ten MAH isolates tested showed different metabolic patterns pointing to high intra-species diversity. Our MAH isolates preferred to use fatty acids such as Tween, caproic, butyric and propionic acid as a carbon source, and L-cysteine as a nitrogen source. Environmental MAH isolates resulted in being more metabolically active than clinical isolates, since the former metabolized more strongly butyric acid (p = 0.0209) and propionic acid (p = 0.00307).DiscussionOur study provides new insight into the metabolism of MAH. Understanding how bacteria utilize substrates during infection might help the developing of strategies to fight such infections. creator: Andrea Sanchini creator: Flavia Dematheis creator: Torsten Semmler creator: Astrid Lewin uri: https://doi.org/10.7717/peerj.2833 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Sanchini et al. title: Pre- versus post-exercise protein intake has similar effects on muscular adaptations link: https://peerj.com/articles/2825 last-modified: 2017-01-03 description: The purpose of this study was to test the anabolic window theory by investigating muscle strength, hypertrophy, and body composition changes in response to an equal dose of protein consumed either immediately pre- versus post-resistance training (RT) in trained men. Subjects were 21 resistance-trained men (>1 year RT experience) recruited from a university population. After baseline testing, participants were randomly assigned to 1 of 2 experimental groups: a group that consumed a supplement containing 25 g protein and 1 g carbohydrate immediately prior to exercise (PRE-SUPP) (n = 9) or a group that consumed the same supplement immediately post-exercise (POST-SUPP) (n = 12). The RT protocol consisted of three weekly sessions performed on non-consecutive days for 10 weeks. A total-body routine was employed with three sets of 8–12 repetitions for each exercise. Results showed that pre- and post-workout protein consumption had similar effects on all measures studied (p > 0.05). These findings refute the contention of a narrow post-exercise anabolic window to maximize the muscular response and instead lends support to the theory that the interval for protein intake may be as wide as several hours or perhaps more after a training bout depending on when the pre-workout meal was consumed. creator: Brad Jon Schoenfeld creator: Alan A. Aragon creator: Colin Wilborn creator: Stacie L. Urbina creator: Sara E. Hayward creator: James Krieger uri: https://doi.org/10.7717/peerj.2825 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Schoenfeld et al. title: The concurrent validity and reliability of the Leg Motion system for measuring ankle dorsiflexion range of motion in older adults link: https://peerj.com/articles/2820 last-modified: 2017-01-03 description: BackgroundNew reliable devices for range of motion (ROM) measures in older adults are necessary to improve knowledge about the functional capability in this population. Dorsiflexion ROM limitation is associated with ankle injuries, foot pain, lower limb disorders, loss of balance, gait control disorders and fall risk in older adults. The aim of the present study was to assess the validity and reliability of the Leg Motion device for measuring ankle dorsiflexion ROM in older adults.MethodsAdescriptive repeated-measures study was designed to test the reliability of Leg Motion in thirty-three healthy elderly patients older than 65 years. The subjects had to meet the following inclusion and exclusion criteria in their medical records: older than 65 years; no lower extremity injury for at least one year prior to evaluation (meniscopathy, or fractures) and any chronic injuries (e.g., osteoarthritis); no previous hip, knee or ankle surgery; no neuropathic alterations and no cognitive conditions (e.g., Alzheimer’s disease or dementia). Participants were recruited through the person responsible for the physiotherapist area from a nursing center. The subjects were evaluated in two different sessions at the same time of day, and there was a break of two weeks between sessions. To test the validity of the Leg Motion system, the participants were measured in a weight-bearing lunge position using a classic goniometer with 1° increments, a smartphone with an inclinometer standard app (iPhone 5S®) with 1° increments and a measuring tape that could measure 0.1 cm. All testing was performed while the patients were barefoot. The researcher had ten years of experience as a physiotherapist using goniometer, tape measure and inclinometer devices.ResultsMean values and standard deviations were as follows: Leg Motion (right 5.15 ± 3.08; left 5.19 ± 2.98), tape measure (right 5.12 ± 3.08; left 5.12 ± 2.80), goniometer (right 45.87° ± 4.98; left 44.50° ± 5.54) and inclinometer app (right 46.53° ± 4.79; left 45.27° ± 5.19). The paired t-test showed no significant differences between the limbs or between the test and re-test values. The test re-test reliability results for Leg Motion were as follows: the standard error of the measurement ranged from 0.29 to 0.43 cm, the minimal detectable difference ranged from 0.79 to 1.19 cm, and the intraclass correlation coefficients (ICC) values ranged from 0.97 to 0.98.ConclusionsThe results of the present study indicated that the Leg Motion device is a valid, reliable, accessible and portable tool as an alternative to the classic weight-bearing lunge test for measuring ankle dorsiflexion ROM in older adults. creator: Carlos Romero Morales creator: César Calvo Lobo creator: David Rodríguez Sanz creator: Irene Sanz Corbalán creator: Beatriz Ruiz Ruiz creator: Daniel López López uri: https://doi.org/10.7717/peerj.2820 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Romero Morales et al. title: A simple rat model of mild traumatic brain injury: a device to reproduce anatomical and neurological changes of mild traumatic brain injury link: https://peerj.com/articles/2818 last-modified: 2017-01-03 description: Mild traumatic brain injury typically involves temporary impairment of neurological function. Previous studies used water pressure or rotational injury for designing the device to make a rat a mild traumatic brain injury model. The objective of this study was to make a simple model of causing mild traumatic brain injury in rats. The device consisted of a free-fall impactor that was targeted onto the rat skull. The weight (175 g) was freely dropped 30 cm to rat’s skull bregma. We installed a safety device made of acrylic panel. To confirm a mild traumatic brain injury in 36 Sprague-Dawley rats, we performed magnetic resonance imaging (MRI) of the brain within 24 h after injury. We evaluated behavior and chemical changes in rats before and after mild traumatic brain injury. The brain MRI did not show high or low signal intensity in 34 rats. The mobility on grid floor was decreased after mild traumatic brain injury. The absolute number of foot-fault and foot-fault ratio were decreased after mild traumatic brain injury. However, the difference of the ratio was a less than absolute number of foot-fault. These results show that the device is capable of reproducing mild traumatic brain injury in rats. Our device can reduce the potential to cause brain hemorrhage and reflect the mechanism of real mild traumatic brain injury compared with existing methods and behaviors. This model can be useful in exploring physiology and management of mild traumatic brain injury. creator: Ho Jeong Kim creator: Soo Jeong Han uri: https://doi.org/10.7717/peerj.2818 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Kim and Han title: Social and environmental factors modulate leucocyte profiles in free-living Greylag geese (Anser anser) link: https://peerj.com/articles/2792 last-modified: 2017-01-03 description: BackgroundBlood parameters such as haematocrit or leucocyte counts are indicators of immune status and health, which can be affected, in a complex way, by exogenous as well as endogenous factors. Additionally, social context is known to be among the most potent stressors in group living individuals, therefore potentially influencing haematological parameters. However, with few exceptions, this potential causal relationship received only moderate scientific attention.MethodsIn a free-living and individually marked population of the highly social and long-lived Greylag goose, Anser anser, we relate variation in haematocrit (HCT), heterophils to lymphocytes ratio (H/L) and blood leucocyte counts to the following factors: intrinsic (sex, age, raising condition, i.e. goose- or hand-raised), social (pair-bond status, pair-bond duration and parental experience) and environmental (biologically relevant periods, ambient temperature) factors. Blood samples were collected repeatedly from a total of 105 focal birds during three biologically relevant seasons (winter flock, mating season, summer).ResultsWe found significant relationships between haematological parameters and social as well as environmental factors. During the mating season, unpaired individuals had higher HCT compared to paired and family individuals and this pattern reversed in fall. Similarly, H/L ratio was positively related to pair-bond status in a seasonally dependent way, with highest values during mating and successful pairs had higher H/L ratio than unsuccessful ones. Also, absolute number of leucocytes tended to vary depending on raising condition in a seasonally dependent way.DiscussionHaematology bears a great potential in ecological and behavioural studies on wild vertebrates. In sum, we found that HTC, H/L ratio and absolute number of leucocytes are modulated by social factors and conclude that they may be considered valid indicators of individual stress load. creator: Didone Frigerio creator: Sonja C. Ludwig creator: Josef Hemetsberger creator: Kurt Kotrschal creator: Claudia A.F. Wascher uri: https://doi.org/10.7717/peerj.2792 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Frigerio et al. title: Comparative analysis of the Rotarix™ vaccine strain and G1P[8] rotaviruses detected before and after vaccine introduction in Belgium link: https://peerj.com/articles/2733 last-modified: 2017-01-03 description: G1P[8] rotaviruses are responsible for the majority of human rotavirus infections worldwide. The effect of universal mass vaccination with rotavirus vaccines on circulating G1P[8] rotaviruses is still poorly understood. Therefore we analyzed the complete genomes of the Rotarix™ vaccine strain, and 70 G1P[8] rotaviruses, detected between 1999 and 2010 in Belgium (36 before and 34 after vaccine introduction) to investigate the impact of rotavirus vaccine introduction on circulating G1P[8] strains. All rotaviruses possessed a complete Wa-like genotype constellation, but frequent intra-genogroup reassortments were observed as well as multiple different cluster constellations circulating in a single season. In addition, identical cluster constellations were found to circulate persistently over multiple seasons. The Rotarix™ vaccine strain possessed a unique cluster constellation that was not present in currently circulating G1P[8] strains. At the nucleotide level, the VP6, VP2 and NSP2 gene segments of Rotarix™ were relatively distantly related to any Belgian G1P[8] strain, but other gene segments of Rotarix™ were found in clusters also containing circulating Belgian strains. At the amino acid level, the genetic distance between Rotarix™ and circulating Belgian strains was considerably lower, except for NSP1. When we compared the Belgian G1P[8] strains collected before and after vaccine introduction a reduction in the proportion of strains that were found in the same cluster as the Rotarix™ vaccine strain was observed for most gene segments. The reduction in the proportion of strains belonging to the same cluster may be the result of the vaccine introduction, although natural fluctuations cannot be ruled out. creator: Mark Zeller creator: Elisabeth Heylen creator: Sana Tamim creator: John K. McAllen creator: Ewen F. Kirkness creator: Asmik Akopov creator: Sarah De Coster creator: Marc Van Ranst creator: Jelle Matthijnssens uri: https://doi.org/10.7717/peerj.2733 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2017 Zeller et al.