title: PeerJ description: Articles published in PeerJ link: https://peerj.com/articles/index.rss3?journal=peerj&month=2016-11 creator: info@peerj.com PeerJ errorsTo: info@peerj.com PeerJ language: en title: Development and evaluation of a novel vaccine against prevalent invasive multi-drug resistant strains of Streptococcus pneumoniae link: https://peerj.com/articles/2737 last-modified: 2016-11-30 description: Streptococcus pneumoniae is a pathogen that causes serious invasive infections, such as septicemia, meningitis and pneumonia in addition to mild upper respiratory tract infections. Protection from pneumococcal diseases is thought to be mediated mainly by serotype-specific antibodies to capsular antigens. Pneumococcal conjugate vaccine consists of sugars (polysaccharides) from the capsule of the bacterium S. pneumoniae that are conjugated to a carrier protein. Three pneumococcal conjugated vaccines, each directed against a group of serotypes, are registered in Egypt; however, local vaccine production is required to cover the most prevalent serotypes. In this work, capsular polysaccharide from the most current and prevalent serotypes in Egypt were extracted, purified and conjugated to bovine serum albumin (BSA). The polysaccharide protein conjugate was purified through ultrafiltration technique and molecular size distribution was compared to an available vaccine. The immunogenicity of the prepared vaccine was examined via two methods: First, by measuring the levels of the elicited antibodies in the sera of the vaccinated mice; second, by challenging the vaccinated groups of mice with approximately 107 CFU of each specific serotype and determining the degree of protection the developled vaccine offers. Our results show that the developed conjugated capsular polysaccharide vaccine is highly immunogenic and protective in mice. This finding illustrates the importance of tracking the most recent and predominant peneumococcal serotypes to generate effective vaccines, instead of using expensive imported vaccines with large number of serotypes which might not be even present in the community. creator: Rehab H. Bahy creator: Hayam M. Hamouda creator: Amal S. Shahat creator: Aymen S. Yassin creator: Magdy A. Amin uri: https://doi.org/10.7717/peerj.2737 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Bahy et al. title: A 4D CT digital phantom of an individual human brain for perfusion analysis link: https://peerj.com/articles/2683 last-modified: 2016-11-30 description: Brain perfusion is of key importance to assess brain function. Modern CT scanners can acquire perfusion maps of the cerebral parenchyma in vivo at submillimeter resolution. These perfusion maps give insights into the hemodynamics of the cerebral parenchyma and are critical for example for treatment decisions in acute stroke. However, the relations between acquisition parameters, tissue attenuation curves, and perfusion values are still poorly understood and cannot be unraveled by studies involving humans because of ethical concerns. We present a 4D CT digital phantom specific for an individual human brain to analyze these relations in a bottom-up fashion. Validation of the signal and noise components was based on 1,000 phantom simulations of 20 patient imaging data. This framework was applied to quantitatively assess the relation between radiation dose and perfusion values, and to quantify the signal-to-noise ratios of penumbra regions with decreasing sizes in white and gray matter. This is the first 4D CT digital phantom that enables to address clinical questions without having to expose the patient to additional radiation dose. creator: Rashindra Manniesing creator: Christoph Brune creator: Bram van Ginneken creator: Mathias Prokop uri: https://doi.org/10.7717/peerj.2683 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Manniesing et al. title: Bioinformatical analysis of eukaryotic shugoshins reveals meiosis-specific features of vertebrate shugoshins link: https://peerj.com/articles/2736 last-modified: 2016-11-30 description: BackgroundShugoshins (SGOs) are proteins that protect cohesins located at the centromeres of sister chromatids from their early cleavage during mitosis and meiosis in plants, fungi, and animals. Their function is to prevent premature sister-chromatid disjunction and segregation. The study focused on the structural differences among SGOs acting during mitosis and meiosis that cause differences in chromosome behavior in these two types of cell division in different organisms.MethodsA bioinformatical analysis of protein domains, conserved amino acid motifs, and physicochemical properties of 32 proteins from 25 species of plants, fungi, and animals was performed.ResultsWe identified a C-terminal amino acid motif that is highly evolutionarily conserved among the SGOs protecting centromere cohesion of sister chromatids in meiotic anaphase I, but not among mitotic SGOs. This meiotic motif is arginine-rich in vertebrates. SGOs differ in different eukaryotic kingdoms by the sets and locations of amino acid motifs and the number of α-helical regions in the protein molecule.DiscussionThese structural differences between meiotic and mitotic SGOs probably could be responsible for the prolonged SGOs resistance to degradation during meiotic metaphase I and anaphase I. We suggest that the “arginine comb” in C-end meiotic motifs is capable of interaction by hydrogen bonds with guanine bases in the minor groove of DNA helix, thus protecting SGOs from hydrolysis. Our findings support independent evolution of meiosis in different lineages of multicellular organisms. creator: Tatiana M. Grishaeva creator: Darya Kulichenko creator: Yuri F. Bogdanov uri: https://doi.org/10.7717/peerj.2736 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Grishaeva et al. title: Effects of arachidonic acid supplementation in maturation diet on female reproductive performance and larval quality of giant river prawn (Macrobrachium rosenbergii) link: https://peerj.com/articles/2735 last-modified: 2016-11-29 description: The giant river prawn (Macrobrachium rosenbergii) is one of the most farmed freshwater crustaceans in the world. Its global production has been stalling in the past decade due to the inconsistent quality of broodstock and hatchery-produced seeds. A better understanding of the role of nutrition in maturation diets will help overcome some of the production challenges. Arachidonic acid (20:4 n-6, ARA) is a fatty acid precursor of signaling molecules important for crustacean reproduction, prostaglandins E and F of the series II (PGE2 and PGF2α), and is often lacking in maturation diets of shrimp and prawns. We examined the effects of ARA in a combination of different fish oil (FO) and soybean oil (SO) blends on females’ reproductive performance and larval quality. Adult females (15.22 ± 0.13 g and 11.12 ± 0.09 cm) were fed six isonitrogenous and isolipidic diets containing one of two different base compositions (A or B), supplemented with one of three levels of Mortierella alpine-derived ARA (containing 40% active ARA): 0, 1 or 2% by ingredient weight. The two base diets differed in the percentages of (FO and SO with diet A containing 2% SO and 2% FO and diet B containing 2.5% SO and 1.5% FO, resulting in differences in proportional contents of dietary linoleic acid (18:2n-6, LOA) and docosahexaenoic acid (22:6n-3, DHA)). After the eight-week experiment, prawns fed diet B with 1 and 2% ARA supplement (B1 and B2) exhibited the highest gonadosomatic index (GSI), hepatosomatic index (HSI), egg clutch weight, fecundity, hatching rate, number of larvae, and reproductive effort compared to those fed other diets (p ≤ 0.05). Larvae from these two dietary treatments also had higher tolerance to low salinity (2 ppt). The maturation period was not significantly different among most treatments (p ≥ 0.05). ARA supplementation, regardless of the base diet, significantly improved GSI, HSI, egg clutch weight and fecundity. However, the diets with an enhanced ARA and LOA (B1 and B2) resulted in the best reproductive performance, egg hatchability and larval tolerance to low salinity. These dietary treatments also allow for effective accumulation of ARA and an n-3 lcPUFA, DHA in eggs and larvae. creator: Chanpim Kangpanich creator: Jarunan Pratoomyot creator: Nisa Siranonthana creator: Wansuk Senanan uri: https://doi.org/10.7717/peerj.2735 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Kangpanich et al. title: The complete chloroplast genome sequence of Helwingia himalaica (Helwingiaceae, Aquifoliales) and a chloroplast phylogenomic analysis of the Campanulidae link: https://peerj.com/articles/2734 last-modified: 2016-11-29 description: Complete chloroplast genome sequences have been very useful for understanding phylogenetic relationships in angiosperms at the family level and above, but there are currently large gaps in coverage. We report the chloroplast genome for Helwingia himalaica, the first in the distinctive family Helwingiaceae and only the second genus to be sequenced in the order Aquifoliales. We then combine this with 36 published sequences in the large (c. 35,000 species) subclass Campanulidae in order to investigate relationships at the order and family levels. The Helwingia genome consists of 158,362 bp containing a pair of inverted repeat (IR) regions of 25,996 bp separated by a large single-copy (LSC) region and a small single-copy (SSC) region which are 87,810 and 18,560 bp, respectively. There are 142 known genes, including 94 protein-coding genes, eight ribosomal RNA genes, and 40 tRNA genes. The topology of the phylogenetic relationships between Apiales, Asterales, and Dipsacales differed between analyses based on complete genome sequences and on 36 shared protein-coding genes, showing that further studies of campanulid phylogeny are needed. creator: Xin Yao creator: Ying-Ying Liu creator: Yun-Hong Tan creator: Yu Song creator: Richard T. Corlett uri: https://doi.org/10.7717/peerj.2734 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Yao et al. title: The effectiveness of cooling conditions on temperature of canine EDTA whole blood samples link: https://peerj.com/articles/2732 last-modified: 2016-11-29 description: BackgroundPreanalytic factors such as time and temperature can have significant effects on laboratory test results. For example, ammonium concentration will increase 31% in blood samples stored at room temperature for 30 min before centrifugation. To reduce preanalytic error, blood samples may be placed in precooled tubes and chilled on ice or in ice water baths; however, the effectiveness of these modalities in cooling blood samples has not been formally evaluated. The purpose of this study was to evaluate the effectiveness of various cooling modalities on reducing temperature of EDTA whole blood samples.MethodsPooled samples of canine EDTA whole blood were divided into two aliquots. Saline was added to one aliquot to produce a packed cell volume (PCV) of 40% and to the second aliquot to produce a PCV of 20% (simulated anemia). Thirty samples from each aliquot were warmed to 37.7 °C and cooled in 2 ml allotments under one of three conditions: in ice, in ice after transfer to a precooled tube, or in an ice water bath. Temperature of each sample was recorded at one minute intervals for 15 min.ResultsWithin treatment conditions, sample PCV had no significant effect on cooling. Cooling in ice water was significantly faster than cooling in ice only or transferring the sample to a precooled tube and cooling it on ice. Mean temperature of samples cooled in ice water was significantly lower at 15 min than mean temperatures of those cooled in ice, whether or not the tube was precooled. By 4 min, samples cooled in an ice water bath had reached mean temperatures less than 4 °C (refrigeration temperature), while samples cooled in other conditions remained above 4.0 °C for at least 11 min. For samples with a PCV of 40%, precooling the tube had no significant effect on rate of cooling on ice. For samples with a PCV of 20%, transfer to a precooled tube resulted in a significantly faster rate of cooling than direct placement of the warmed tube onto ice.DiscussionCanine EDTA whole blood samples cool most rapidly and to a greater degree when placed in an ice-water bath rather than in ice. Samples stored on ice water can rapidly drop below normal refrigeration temperatures; this should be taken into consideration when using this cooling modality. creator: Karen M. Tobias creator: Leslie Serrano creator: Xiaocun Sun creator: Bente Flatland uri: https://doi.org/10.7717/peerj.2732 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Tobias et al. title: Disentangling urban habitat and matrix effects on wild bee species link: https://peerj.com/articles/2729 last-modified: 2016-11-29 description: In face of a dramatic decline of wild bee species in many rural landscapes, potential conservation functions of urban areas gain importance. Yet effects of urbanization on pollinators, and in particular on wild bees, remain ambiguous and not comprehensively understood. This is especially true for amenity grassland and extensively managed wastelands within large-scale residential housing areas. Using Berlin as a study region, we aimed to investigate (a) if these greenspaces are accepted by wild bee assemblages as foraging habitats; (b) how assemblage structure of bees and individual bee species are affected by different habitat (e.g., management, flower density) and urban matrix variables (e.g., isolation, urbanization); and (c) to what extent grassland restoration can promote bees in urban environments. In summer 2012, we collected 62 bee species belonging to more than 20% of the taxa known for Berlin. Urbanization significantly affected species composition of bees; 18 species were affiliated to different levels of urbanization. Most bee species were not affected by any of the environmental variables tested, and urbanization had a negative effect only for one bee species. Further, we determined that restoration of diverse grasslands positively affected bee species richnesss in urban environments. We conclude that differently structured and managed greenspaces in large-scale housing areas can provide additional foraging habitats and refuges for pollinators. This supports approaches towards a biodiversity friendly management within urban regions and may be of particular importance given that anthropogenic pressure is increasing in many rural landscapes. creator: Leonie K. Fischer creator: Julia Eichfeld creator: Ingo Kowarik creator: Sascha Buchholz uri: https://doi.org/10.7717/peerj.2729 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Fischer et al. title: Flexible ureteroscopy for renal stone without preoperative ureteral stenting shows good prognosis link: https://peerj.com/articles/2728 last-modified: 2016-11-29 description: PurposeTo clarify the outcome of flexible ureteroscopy (fURS) for management of renal calculi without preoperative stenting.MethodsA total of 171 patients who received 176 fURS procedures for unilateral renal stones were reviewed. All procedures were divided into two groups depending on whether they received ureteral stenting preoperatively. Baseline characteristics of patients, stone burden, operation time, stone-free rates, and complications were compared between both groups.ResultsSuccessful primary access to the renal pelvis was achieved in 104 of 114 (91.2%) patients without preoperative stenting, while all procedures with preoperative stenting (n = 62) were successfully performed. A total of 156 procedures were included for further data analysis (56 procedures in stenting group and 100 in non-stenting group). No significant differences was found regardless of a preoperative stent placement in terms of stone-free rate (73.2% with stenting vs. 71.0% without, P = 0.854), operative time (70.4 ± 32.8 with stenting vs. 70.2 ± 32.1 without, P = 0.969).ConclusionsfURS for management of renal stone without preoperative ureteral stenting are associated with well outcome in short term follow-up. Our study may help patients and doctors to decide if an optional stent is placed or not. creator: Jiaqiao Zhang creator: Chuou Xu creator: Deng He creator: Yuchao Lu creator: Henglong Hu creator: Baolong Qin creator: Yufeng Wang creator: Qing Wang creator: Cong Li creator: Shaogang Wang creator: Jihong Liu uri: https://doi.org/10.7717/peerj.2728 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Zhang et al. title: Quantitative analysis of rat adipose tissue cell recovery, and non-fat cell volume, in primary cell cultures link: https://peerj.com/articles/2725 last-modified: 2016-11-29 description: BackgroundWhite adipose tissue (WAT) is a complex, diffuse, multifunctional organ which contains adipocytes, and a large proportion of fat, but also other cell types, active in defense, regeneration and signalling functions. Studies with adipocytes often require their isolation from WAT by breaking up the matrix of collagen fibres; however, it is unclear to what extent adipocyte number in primary cultures correlates with their number in intact WAT, since recovery and viability are often unknown.Experimental DesignEpididymal WAT of four young adult rats was used to isolate adipocytes with collagenase. Careful recording of lipid content of tissue, and all fraction volumes and weights, allowed us to trace the amount of initial WAT fat remaining in the cell preparation. Functionality was estimated by incubation with glucose and measurement of glucose uptake and lactate, glycerol and NEFA excretion rates up to 48 h. Non-adipocyte cells were also recovered and their sizes (and those of adipocytes) were measured. The presence of non-nucleated cells (erythrocytes) was also estimated.ResultsCell numbers and sizes were correlated from all fractions to intact WAT. Tracing the lipid content, the recovery of adipocytes in the final, metabolically active, preparation was in the range of 70–75%. Cells showed even higher metabolic activity in the second than in the first day of incubation. Adipocytes were 7%, erythrocytes 66% and other stromal (nucleated cells) 27% of total WAT cells. However, their overall volumes were 90%, 0.05%, and 0.2% of WAT. Non-fat volume of adipocytes was 1.3% of WAT.ConclusionsThe methodology presented here allows for a direct quantitative reference to the original tissue of studies using isolated cells. We have also found that the “live cell mass” of adipose tissue is very small: about 13 µL/g for adipocytes and 2 µL/g stromal, plus about 1 µL/g blood (the rats were killed by exsanguination). These data translate (with respect to the actual “live cytoplasm” size) into an extremely high metabolic activity, which make WAT an even more significant agent in the control of energy metabolism. creator: Floriana Rotondo creator: María del Mar Romero creator: Ana Cecilia Ho-Palma creator: Xavier Remesar creator: José Antonio Fernández-López creator: Marià Alemany uri: https://doi.org/10.7717/peerj.2725 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Rotondo et al. title: The first complete plastid genomes of Melastomataceae are highly structurally conserved link: https://peerj.com/articles/2715 last-modified: 2016-11-29 description: BackgroundIn the past three decades, several studies have predominantly relied on a small sample of the plastome to infer deep phylogenetic relationships in the species-rich Melastomataceae. Here, we report the first full plastid sequences of this family, compare general features of the sampled plastomes to other sequenced Myrtales, and survey the plastomes for highly informative regions for phylogenetics.MethodsGenome skimming was performed for 16 species spread across the Melastomataceae. Plastomes were assembled, annotated and compared to eight sequenced plastids in the Myrtales. Phylogenetic inference was performed using Maximum Likelihood on six different data sets, where putative biases were taken into account. Summary statistics were generated for all introns and intergenic spacers with suitable size for polymerase chain reaction (PCR) amplification and used to rank the markers by phylogenetic information.ResultsThe majority of the plastomes sampled are conserved in gene content and order, as well as in sequence length and GC content within plastid regions and sequence classes. Departures include the putative presence of rps16 and rpl2 pseudogenes in some plastomes. Phylogenetic analyses of the majority of the schemes analyzed resulted in the same topology with high values of bootstrap support. Although there is still uncertainty in some relationships, in the highest supported topologies only two nodes received bootstrap values lower than 95%.DiscussionMelastomataceae plastomes are no exception for the general patterns observed in the genomic structure of land plant chloroplasts, being highly conserved and structurally similar to most other Myrtales. Despite the fact that the full plastome phylogeny shares most of the clades with the previously widely used and reduced data set, some changes are still observed and bootstrap support is higher. The plastome data set presented here is a step towards phylogenomic analyses in the Melastomataceae and will be a useful resource for future studies. creator: Marcelo Reginato creator: Kurt M. Neubig creator: Lucas C. Majure creator: Fabian A. Michelangeli uri: https://doi.org/10.7717/peerj.2715 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Reginato et al. title: Pentylenetetrazole-induced seizures in developing rats prenatally exposed to valproic acid link: https://peerj.com/articles/2709 last-modified: 2016-11-29 description: BackgroundEpidemiological evidence indicates epilepsy is more common in patients with autism spectrum disorders (ASD) (20–25%) than in the general population. The aim of this project was to analyze seizure susceptibility in developing rats prenatally exposed to valproic acid (VPA) as autism model.MethodsPregnant females were injected with VPA during the twelfth embryonic day. Seizures were induced in fourteen-days-old rat pups using two models of convulsions: pentylenetetrazole (PTZ) and lithium-pilocarpine (Li-Pilo).ResultsTwo subgroups with different PTZ-induced seizure susceptibility in rats exposed to VPA were found: a high susceptibility (VPA+) (28/42, seizure severity 5) and a low susceptibility (VPA−) (14/42, seizure severity 2). The VPA+ subgroup exhibited an increased duration of the generalized tonic-clonic seizure (GTCS; 45 ± 2.7 min), a higher number of rats showed several GTCS (14/28) and developed status epilepticus (SE) after PTZ injection (19/27) compared with control animals (36.6 ± 1.9 min; 10/39; 15/39, respectively). No differences in seizure severity, latency or duration of SE induced by Li-Pilo were detected between VPA and control animals.DiscussionPrenatal VPA modifies the susceptibility to PTZ-induced seizures in developing rats, which may be linked to an alteration in the GABAergic transmission. These findings contribute to a better understanding of the comorbidity between autism and epilepsy. creator: Angel A. Puig-Lagunes creator: Jorge Manzo creator: Luis Beltrán-Parrazal creator: Consuelo Morgado-Valle creator: Rebeca Toledo-Cárdenas creator: Maria-Leonor López-Meraz uri: https://doi.org/10.7717/peerj.2709 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Puig-Lagunes et al. title: Open surgery versus retroperitoneal laparoscopic nephrectomy for renal tuberculosis: a retrospective study of 120 patients link: https://peerj.com/articles/2708 last-modified: 2016-11-29 description: BackgroundLaparoscopic renal surgery has been widely used in the treatment of renal diseases. However, there is still little research about its application in addressing renal tuberculosis. The purpose of this study is to retrospectively investigate the surgical results of laparoscopic and open surgery for nonfunctional tuberculous kidneys.MethodsBetween May 2011 and June 2016, 120 nephrectomies were performed in patients with a nonfunctional tuberculous kidney. Of these, 69 patients underwent retroperitoneal laparoscopic nephrectomy, and 51 patients underwent open nephrectomy. Data about the patients’ characteristics and surgical outcomes were collected from their electronic medical records. Outcomes were compared between these two groups.ResultsOur results showed that a number of renal tuberculosis patients presented no significant symptoms during their disease. Lower urinary tract symptoms (LUTS) were the most common at a rate of 73/120, followed by flank pain or accidently discovery (66/120), urine abnormality (30/120) and fever (27/120). Patients who underwent open surgery were similar to laparoscopic patients with regard to sex, BMI, location, previous tuberculous history, grade, anemia, adhesion, hypertension, diabetes and preoperative serum creatinine level, but were generally older than laparoscopic patients. There were no significant differences between open and laparoscopic surgery in estimated blood loss, transfusion, postoperative hospital days and perioperative complication rate. However, the median operation time of laparoscopic operation was much longer than open surgery (180 [150–225] vs 135 [120–165] minutes, P < 0.01). Seven of the 69 laparoscopic operations were converted to open surgery because of severe adhesions.ConclusionLaparoscopic nephrectomy is as an effective treatment as open surgery for a nonfunctional tuberculous kidney, although it requires more time during the surgical procedure. No significant differences in other surgical outcomes were observed. creator: Su Zhang creator: You Luo creator: Cheng Wang creator: Hu Xiong creator: Sheng-Jun Fu creator: Li Yang uri: https://doi.org/10.7717/peerj.2708 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Zhang et al. title: Abundance and physiology of dominant soft corals linked to water quality in Jakarta Bay, Indonesia link: https://peerj.com/articles/2625 last-modified: 2016-11-29 description: Declining water quality is one of the main reasons of coral reef degradation in the Thousand Islands off the megacity Jakarta, Indonesia. Shifts in benthic community composition to higher soft coral abundances have been reported for many degraded reefs throughout the Indo-Pacific. However, it is not clear to what extent soft coral abundance and physiology are influenced by water quality. In this study, live benthic cover and water quality (i.e. dissolved inorganic nutrients (DIN), turbidity (NTU), and sedimentation) were assessed at three sites (< 20 km north of Jakarta) in Jakarta Bay (JB) and five sites along the outer Thousand Islands (20–60 km north of Jakarta). This was supplemented by measurements of photosynthetic yield and, for the first time, respiratory electron transport system (ETS) activity of two dominant soft coral genera, Sarcophyton spp. and Nephthea spp. Findings revealed highly eutrophic water conditions in JB compared to the outer Thousand Islands, with 44% higher DIN load (7.65 μM/L), 67% higher NTU (1.49 NTU) and 47% higher sedimentation rate (30.4 g m−2 d−1). Soft corals were the dominant type of coral cover within the bay (2.4% hard and 12.8% soft coral cover) compared to the outer Thousand Islands (28.3% hard and 6.9% soft coral cover). Soft coral abundances, photosynthetic yield, and ETS activity were highly correlated with key water quality parameters, particularly DIN and sedimentation rates. The findings suggest water quality controls the relative abundance and physiology of dominant soft corals in JB and may thus contribute to phase shifts from hard to soft coral dominance, highlighting the need to better manage water quality in order to prevent or reverse phase shifts. creator: Gunilla Baum creator: Indra Januar creator: Sebastian C.A. Ferse creator: Christian Wild creator: Andreas Kunzmann uri: https://doi.org/10.7717/peerj.2625 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Baum et al. title: Legionella shows a diverse secondary metabolism dependent on a broad spectrum Sfp-type phosphopantetheinyl transferase link: https://peerj.com/articles/2720 last-modified: 2016-11-24 description: Several members of the genus Legionella cause Legionnaires’ disease, a potentially debilitating form of pneumonia. Studies frequently focus on the abundant number of virulence factors present in this genus. However, what is often overlooked is the role of secondary metabolites from Legionella. Following whole genome sequencing, we assembled and annotated the Legionella parisiensis DSM 19216 genome. Together with 14 other members of the Legionella, we performed comparative genomics and analysed the secondary metabolite potential of each strain. We found that Legionella contains a huge variety of biosynthetic gene clusters (BGCs) that are potentially making a significant number of novel natural products with undefined function. Surprisingly, only a single Sfp-like phosphopantetheinyl transferase is found in all Legionella strains analyzed that might be responsible for the activation of all carrier proteins in primary (fatty acid biosynthesis) and secondary metabolism (polyketide and non-ribosomal peptide synthesis). Using conserved active site motifs, we predict some novel compounds that are probably involved in cell-cell communication, differing to known communication systems. We identify several gene clusters, which may represent novel signaling mechanisms and demonstrate the natural product potential of Legionella. creator: Nicholas J. Tobias creator: Tilman Ahrendt creator: Ursula Schell creator: Melissa Miltenberger creator: Hubert Hilbi creator: Helge B. Bode uri: https://doi.org/10.7717/peerj.2720 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Tobias et al. title: A revision of tetrapod footprints from the late Carboniferous of the West Midlands, UK link: https://peerj.com/articles/2718 last-modified: 2016-11-24 description: A series of sandstone slabs from Hamstead, Birmingham (West Midlands, UK), preserve an assemblage of tetrapod trackways and individual tracks from the Enville Member of the Salop Formation (late Carboniferous: late Moscovian–Kasimovian). This material has received limited previous study, despite being one of the few British sites to preserve Carboniferous tetrapod footprints. Here, we restudy and revise the taxonomy of this material, and document it using 3D models produced using photogrammetry. The assemblage is dominated by large tracks assigned to Limnopus isp., which were made by early amphibians (temnospondyls). A number of similar but smaller tracks are assigned to Batrachichnus salamandroides (also made by temnospondyls). Dimetropus leisnerianus (made by early synapsids) and Dromopus lacertoides (made by lizard-like sauropsids such as araeoscelids) are also present. This ichnofauna contrasts with a slightly stratigraphically older, more extensive and better-studied assemblage from Alveley (Shropshire), which is dominated by small amphibians with relatively rare reptiliomorphs, but which lacks Dromopus tracks. The presence of Dromopus lacertoides at Hamstead is consistent with the trend towards increasing aridity through the late Carboniferous. It is possible that the assemblage is the stratigraphically oldest occurrence of this important amniote ichnotaxon. creator: Luke E. Meade creator: Andrew S. Jones creator: Richard J. Butler uri: https://doi.org/10.7717/peerj.2718 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Meade et al. title: Mining SNPs in extracellular vesicular transcriptome of Trypanosoma cruzi: a step closer to early diagnosis of neglected Chagas disease link: https://peerj.com/articles/2693 last-modified: 2016-11-24 description: One of the newest and strongest members of intercellular communicators, the Extracellular vesicles (EVs) and their enclosed RNAs; Extracellular RNAs (exRNAs) have been acknowledged as putative biomarkers and therapeutic targets for various diseases. Although a very deep insight has not been possible into the physiology of these vesicles, they are believed to be involved in cell-to-cell communication and host-pathogen interactions. EVs might be significantly helpful in discovering biomarkers for possible target identification as well as prognostics, diagnostics and developing vaccines. In recent studies, highly bioactive EVs have drawn attention of parasitologists for being able to communicate between different cells and having likeliness of reflecting both source and target environments. Next-generation sequencing (NGS) has eased the way to have a deeper insight into these vesicles and their roles in various diseases. This article arises from bioinformatics-based analysis and predictive data mining of transcriptomic (RNA-Seq) data of EVs, derived from different life stages of Trypanosoma cruzi; a causing agent of neglected Chagas disease. Variants (Single Nucleotide Polymorphisms (SNPs)) were mined from Extracellular vesicular transcriptomic data and functionally analyzed using different bioinformatics based approaches. Functional analysis showed the association of these variants with various important factors like Trans-Sialidase (TS), Alpha Tubulin, P-Type H+-ATPase, etc. which, in turn, are associated with disease in different ways. Some of the ‘candidate SNPs’ were found to be stage-specific, which strengthens the probability of finding stage-specific biomarkers. These results may lead to a better understanding of Chagas disease, and improved knowledge may provide further development of the biomarkers for prognosis, diagnosis and drug development for treating Chagas disease. creator: Pallavi Gaur creator: Anoop Chaturvedi uri: https://doi.org/10.7717/peerj.2693 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Gaur and Chaturvedi title: Building healthy eating habits in childhood: a study of the attitudes, knowledge and dietary habits of schoolchildren in Malaysia link: https://peerj.com/articles/2651 last-modified: 2016-11-24 description: BackgroundOverweight and obesity have increased rapidly in incidence to become a global issue today. Overweight and obesity problems are significantly linked to unhealthy dietary patterns, physical inactivity and misperception of body image. This study aimed to determine whether Malaysian children build healthy eating habits from childhood.MethodsA survey on eating habits was conducted among primary school students in standards 4 to 6 in the state of Selangor, Malaysia. The findings of the study were reported in the form of descriptive statistics involving frequencies and percentages. Data from 400 respondents were analyzed.ResultsOur findings showed that the students understood the definition of healthy food and the types of food that are considered healthy. Although the students knew that food such as deep-fried drumsticks and hamburgers contain a high amount of saturated fat and cholesterol, these foods were still consumed by them. There was also a high consumption of foods that are fried and contain sugar, salt and saturated fat. In choosing food, two major factors contributed to the students’ decisions: cleanliness (65.8%) and the preference of their parents (12.3%).DiscussionOur findings indicate that by implementing the Integrated School Health Program (ISHP) properly, students’ eating habits can be improved by creating a school with a healthy environment. creator: Kazi Enamul Hoque creator: Megat Ahmad Kamaluddin creator: Ahmad Zabidi Abdul Razak creator: Afiq Athari Abdul Wahid uri: https://doi.org/10.7717/peerj.2651 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Hoque et al. title: Predictor effect of Locus Of Control (LOC) on self-care activities and metabolic control in individuals with type 2 diabetes link: https://peerj.com/articles/2722 last-modified: 2016-11-23 description: BackgroundPrevious studies have examined the role of individuals’ personal characteristics in diabetes management and used the locus of control theory to assess adherence to a diabetes management regimen. These studies have emphasized that having internal locus of control may be a protective factor in diabetes management.ObjectiveThe purpose of this study is to determine the predictor effect of locus of control on self-care activities and A1c level.MethodThe study is descriptive and relational. Researchers used a Diabetes Self-Care Activities Scale and a Locus of Control Scale to collect data. The study sample consisted of 129 individuals with type 2 diabetes.ResultsThe average score of locus of control of individuals with diabetes was 10.26, and the frequency of self-care activities in the past week was 2.9 days. A weak but statistically significant negative relation was found between the locus of control level and self-care activities of individuals with diabetes, which had no effect on A1c. It was determined that locus of control predicts 19% of self-care activities.ConclusionAccording to the study results, having internal locus of control had positive effects on self-care activities. Training and planning activities to improve internal locus of control can improve diabetes management. creator: Dilek Büyükkaya Besen creator: Neslihan Günüşen creator: Hamdiye Arda Sürücü creator: Cansu Koşar uri: https://doi.org/10.7717/peerj.2722 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Büyükkaya Besen et al. title: A qualitative view of the HIV epidemic in coastal Ecuador link: https://peerj.com/articles/2726 last-modified: 2016-11-22 description: In 2013 approximately 37,000 people were living with HIV in Ecuador (prevalence 0.4%), representing a generalized epidemic where most new infections arise from sexual interactions in the general population. Studies that examine attitudes towards people living with HIV (PLWH), individual risk perception of acquiring HIV amongst Ecuadorians, and the ways in which levels of risk perception may affect risk behaviors are lacking. This qualitative study aimed to fill this gap in the literature by investigating these issues in the rural, coastal community of Manglaralto, Ecuador, which has among the highest incidence of HIV in Ecuador. We conducted interviews with 15 patients at Manglaralto Hospital. Analysis of interview transcripts revealed widespread negative attitudes towards PLWH, prevalent risk behaviors such as multiple sex partners and lack of condom use, and low individual risk-perception of contracting HIV. These findings underscore the need for increased efforts to prevent further growth of the HIV epidemic in Ecuador. creator: Adam L. Beckman creator: Magdalena M. Wilson creator: Vishaal Prabhu creator: Nicola Soekoe creator: Humberto Mata creator: Lauretta E. Grau uri: https://doi.org/10.7717/peerj.2726 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Beckman et al. title: Glycyrrhetinic acid might increase the nephrotoxicity of bakuchiol by inhibiting cytochrome P450 isoenzymes link: https://peerj.com/articles/2723 last-modified: 2016-11-22 description: BackgroundLicorice, a popular traditional Chinese medicine (TCM), is widely used to moderate the effects (detoxification) of other herbs in TCM and often combined with Fructus Psoraleae. However, the classical TCM book states that Fructus Psoraleae is incompatible with licorice; the mechanism underlying this incompatibility has not been identified. Glycyrrhetinic acid (GA), the active metabolite of licorice, may increase the toxicity of bakuchiol (BAK), the main chemical ingredient in Psoralea corylifolia, by inhibiting its detoxification enzymes CYP450s.MethodsThe effect of concomitant GA administration on BAK-induced nephrotoxicity was investigated, and the metabolic interaction between BAK and GA was further studied in vitro and in vivo. The cytotoxicity was assessed using an MTT assay in a co-culture model of HK-2 cell and human liver microsomes (HLMs). The effect of GA on the metabolism of BAK, and on the activities of CYP isoforms were investigated in HLMs. The toxicokinetics and tissue exposure of BAK as well as the renal and hepatic functional markers were measured after the administration of a single oral dose in rats.ResultsIn vitro studies showed that the metabolic detoxification of BAK was significantly reduced by GA, and BAK was toxic to HK-2 cells, as indicated by 25∼40% decreases in viability when combined with GA. Further investigation revealed that GA significantly inhibited the metabolism of BAK in HLMs in a dose-dependent manner. GA strongly inhibits CYP3A4 and weakly inhibits CYP2C9 and CYP1A2; these CYP isoforms are involved in the metabolism of BAK. In vivo experiment found that a single oral dose of BAK combined with GA or in the presence of 1-aminobenzotriazole (ABT), altered the toxicokinetics of BAK in rats, increased the internal exposure, suppressed the elimination of BAK prototype, and therefore may have enhanced the renal toxicity.ConclusionThe present study demonstrated that GA inhibits CYP isoforms and subsequently may increase the nephrotoxicity of BAK, which underlie one of the possible mechanisms responsible for the incompatibility of Licorice with Fructus Psoraleae. creator: Aifang Li creator: Nana Ma creator: Zijing Zhao creator: Mei Yuan creator: Hua Li creator: Qi Wang uri: https://doi.org/10.7717/peerj.2723 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Li et al. title: A preliminary identification of Rf*-A619, a novel restorer gene for CMS-C in maize (Zea mays L.) link: https://peerj.com/articles/2719 last-modified: 2016-11-22 description: C-type cytoplasmic male sterility (CMS-C) is widely utilized for hybrid maize seed production. However, genetic mechanisms underlying the fertility restoration are very complicated. At present, there is a divergence on the number of fertility restorer genes in maize inbred line A619 for CMS-C. To further elucidate the restoring mechanism of A619, we used genetic analysis and molecular markers to confirm the restorer genes of maize inbred line A619 for C-type male sterile line C48-2 in this study. Firstly, the fertility segregations of (C48-2 × A619)F2 populations were investigated under three environments during 2013–2015. The segregation ratio of fertile and sterile plants in the F2 population fit to 15:1 via chi-square test and this result suggested that there are two dominant restorer genes in A619 for CMS-C, i.e., Rf4 and a novel gene named Rf*-A619. Next, based on the sequence differences between Rf4 and its recessive allelic rf4, a novel dominant marker F2/R2 was developed and validated to genotyping Rf4 in the F2 population. Through genotypic analysis, we found that there were a certain amount of fertile individuals without Rf4 which accounted for 3/16 in the F2 population via chi-square test at the 0.05 level. These results provided another proof to sustain that the inbred line A619 contains one additional restorer gene for CMS-C fertility restoration except Rf4. At last, we used one SSR marker which is tightly linked with the dominant restorer gene Rf5 to analyze those fertile plants without Rf4 in the F2 population. The PCR amplification results showed that Rf*-A619 is not allelic to Rf5 but a novel restorer gene for CMS-C. These results not only provide a basis for the mapping and characterization of a novel restorer gene but also give a new insight into the mechanism of CMS-C fertility restoration. creator: Liu Yongming creator: Zhao Zhuofan creator: Lu Yanli creator: Li Chuan creator: Wang Jing creator: Dong Boxiao creator: Liang Bing creator: Qiu Tao creator: Zeng Wenbing creator: Cao Moju uri: https://doi.org/10.7717/peerj.2719 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Yongming et al. title: Effect of common and experimental anti-tuberculosis treatments on Mycobacterium tuberculosis growing as biofilms link: https://peerj.com/articles/2717 last-modified: 2016-11-22 description: Much is known regarding the antibiotic susceptibility of planktonic cultures of Mycobacterium tuberculosis, the bacterium responsible for the lung disease tuberculosis (TB). As planktonically-grown M. tuberculosis are unlikely to be entirely representative of the bacterium during infection, we set out to determine how effective a range of anti-mycobacterial treatments were against M. tuberculosis growing as a biofilm, a bacterial phenotype known to be more resistant to antibiotic treatment. Light levels from bioluminescently-labelled M. tuberculosis H37Rv (strain BSG001) were used as a surrogate for bacterial viability, and were monitored before and after one week of treatment. After treatment, biofilms were disrupted, washed and inoculated into fresh broth and plated onto solid media to rescue any surviving bacteria. We found that in this phenotypic state M. tuberculosis was resistant to the majority of the compounds tested. Minimum inhibitory concentrations (MICs) increased by 20-fold to greater than 1,000-fold, underlying the potential of this phenotype to cause significant problems during treatment. creator: James P. Dalton creator: Benedict Uy creator: Narisa Phummarin creator: Brent R. Copp creator: William A. Denny creator: Simon Swift creator: Siouxsie Wiles uri: https://doi.org/10.7717/peerj.2717 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Dalton et al. title: Pathogen reduction co-benefits of nutrient best management practices link: https://peerj.com/articles/2713 last-modified: 2016-11-22 description: BackgroundMany of the practices currently underway to reduce nitrogen, phosphorus, and sediment loads entering the Chesapeake Bay have also been observed to support reduction of disease-causing pathogen loadings. We quantify how implementation of these practices, proposed to meet the nutrient and sediment caps prescribed by the Total Maximum Daily Load (TMDL), could reduce pathogen loadings and provide public health co-benefits within the Chesapeake Bay system.MethodsWe used published data on the pathogen reduction potential of management practices and baseline fecal coliform loadings estimated as part of prior modeling to estimate the reduction in pathogen loadings to the mainstem Potomac River and Chesapeake Bay attributable to practices implemented as part of the TMDL. We then compare the estimates with the baseline loadings of fecal coliform loadings to estimate the total pathogen reduction potential of the TMDL.ResultsWe estimate that the TMDL practices have the potential to decrease disease-causing pathogen loads from all point and non-point sources to the mainstem Potomac River and the entire Chesapeake Bay watershed by 19% and 27%, respectively. These numbers are likely to be underestimates due to data limitations that forced us to omit some practices from analysis.DiscussionBased on known impairments and disease incidence rates, we conclude that efforts to reduce nutrients may create substantial health co-benefits by improving the safety of water-contact recreation and seafood consumption. creator: Jennifer Richkus creator: Lisa A. Wainger creator: Mary C. Barber uri: https://doi.org/10.7717/peerj.2713 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Richkus et al. title: Dominance relationships in a family pack of captive arctic wolves (Canis lupus arctos): the influence of competition for food, age and sex link: https://peerj.com/articles/2707 last-modified: 2016-11-22 description: BackgroundDominance is one of the most pervasive concepts in the study of wolf social behaviour but recently its validity has been questioned. For some authors, the bonds between members of wolf families are better described as parent-offspring relationships and the concept of dominance should be used just to evaluate the social dynamics of non-familial captive pack members (e.g., Mech & Cluff, 2010). However, there is a dearth of studies investigating dominance relationships and its correlates in wolf family packs.MethodsHere, we applied a combination of the most commonly used quantitative methods to evaluate the dominance relationships in a captive family pack of 19 Arctic wolves.ResultsWe found a significant linear and completely transitive hierarchy based on the direction of submissive behaviours and found that dominance relationships were not influenced by the competitive contexts (feeding vs. non-feeding context). A significant linear hierarchy also emerges amongst siblings once the breeding pair (the two top-ranking individuals) is removed from analyses. Furthermore, results suggest that wolves may use greeting behaviour as a formal signal of subordination. Whereas older wolves were mostly dominant over younger ones, no clear effect of sex was found. However, frequency of agonistic (submissive, dominant and aggressive) behaviours was higher between female–female and male–male dyads than female–male dyads and sex-separated linear hierarchies showed a stronger linearity than the mixed one. Furthermore, dominance status was conveyed through different behavioural categories during intra-sexual and inter-sexual interactions.DiscussionCurrent results highlight the importance of applying a systematic methodology considering the individuals’ age and sex when evaluating the hierarchical structure of a social group. Moreover, they confirm the validity of the concept of dominance relationships in describing the social bonds within a family pack of captive wolves. creator: Simona Cafazzo creator: Martina Lazzaroni creator: Sarah Marshall-Pescini uri: https://doi.org/10.7717/peerj.2707 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Cafazzo et al. title: Hypothesis on Serenoa repens (Bartram) small extract inhibition of prostatic 5α-reductase through an in silico approach on 5β-reductase x-ray structure link: https://peerj.com/articles/2698 last-modified: 2016-11-22 description: Benign prostatic hyperplasia is a common disease in men aged over 50 years old, with an incidence increasing to more than 80% over the age of 70, that is increasingly going to attract pharmaceutical interest. Within conventional therapies, such as α-adrenoreceptor antagonists and 5α-reductase inhibitor, there is a large requirement for treatments with less adverse events on, e.g., blood pressure and sexual function: phytotherapy may be the right way to fill this need. Serenoa repens standardized extract has been widely studied and its ability to reduce lower urinary tract symptoms related to benign prostatic hyperplasia is comprehensively described in literature. An innovative investigation on the mechanism of inhibition of 5α-reductase by Serenoa repens extract active principles is proposed in this work through computational methods, performing molecular docking simulations on the crystal structure of human liver 5β-reductase. The results confirm that both sterols and fatty acids can play a role in the inhibition of the enzyme, thus, suggesting a competitive mechanism of inhibition. This work proposes a further confirmation for the rational use of herbal products in the management of benign prostatic hyperplasia, and suggests computational methods as an innovative, low cost, and non-invasive process for the study of phytocomplex activity toward proteic targets. creator: Paolo Governa creator: Daniela Giachetti creator: Marco Biagi creator: Fabrizio Manetti creator: Luca De Vico uri: https://doi.org/10.7717/peerj.2698 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Governa et al. title: Detecting sedimentation impacts to coral reefs resulting from dredging the Port of Miami, Florida USA link: https://peerj.com/articles/2711 last-modified: 2016-11-17 description: The federal channel at Port of Miami, Florida, USA, was dredged between late 2013 and early 2015 to widen and deepen the channel. Due to the limited spatial extent of impact-assessment monitoring associated with the project, the extent of the dredging impacts on surrounding coral reefs has not been well quantified. Previously published remote sensing analyses, as well as agency and anecdotal reports suggest the most severe and largest area of sedimentation occurred on a coral reef feature referred to as the Inner Reef, particularly in the sector north of the channel. A confounding regional warm-water mass bleaching event followed by a coral disease outbreak during this same time frame made the assessment of dredging-related impacts to coral reefs adjacent to the federal channel difficult but still feasible. The current study sought to better understand the sedimentation impacts that occurred in the coral reef environment surrounding Port of Miami, to distinguish those impacts from other regional events or disturbances, and provide supplemental information on impact assessment that will inform discussions on compensatory mitigation requirements. To this end, in-water field assessments conducted after the completion of dredging and a time series analysis of tagged corals photographed pre-, during, and post-dredging, are used to discern dredging-related sedimentation impacts for the Inner Reef north. Results indicate increased sediment accumulation, severe in certain times and places, and an associated biological response (e.g., higher prevalence of partial mortality of corals) extended up to 700 m from the channel, whereas project-associated monitoring was limited to 50 m from the channel. These results can contribute to more realistic prediction of areas of indirect effect from dredging projects needed to accurately evaluate proposed projects and design appropriate compliance monitoring. Dredging projects near valuable and sensitive habitats subject to local and global stressors require monitoring methods capable of discerning non-dredging related impacts and adaptive management to ensure predicted and unpredicted project-related impacts are quantified. Anticipated increasing frequency and intensity of seasonal warming stress also suggests that manageable- but- unavoidable local stressors such as dredging should be partitioned from such seasonal thermal stress events. creator: Margaret W. Miller creator: Jocelyn Karazsia creator: Carolyn E. Groves creator: Sean Griffin creator: Tom Moore creator: Pace Wilber creator: Kurtis Gregg uri: https://doi.org/10.7717/peerj.2711 license: http://creativecommons.org/publicdomain/zero/1.0/ rights: title: Ficolin-2 inhibitors are present in sera after prolonged storage at −80 °C link: https://peerj.com/articles/2705 last-modified: 2016-11-17 description: Ficolins can activate the lectin pathway of the complement system that provides innate immune protection against pathogens, marks host cellular debris for clearance, and promotes inflammation. Baseline inflammation increases with aging in a phenomenon known as “inflammaging.” Although IL-6 and C-reactive protein are known to increase with age, contributions of many complement factors, including ficolins, to inflammaging have been little studied. Ficolin-2 is abundant in human serum and can recognize many target structures; therefore, ficolin-2 has potential to contribute to inflammaging. We hypothesized that inflammaging would alter ficolin-2 levels among older adults and examined 360 archived sera collected from older individuals. We found that these sera had apparently reduced ficolin-2 levels and that 84.2% of archived sera exhibited ficolin-2 inhibitors, which suppressed apparent amounts of ficolin-2 detected by enzyme-linked immunosorbent assay. Fresh serum samples were obtained from donors whose archived sera showed inhibitors, but the fresh sera did not have ficolin-2 inhibitors. Ficolin-2 inhibitors were present in other long-stored sera from younger persons. Furthermore, noninhibiting samples and fresh sera from older adults had apparently normal amounts of ficolin-2. Thus, ficolin-2 inhibitors may arise as an artifact of long-term storage of serum at −80 °C. creator: Kimball Aaron Geno creator: Richard E. Kennedy creator: Patricia Sawyer creator: Cynthia J. Brown creator: Moon H. Nahm uri: https://doi.org/10.7717/peerj.2705 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Geno et al. title: Multiple, independent colonizations of the Hawaiian Archipelago by the family Dolichopodidae (Diptera) link: https://peerj.com/articles/2704 last-modified: 2016-11-17 description: The family Dolichopodidae forms two of the four largest evolutionary radiations in the Hawaiian Islands across all flies: Campsicnemus (183 spp) and the Eurynogaster complex (66 spp). They also include a small radiation of Conchopus (6 spp). A handful of other dolichopodid species are native to the islands in singleton lineages or small radiations. This study provides a phylogenetic perspective on the colonization history of the dolichopodid fauna in the islands. We generated a multi-gene data set including representatives from 11 of the 14 endemic Hawaiian dolichopodid genera to examine the history of colonization to the islands, and analyzed it using Bayesian and maximum likelihood phylogenetic methods. We used a subset of the data that included Conchopus and the eight genera comprising the Eurynogaster complex to estimate the first phylogenetic hypothesis for these endemic groups, then used Beast to estimate their age of arrival to the archipelago. The Eurynogaster complex, Campsicnemus and Conchopus are clearly the result of independent colonizations. The results strongly support the Eurynogaster complex as a monophyletic group, and also supports the monophyly of 4 of the 8 described genera within the complex (Adachia, Arciellia, Uropachys and Eurynogaster). Members of the family Dolichopodidae have been dispersing over vast distances to colonize the Hawaiian Archipelago for millions of years, leading to multiple independent evolutionary diversification events. The Eurynogaster complex arrived in the Hawaiian Archipelago 11.8 Ma, well before the arrival of Campsicnemus (4.5 Ma), and the even more recent Conchopus (1.8 Ma). Data presented here demonstrate that the Hawaiian Dolichopodidae both disperse and diversify easily, a rare combination that lays the groundwork for field studies on the reproductive isolating mechanisms and ecological partitioning of this group. creator: Kari Roesch Goodman creator: Neal Evenhuis creator: Pavla Bartošová-Sojková creator: Patrick Michael O’Grady uri: https://doi.org/10.7717/peerj.2704 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Goodman et al. title: Salinity modulates thermotolerance, energy metabolism and stress response in amphipods Gammarus lacustris link: https://peerj.com/articles/2657 last-modified: 2016-11-17 description: Temperature and salinity are important abiotic factors for aquatic invertebrates. We investigated the influence of different salinity regimes on thermotolerance, energy metabolism and cellular stress defense mechanisms in amphipods Gammarus lacustris Sars from two populations. We exposed amphipods to different thermal scenarios and determined their survival as well as activity of major antioxidant enzymes (peroxidase, catalase, glutathione S-transferase) and parameters of energy metabolism (content of glucose, glycogen, ATP, ADP, AMP and lactate). Amphipods from a freshwater population were more sensitive to the thermal challenge, showing higher mortality during acute and gradual temperature change compared to their counterparts from a saline lake. A more thermotolerant population from a saline lake had high activity of antioxidant enzymes. The energy limitations of the freshwater population (indicated by low baseline glucose levels, downward shift of the critical temperature of aerobic metabolism and inability to maintain steady-state ATP levels during warming) was observed, possibly reflecting a trade-off between the energy demands for osmoregulation under the hypo-osmotic condition of a freshwater environment and protection against temperature stress. creator: Kseniya P. Vereshchagina creator: Yulia A. Lubyaga creator: Zhanna Shatilina creator: Daria Bedulina creator: Anton Gurkov creator: Denis V. Axenov-Gribanov creator: Boris Baduev creator: Elizaveta S. Kondrateva creator: Mikhail Gubanov creator: Egor Zadereev creator: Inna Sokolova creator: Maxim Timofeyev uri: https://doi.org/10.7717/peerj.2657 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Vereshchagina et al. title: Ceratopogonidae (Diptera: Nematocera) of the piedmont of the Yungas forests of Tucumán: ecology and distribution link: https://peerj.com/articles/2655 last-modified: 2016-11-17 description: Within the Ceratopogonidae family, many genera transmit numerous diseases to humans and animals, while others are important pollinators of tropical crops. In the Yungas ecoregion of Argentina, previous systematic and ecological research on Ceratopogonidae focused on Culicoides, since they are the main transmitters of mansonelliasis in northwestern Argentina; however, few studies included the genera Forcipomyia, Dasyhelea, Atrichopogon, Alluaudomyia, Echinohelea, and Bezzia. Therefore, the objective of this study was to determine the presence and abundance of Ceratopogonidae in this region, their association with meteorological variables, and their variation in areas disturbed by human activity. Monthly collection of specimens was performed from July 2008 to July 2009 using CDC miniature light traps deployed for two consecutive days. A total of 360 specimens were collected, being the most abundant Dasyhelea genus (48.06%) followed by Forcipomyia (26.94%) and Atrichopogon (13.61%). Bivariate analyses showed significant differences in the abundance of the genera at different sampling sites and climatic conditions, with the summer season and El Corralito site showing the greatest abundance of specimens. Accumulated rainfall was the variable that related the most to the abundance of Culicoides (10.56%), while temperature was the most closely related variable to the abundance of Forcipomyia, Dasyhelea, and Atrichopogon. creator: José Manuel Direni Mancini creator: Cecilia Adriana Veggiani-Aybar creator: Ana Denise Fuenzalida creator: Mercedes Sara Lizarralde de Grosso creator: María Gabriela Quintana uri: https://doi.org/10.7717/peerj.2655 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Direni Mancini et al. title: PDIA3 gene induces visceral hypersensitivity in rats with irritable bowel syndrome through the dendritic cell-mediated activation of T cells link: https://peerj.com/articles/2644 last-modified: 2016-11-17 description: This study investigated the mechanism of protein disulfide-isomerase A3 (PDIA3)-induced visceral hypersensitivity in irritable bowel syndrome (IBS). Rats were treated with saline (control), acetic acid and restraint stress (IBS model), empty vector (RNAi control) and PDIA3-RNAi vector (PDIA3-RNAi). Mesenteric lymph node DCs (MLNDCs) and splenic CD4+/CD8+ T cells were isolated for co-cultivation. Compared with control, MLNDCs co-cultured with CD4+ or CD8+ T cells showed an increased ability to promote T cell proliferation and produced more IL-4 or IL-9 secretion. Compared with the RNAi control, MLNDCs from the PDIA3 knockdown models were less effective in promoting the proliferation of CD4+/CD8+ T cells. It is concluded that PDIA3 plays an important role in the development of IBS through the DC-mediated activation of T cells, resulting in degranulation of MCs and visceral hypersensitivity. creator: Zhaomeng Zhuang creator: Lu Zhang creator: Xiaoteng Wang creator: Liyuan Tao creator: Bin Lv uri: https://doi.org/10.7717/peerj.2644 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Zhuang et al. title: A large scale test of the gaming-enhancement hypothesis link: https://peerj.com/articles/2710 last-modified: 2016-11-16 description: A growing research literature suggests that regular electronic game play and game-based training programs may confer practically significant benefits to cognitive functioning. Most evidence supporting this idea, the gaming-enhancement hypothesis, has been collected in small-scale studies of university students and older adults. This research investigated the hypothesis in a general way with a large sample of 1,847 school-aged children. Our aim was to examine the relations between young people’s gaming experiences and an objective test of reasoning performance. Using a Bayesian hypothesis testing approach, evidence for the gaming-enhancement and null hypotheses were compared. Results provided no substantive evidence supporting the idea that having preference for or regularly playing commercially available games was positively associated with reasoning ability. Evidence ranged from equivocal to very strong in support for the null hypothesis over what was predicted. The discussion focuses on the value of Bayesian hypothesis testing for investigating electronic gaming effects, the importance of open science practices, and pre-registered designs to improve the quality of future work. creator: Andrew K. Przybylski creator: John C. Wang uri: https://doi.org/10.7717/peerj.2710 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Przybylski and Wang title: Historical reconstruction of climatic and elevation preferences and the evolution of cloud forest-adapted tree ferns in Mesoamerica link: https://peerj.com/articles/2696 last-modified: 2016-11-16 description: BackgroundCloud forests, characterized by a persistent, frequent or seasonal low-level cloud cover and fragmented distribution, are one of the most threatened habitats, especially in the Neotropics. Tree ferns are among the most conspicuous elements in these forests, and ferns are restricted to regions in which minimum temperatures rarely drop below freezing and rainfall is high and evenly distributed around the year. Current phylogeographic data suggest that some of the cloud forest-adapted species remained in situ or expanded to the lowlands during glacial cycles and contracted allopatrically during the interglacials. Although the observed genetic signals of population size changes of cloud forest-adapted species including tree ferns correspond to predicted changes by Pleistocene climate change dynamics, the observed patterns of intraspecific lineage divergence showed temporal incongruence.MethodsHere we combined phylogenetic analyses, ancestral area reconstruction, and divergence time estimates with climatic and altitudinal data (environmental space) for phenotypic traits of tree fern species to make inferences about evolutionary processes in deep time. We used phylogenetic Bayesian inference and geographic and altitudinal distribution of tree ferns to investigate ancestral area and elevation and environmental preferences of Mesoamerican tree ferns. The phylogeny was then used to estimate divergence times and ask whether the ancestral area and elevation and environmental shifts were linked to climatic events and historical climatic preferences.ResultsBayesian trees retrieved Cyathea, Alsophyla, Gymnosphaera and Sphaeropteris in monophyletic clades. Splits for species in these genera found in Mesoamerican cloud forests are recent, from the Neogene to the Quaternary, Australia was identified as the ancestral area for the clades of these genera, except for Gymnosphaera that was Mesoamerica. Climate tolerance was not divergent from hypothesized ancestors for the most significant variables or elevation. For elevational shifts, we found repeated change from low to high elevations.ConclusionsOur data suggest that representatives of Cyatheaceae main lineages migrated from Australia to Mesoamerican cloud forests in different times and have persisted in these environmentally unstable areas but extant species diverged recentrly from their ancestors. creator: Victoria Sosa creator: Juan Francisco Ornelas creator: Santiago Ramírez-Barahona creator: Etelvina Gándara uri: https://doi.org/10.7717/peerj.2696 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Sosa et al. title: Biology of tiny animals: three new species of minute salamanders (Plethodontidae: Thorius) from Oaxaca, Mexico link: https://peerj.com/articles/2694 last-modified: 2016-11-15 description: We describe three new species of minute salamanders, genus Thorius, from the Sierra Madre del Sur of Oaxaca, Mexico. Until now only a single species, T. minutissimus, has been reported from this region, although molecular data have long shown extensive genetic differentiation among geographically disjunct populations. Adult Thorius pinicola sp. nov., T. longicaudus sp. nov., and T. tlaxiacus sp. nov. are larger than T. minutissimus and possess elliptical rather than oval nostrils; T. pinicola and T. longicaudus also have longer tails. All three new species occur west of the range of T. minutissimus, which has the easternmost distribution of any member of the genus. The new species are distinguished from each other and from other named Thorius in Oaxaca by a combination of adult body size, external morphology and osteology, and by protein characters (allozymes) and differences in DNA sequences. In addition, we redescribe T. minutissimus and a related species, T. narisovalis, to further clarify the taxonomic status of Oaxacan populations and to facilitate future studies of the remaining genetically differentiated Thorius that cannot be satisfactorily assigned to any named species. Populations of all five species considered here appear to have declined dramatically over the last one or two decades and live specimens are difficult to find in nature. Thorius may be the most endangered genus of amphibians in the world. All species may go extinct before the end of this century. creator: Gabriela Parra-Olea creator: Sean M. Rovito creator: Mario García-París creator: Jessica A. Maisano creator: David B. Wake creator: James Hanken uri: https://doi.org/10.7717/peerj.2694 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Parra-Olea et al. title: jicbioimage: a tool for automated and reproducible bioimage analysis link: https://peerj.com/articles/2674 last-modified: 2016-11-15 description: There has been steady improvement in methods for capturing bioimages. However analysing these images still remains a challenge. The Python programming language provides a powerful and flexible environment for scientific computation. It has a wide range of supporting libraries for image processing but lacks native support for common bioimage formats, and requires specific code to be written to ensure that suitable audit trails are generated and analyses are reproducible. Here we describe the development of a Python tool that: (1) allows users to quickly view and explore microscopy data; (2) generate reproducible analyses, encoding a complete history of image transformations from raw data to final result; and (3) scale up analyses from initial exploration to high throughput processing pipelines, with a minimal amount of extra effort. The tool, jicbioimage, is open source and freely available online at http://jicbioimage.readthedocs.io. creator: Tjelvar S. G. Olsson creator: Matthew Hartley uri: https://doi.org/10.7717/peerj.2674 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Olsson and Hartley title: Gene expression profile of sodium channel subunits in the anterior cingulate cortex during experimental paclitaxel-induced neuropathic pain in mice link: https://peerj.com/articles/2702 last-modified: 2016-11-15 description: Paclitaxel, a chemotherapeutic agent, causes neuropathic pain whose supraspinal pathophysiology is not fully understood. Dysregulation of sodium channel expression, studied mainly in the periphery and spinal cord level, contributes to the pathogenesis of neuropathic pain. We examined gene expression of sodium channel (Nav) subunits by real time polymerase chain reaction (PCR) in the anterior cingulate cortex (ACC) at day 7 post first administration of paclitaxel, when mice had developed paclitaxel-induced thermal hyperalgesia. The ACC was chosen because increased activity in the ACC has been observed during neuropathic pain. In the ACC of vehicle-treated animals the threshold cycle (Ct) values for Nav1.4, Nav1.5, Nav1.7, Nav1.8 and Nav1.9 were above 30 and/or not detectable in some samples. Thus, comparison in mRNA expression between untreated control, vehicle-treated and paclitaxel treated animals was done for Nav1.1, Nav1.2, Nav1.3, Nav1.6, Nax as well as Navβ1–Navβ4. There were no differences in the transcript levels of Nav1.1–Nav1.3, Nav1.6, Nax, Navβ1–Navβ3 between untreated and vehicle-treated mice, however, vehicle treatment increased Navβ4 expression. Paclitaxel treatment significantly increased the mRNA expression of Nav1.1, Nav1.2, Nav1.6 and Nax, but not Nav1.3, sodium channel alpha subunits compared to vehicle-treated animals. Treatment with paclitaxel significantly increased the expression of Navβ1 and Navβ3, but not Navβ2 and Navβ4, sodium channel beta subunits compared to vehicle-treated animals. These findings suggest that during paclitaxel-induced neuropathic pain (PINP) there is differential upregulation of sodium channels in the ACC, which might contribute to the increased neuronal activity observed in the area during neuropathic pain. creator: Willias Masocha uri: https://doi.org/10.7717/peerj.2702 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Masocha title: Salinity and temperature significantly influence seed germination, seedling establishment, and seedling growth of eelgrass Zostera marina L. link: https://peerj.com/articles/2697 last-modified: 2016-11-15 description: Globally, seagrass beds have been recognized as critical yet declining coastal habitats. To mitigate seagrass losses, seagrass restorations have been conducted in worldwide over the past two decades. Seed utilization is considered to be an important approach in seagrass restoration efforts. In this study, we investigated the effects of salinity and temperature on seed germination, seedling establishment, and seedling growth of eelgrass Zostera marina L. (Swan Lake, northern China). We initially tested the effects of salinity (0, 5, 10, 15, 20, 25, 30, 35, and 40 ppt) and water temperature (5, 10, 15, and 20 °C) on seed germination to identify optimal levels. To identify levels of salinity that could potentially limit survival and growth, and, consequently, the spatial distribution of seedlings in temperate estuaries, we then examined the effect of freshwater and other salinity levels (10, 20, and 30 ppt) on seedling growth and establishment to confirm suitable conditions for seedling development. Finally, we examined the effect of transferring germinated seeds from freshwater or low salinity levels (1, 5, and 15 ppt) to natural seawater (32 ppt) on seedling establishment rate (SER) at 15 °C. In our research, we found that: (1) Mature seeds had a considerably lower moisture content than immature seeds; therefore, moisture content may be a potential indicator of Z. marina seed maturity; (2) Seed germination significantly increased at low salinity (p < 0.001) and high temperature (p < 0.001). Salinity had a much stronger influence on seed germination than temperature. Maximum seed germination (88.67 ± 5.77%) was recorded in freshwater at 15 °C; (3) Freshwater and low salinity levels (< 20 ppt) increased germination but had a strong negative effect on seedling morphology (number of leaves per seedling reduced from 2 to 0, and maximum seedling leaf length reduced from 4.48 to 0 cm) and growth (seedling biomass reduced by 46.15–66.67% and maximum seedling length reduced by 21.16–69.50%). However, Z. marina performed almost equally well at salinities of 20 and 30 ppt. Very few germinated seeds completed leaf differentiation and seedling establishment in freshwater or at low salinity, implying that freshwater and low salinity may potentially limit the distribution of this species in coastal and estuarine waters. Therefore, the optimum salinity for Z. marina seedling establishment and colonization appears to be above 20 ppt in natural beds; (4) Seeds germinated in freshwater or at low salinity levels could be transferred to natural seawater to accomplish seedling establishment and colonization. This may be the optimal method for the adoption of seed utilization in seagrass restoration. We also identified seven stages of seed germination and seedling metamorphosis in order to characterize growth and developmental characteristics. Our results may serve as useful information for Z. marina habitat establishment and restoration programs. creator: Shaochun Xu creator: Yi Zhou creator: Pengmei Wang creator: Feng Wang creator: Xiaomei Zhang creator: Ruiting Gu uri: https://doi.org/10.7717/peerj.2697 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Xu et al. title: The scavenger receptor repertoire in six cnidarian species and its putative role in cnidarian-dinoflagellate symbiosis link: https://peerj.com/articles/2692 last-modified: 2016-11-15 description: Many cnidarians engage in a mutualism with endosymbiotic photosynthetic dinoflagellates that forms the basis of the coral reef ecosystem. Interpartner interaction and regulation includes involvement of the host innate immune system. Basal metazoans, including cnidarians have diverse and complex innate immune repertoires that are just beginning to be described. Scavenger receptors (SR) are a diverse superfamily of innate immunity genes that recognize a broad array of microbial ligands and participate in phagocytosis of invading microbes. The superfamily includes subclades named SR-A through SR-I that are categorized based on the arrangement of sequence domains including the scavenger receptor cysteine rich (SRCR), the C-type lectin (CTLD) and the CD36 domains. Previous functional and gene expression studies on cnidarian-dinoflagellate symbiosis have implicated SR-like proteins in interpartner communication and regulation. In this study, we characterized the SR repertoire from a combination of genomic and transcriptomic resources from six cnidarian species in the Class Anthozoa. We combined these bioinformatic analyses with functional experiments using the SR inhibitor fucoidan to explore a role for SRs in cnidarian symbiosis and immunity. Bioinformatic searches revealed a large diversity of SR-like genes that resembled SR-As, SR-Bs, SR-Es and SR-Is. SRCRs, CTLDs and CD36 domains were identified in multiple sequences in combinations that were highly homologous to vertebrate SRs as well as in proteins with novel domain combinations. Phylogenetic analyses of CD36 domains of the SR-B-like sequences from a diversity of metazoans grouped cnidarian with bilaterian sequences separate from other basal metazoans. All cnidarian sequences grouped together with moderate support in a subclade separately from bilaterian sequences. Functional experiments were carried out on the sea anemone Aiptasia pallida that engages in a symbiosis with Symbiodinium minutum (clade B1). Experimental blocking of the SR ligand binding site with the inhibitor fucoidan reduced the ability of S. minutum to colonize A. pallida suggesting that host SRs play a role in host-symbiont recognition. In addition, incubation of symbiotic anemones with fucoidan elicited an immune response, indicating that host SRs function in immune modulation that results in host tolerance of the symbionts. creator: Emilie F. Neubauer creator: Angela Z. Poole creator: Olivier Detournay creator: Virginia M. Weis creator: Simon K. Davy uri: https://doi.org/10.7717/peerj.2692 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Neubauer et al. title: Assessing pollinators’ use of floral resource subsidies in agri-environment schemes: An illustration using Phacelia tanacetifolia and honeybees link: https://peerj.com/articles/2677 last-modified: 2016-11-15 description: BackgroundHoneybees (Apis mellifera L.) are frequently used in agriculture for pollination services because of their abundance, generalist floral preferences, ease of management and hive transport. However, their populations are declining in many countries. Agri-Environment Schemes (AES) are being implemented in agricultural systems to combat the decline in populations of pollinators and other insects. Despite AES being increasingly embedded in policy and budgets, scientific assessments of many of these schemes still are lacking, and only a few studies have examined the extent to which insect pollinators use the floral enhancements that are part of AES and on which floral components they feed (i.e., pollen and/or nectar).MethodsIn the present work, we used a combination of observations on honeybee foraging for nectar/pollen from the Californian annual plant Phacelia tanacetifolia in the field, collection of pollen pellets from hives, and pollen identification, to assess the value of adding phacelia to an agro-ecosystem to benefit honeybees.ResultsIt was found that phacelia pollen was almost never taken by honeybees. The work here demonstrates that honeybees may not use the floral enhancements added to a landscape as expected and points to the need for more careful assessments of what resources are used by honeybees in AES and understanding the role, if any, which AES play in enhancing pollinator fitness.DiscussionWe recommend using the methodology in this paper to explore the efficacy of AES before particular flowering species are adopted more widely to give a more complete illustration of the actual efficacy of AES. creator: Rowan Sprague creator: Stéphane Boyer creator: Georgia M. Stevenson creator: Steve D. Wratten uri: https://doi.org/10.7717/peerj.2677 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Sprague et al. title: Zebra mussel beds: an effective feeding ground for Ponto-Caspian gobies or suitable shelter for their prey? link: https://peerj.com/articles/2672 last-modified: 2016-11-15 description: Aggregations of the Ponto-Caspian invasive zebra mussel (Dreissena polymorpha) constitute a suitable habitat for macroinvertebrates, considerably increasing their abundance and providing effective antipredator protection. Thus, the overall effect of a mussel bed on particular predator species may vary from positive to negative, depending on both prey density increase and predator ability to prey in a structurally complex habitat. Alien Ponto-Caspian goby fish are likely to be facilitated when introduced into new areas by zebra mussels, provided that they are capable of utilizing mussel beds as habitat and feeding grounds. We ran laboratory experiments to find which prey (chironomid larvae) densities (from ca. 500 to 2,000 individuals m−2) in a mussel bed make it a more beneficial feeding ground for the racer goby Babka gymnotrachelus (RG) and western tubenose goby Proterorhinus semilunaris (WTG) compared to sandy and stone substrata (containing the basic prey density of 500 ind. m−2). Moreover, we checked how food availability affects habitat selection by fish. Mussel beds became more suitable for fish than alternative mineral substrata when food abundance was at least two times higher (1,000 vs. 500 ind. m−2), regardless of fish size and species. WTG was associated with mussel beds regardless of its size and prey density, whereas RG switched to this habitat when it became a better feeding ground than alternative substrata. Larger RG exhibited a stronger affinity for mussels than small individuals. WTG fed more efficiently from a mussel bed at high food abundances than RG. A literature review has shown that increasing chironomid density, which in our study was sufficient to make a mussel habitat an attractive feeding ground for the gobies, is commonly observed in mussel beds in the field. Therefore, we conclude that zebra mussels may positively affect the alien goby species and are likely to facilitate their establishment in novel areas, contributing to an invasional meltdown in the Ponto-Caspian invasive community. creator: Jarosław Kobak creator: Małgorzata Poznańska creator: Łukasz Jermacz creator: Tomasz Kakareko creator: Daniel Prądzynski creator: Małgorzata Łodygowska creator: Karolina Montowska creator: Karolina Bącela-Spychalska uri: https://doi.org/10.7717/peerj.2672 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Kobak et al. title: Lithocholic acid induces endoplasmic reticulum stress, autophagy and mitochondrial dysfunction in human prostate cancer cells link: https://peerj.com/articles/2445 last-modified: 2016-11-15 description: Lithocholic acid (LCA) is a secondary bile acid that is selectively toxic to human neuroblastoma, breast and prostate cancer cells, whilst sparing normal cells. We previously reported that LCA inhibited cell viability and proliferation and induced apoptosis and necrosis of androgen-dependent LNCaP and androgen-independent PC-3 human prostate cancer cells. In the present study, we investigated the roles of endoplasmic reticulum (ER) stress, autophagy and mitochondrial dysfunction in the toxicity of LCA in PC-3 and autophagy deficient, androgen-independent DU-145 cells. LCA induced ER stress-related proteins, such as CCAAT-enhancer-binding protein homologous protein (CHOP), and the phosphorylation of eukaryotic initiation factor 2-alpha (p-eIF2α) and c-Jun N-terminal kinases (p-JNK) in both cancer cell-types. The p53 upregulated modulator of apoptosis (PUMA) and B cell lymphoma-like protein 11 (BIM) levels were decreased at overtly toxic LCA concentrations, although PUMA levels increased at lower LCA concentrations in both cell lines. LCA induced autophagy-related conversion of microtubule-associated proteins 1A/1B light chain 3B (LC3BI–LC3BII), and autophagy-related protein ATG5 in PC-3 cells, but not in autophagy-deficient DU-145 cells. LCA (>10 µM) increased levels of reactive oxygen species (ROS) concentration-dependently in PC-3 cells, whereas ROS levels were not affected in DU-145 cells. Salubrinal, an inhibitor of eIF2α dephosphorylation and ER stress, reduced LCA-induced CHOP levels slightly in PC-3, but not DU-145 cells. Salubrinal pre-treatment increased the cytotoxicity of LCA in PC-3 and DU-145 cells and resulted in a statistically significant loss of cell viability at normally non-toxic concentrations of LCA. The late-stage autophagy inhibitor bafilomycin A1 exacerbated LCA toxicity at subtoxic LCA concentrations in PC-3 cells. The antioxidant α-tocotrienol strongly inhibited the toxicity of LCA in PC-3 cells, but not in DU-145 cells. Collectively, although LCA induces autophagy and ER stress in PC-3 cells, these processes appear to be initially of protective nature and subsequently consequential to, but not critical for the ROS-mediated mitochondrial dysfunction and cytotoxicity of LCA. The full mechanism of LCA-induced mitochondrial dysfunction and cytotoxicity in the similarly sensitive DU-145 cells remains to be elucidated. creator: Ahmed A. Gafar creator: Hossam M. Draz creator: Alexander A. Goldberg creator: Mohamed A. Bashandy creator: Sayed Bakry creator: Mahmoud A. Khalifa creator: Walid AbuShair creator: Vladimir I. Titorenko creator: J. Thomas Sanderson uri: https://doi.org/10.7717/peerj.2445 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Gafar et al. title: Identification of phenolic secondary metabolites from Schotia brachypetala Sond. (Fabaceae) and demonstration of their antioxidant activities in Caenorhabditis elegans link: https://peerj.com/articles/2404 last-modified: 2016-11-15 description: BackgroundSchotia brachypetala Sond. (Fabaceae) is an endemic tree of Southern Africa whose phytochemistry and pharmacology were slightly studied. The present work aimed at profiling the major phenolics compounds present in the hydro-alcohol extract from S. brachypetala leaves (SBE) using LC/HRESI/MS/MS and NMR and prove their antioxidant capabilities using novel methods.MethodsIn vitro assays; DPPH, TEAC persulfate decolorizing kinetic and FRAP assays, and in vivo assays: Caenorhabditis elegans strains maintenance, Intracellular ROS in C. elegans, Survival assay, GFP expression and Subcellular DAF-16 localization were employed to evaluate the antioxidant activity.ResultsMore than forty polyphenols, including flavonoid glycosides, galloylated flavonoid glycosides, isoflavones, dihydrochalcones, procyanidins, anthocyanins, hydroxy benzoic acid derivatives, hydrolysable tannins, and traces of methylated and acetylated flavonoid derivatives were identified. Three compounds were isolated and identified from the genus Schotia for the first time, namely gallic acid, myricetin-3-O-α-L-1C4-rhamnoside and quercetin-3-O-L-1C4-rhamnoside. The total phenolics content of SBE was (376 mg CAE/g), followed by flavonoids (67.87 QE/g). In vitro antioxidant activity of SBE was evidenced by DPPH radical scavenging activity (IC50 of 9 µg/mL), FRAP ferric reducing activity (5,000 mol Fe2+ E/mg) and ABTS peroxide inhibiting activity (1,054 mM Trolox E/mg). The tested extract was able to protect the worms against juglone induced oxidative stress, an increased survival rate (up to 41%) was recorded, when compared with the control group (11%) and attenuate the reactive oxygen species (ROS) accumulation in dose-dependent and reached up to 72% for the highest tested concentration. SBE was also able to attenuate the levels of heat shock protein (HSP) expression in dose-dependent up to 60% in the 150 µg SBE/mL group. In DAF-16 Subcellular localization SBE treated worms showed nuclear localization pattern up to 78%, while it was only 5% in the untreated control group.DiscussionA pronounced antioxidant activity in vivo, which can be attributed to its ability to promote the nuclear translocation of DAF-16/FOXO, the main transcription factor regulating the expression of stress response genes. The remarkable antioxidant activity in vitro and in vivo correlates to SBE rich phenolic profile. creator: Mansour Sobeh creator: Esraa ElHawary creator: Herbenya Peixoto creator: Rola M. Labib creator: Heba Handoussa creator: Noha Swilam creator: Ahmed H. El-Khatib creator: Farukh Sharapov creator: Tamer Mohamed creator: Sonja Krstin creator: Michael W. Linscheid creator: Abdel Nasser Singab creator: Michael Wink creator: Nahla Ayoub uri: https://doi.org/10.7717/peerj.2404 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Sobeh et al. title: Comparative analysis of the complete chloroplast genome sequences in psammophytic Haloxylon species (Amaranthaceae) link: https://peerj.com/articles/2699 last-modified: 2016-11-10 description: The Haloxylon genus belongs to the Amaranthaceae (formerly Chenopodiaceae) family. The small trees or shrubs in this genus are referred to as the King of psammophytic plants, and perform important functions in environmental protection, including wind control and sand fixation in deserts. To better understand these beneficial plants, we sequenced the chloroplast (cp) genomes of Haloxylon ammodendron (HA) and Haloxylon persicum (HP) and conducted comparative genomic analyses on these and two other representative Amaranthaceae species. Similar to other higher plants, we found that the Haloxylon cp genome is a quadripartite, double-stranded, circular DNA molecule of 151,570 bp in HA and 151,586 bp in HP. It contains a pair of inverted repeats (24,171 bp in HA and 24,177 bp in HP) that separate the genome into a large single copy region of 84,214 bp in HA and 84,217 bp in HP, and a small single copy region of 19,014 bp in HA and 19,015 bp in HP. Each Haloxylon cp genome contains 112 genes, including 78 coding, 30 tRNA, and four ribosomal RNA genes. We detected 59 different simple sequence repeat loci, including 44 mono-nucleotide, three di-nucleotide, one tri-nucleotide, and 11 tetra-nucleotide repeats. Comparative analysis revealed only 67 mutations between the two species, including 44 substitutions, 23 insertions/deletions, and two micro-inversions. The two inversions, with lengths of 14 and 3 bp, occur in the petA-psbJ intergenic region and rpl16 intron, respectively, and are predicted to form hairpin structures with repeat sequences of 27 and 19 bp, respectively, at the two ends. The ratio of transitions to transversions was 0.76. These results are valuable for future studies on Haloxylon genetic diversity and will enhance our understanding of the phylogenetic evolution of Amaranthaceae. creator: Wenpan Dong creator: Chao Xu creator: Delu Li creator: Xiaobai Jin creator: Ruili Li creator: Qi Lu creator: Zhili Suo uri: https://doi.org/10.7717/peerj.2699 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Dong et al. title: Soymilk residue (okara) as a natural immobilization carrier for Lactobacillus plantarum cells enhances soymilk fermentation, glucosidic isoflavone bioconversion, and cell survival under simulated gastric and intestinal conditions link: https://peerj.com/articles/2701 last-modified: 2016-11-10 description: Cell immobilization is an alternative to microencapsulation for the maintenance of cells in a liquid medium. However, artificial immobilization carriers are expensive and pose a high safety risk. Okara, a food-grade byproduct from soymilk production, is rich in prebiotics. Lactobacilli could provide health enhancing effects to the host. This study aimed to evaluate the potential of okara as a natural immobilizer for L. plantarum 70810 cells. The study also aimed to evaluate the effects of okara-immobilized L. plantarum 70810 cells (IL) on soymilk fermentation, glucosidic isoflavone bioconversion, and cell resistance to simulated gastric and intestinal stresses. Scanning electron microscopy (SEM) was used to show cells adherence to the surface of okara. Lactic acid, acetic acid and isoflavone analyses in unfermented and fermented soymilk were performed by HPLC with UV detection. Viability and growth kinetics of immobilized and free L. plantarum 70810 cells (FL) were followed during soymilk fermentation. Moreover, changes in pH, titrable acidity and viscosity were measured by conventional methods. For in vitro testing of simulated gastrointestinal resistance, fermented soymilk was inoculated with FL or IL and an aliquot incubated into acidic MRS broth which was conveniently prepared to simulate gastric, pancreatic juices and bile salts. Survival to simulated gastric and intestinal stresses was evaluated by plate count of colony forming units on MRS agar. SEM revealed that the lactobacilli cells attached and bound to the surface of okara. Compared with FL, IL exhibited a significantly higher specific growth rate, shorter lag phase of growth, higher productions of lactic and acetic acids, a faster decrease in pH and increase in titrable acidity, and a higher soymilk viscosity. Similarly, IL in soymilk showed higher productions of daizein and genistein compared with the control. Compared with FL, IL showed reinforced resistance to simulatedgastric and intestinal stresses in vitro that included low pH, low pH plus pepsin, pancreatin, and bile salt. Our results indicate that okara is a new potential immobilization carrier to enhance the growth and glucosidic isoflavone bioconversion activities of L. plantarum in soymilk and improve cell survivability following simulated gastric and intestinal conditions. creator: Xia Xiudong creator: Wang Ying creator: Liu Xiaoli creator: Li Ying creator: Zhou Jianzhong uri: https://doi.org/10.7717/peerj.2701 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Xiudong et al. title: Induced pluripotent stem cells from human hair follicle keratinocytes as a potential source for in vitro hair follicle cloning link: https://peerj.com/articles/2695 last-modified: 2016-11-10 description: BackgroundHuman hair follicles are important for the renewal of new hairs and their development. The generation of induced pluripotent stem cells (iPSCs) from hair follicles is easy due to its accessibility and availability. The pluripotent cells derived from hair follicles not only have a higher tendency to re-differentiate into hair follicles, but are also more suited for growth in hair scalp tissue microenvironment.MethodsIn this study, human hair follicular keratinocytes were used to generate iPSCs, which were then further differentiated in vitro into keratinocytes. The derived iPSCs were characterised by using immunofluorescence staining, flow cytometry, and reverse-transcription PCR to check for its pluripotency markers expression.ResultsThe iPSC clones expressed pluripotency markers such as TRA-1-60, TRA-1-81, SSEA4, OCT4, SOX2, NANOG, LEFTY, and GABRB. The well-formed three germ layers were observed during differentiation using iPSCs derived from hair follicles. The successful formation of keratioctyes from iPSCs was confirmed by the expression of cytokeratin 14 marker.DiscussionHair follicles represent a valuable keratinocytes source for in vitro hair cloning for use in treating hair balding or grafting in burn patients. Our significant findings in this report proved that hair follicles could be used to produce pluripotent stem cells and suggested that the genetic and micro-environmental elements of hair follicles might trigger higher and more efficient hair follicles re-differentiation. creator: Sheng Jye Lim creator: Shu Cheow Ho creator: Pooi Ling Mok creator: Kian Lee Tan creator: Alan H.K. Ong creator: Seng Chiew Gan uri: https://doi.org/10.7717/peerj.2695 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Lim et al. title: Reanalysis of the anthrax epidemic in Rhodesia, 1978–1984 link: https://peerj.com/articles/2686 last-modified: 2016-11-10 description: In the mid-1980s, the largest epidemic of anthrax of the last 200 years was documented in a little known series of studies by Davies in The Central African Journal of Medicine. This epidemic involved thousands of cattle and 10,738 human cases with 200 fatalities in Rhodesia during the Counterinsurgency. Grossly unusual epidemiological features were noted that, to this day, have not been definitively explained. This study performed a historical reanalysis of the data to reveal an estimated geographic involvement of 245,750 km2, with 171,990 cattle and 17,199 human cases. Here we present the first documented geotemporal visualization of the human anthrax epidemic. creator: James M. Wilson creator: Walter Brediger creator: Thomas P. Albright creator: Julie Smith-Gagen uri: https://doi.org/10.7717/peerj.2686 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Wilson et al. title: Sexual reproduction in the Caribbean coral genus Isophyllia (Scleractinia: Mussidae) link: https://peerj.com/articles/2665 last-modified: 2016-11-10 description: The sexual pattern, reproductive mode, and timing of reproduction of Isophyllia sinuosa and Isophyllia rigida, two Caribbean Mussids, were assessed by histological analysis of specimens collected monthly during 2000–2001. Both species are simultaneous hermaphroditic brooders characterized by a single annual gametogenetic cycle. Spermatocytes and oocytes of different stages were found to develop within the same mesentery indicating sequential maturation for extended planulation. Oogenesis took place during May through April in I. sinuosa and from August through June in I. rigida. Oocytes began development 7–8 months prior to spermaries but both sexes matured simultaneously. Zooxanthellate planulae were observed in I. sinuosa during April and in I. rigida from June through September. Higher polyp and mesenterial fecundity were found in I. rigida compared to I. sinuosa. Larger oocyte sizes were found in I. sinuosa than in I. rigida, however larger planula sizes were found in I. rigida. Hermaphroditism is the exclusive sexual pattern within the Mussidae while brooding has been documented within the related genera Mussa, Scolymia and Mycetophyllia. This study represents the first description of the sexual characteristics of I. rigida and provides an updated description of I. sinuosa. creator: Derek Soto creator: Ernesto Weil uri: https://doi.org/10.7717/peerj.2665 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Soto and Weil title: Hyainailourine and teratodontine cranial material from the late Eocene of Egypt and the application of parsimony and Bayesian methods to the phylogeny and biogeography of Hyaenodonta (Placentalia, Mammalia) link: https://peerj.com/articles/2639 last-modified: 2016-11-10 description: Hyaenodonta is a diverse, extinct group of carnivorous mammals that included weasel- to rhinoceros-sized species. The oldest-known hyaenodont fossils are from the middle Paleocene of North Africa and the antiquity of the group in Afro-Arabia led to the hypothesis that it originated there and dispersed to Asia, Europe, and North America. Here we describe two new hyaenodont species based on the oldest hyaenodont cranial specimens known from Afro-Arabia. The material was collected from the latest Eocene Locality 41 (L-41, ∼34 Ma) in the Fayum Depression, Egypt. Akhnatenavus nefertiticyon sp. nov. has specialized, hypercarnivorous molars and an elongate cranial vault. In A. nefertiticyon the tallest, piercing cusp on M1–M2 is the paracone. Brychotherium ephalmos gen. et sp. nov. has more generalized molars that retain the metacone and complex talonids. In B. ephalmos the tallest, piercing cusp on M1–M2 is the metacone. We incorporate this new material into a series of phylogenetic analyses using a character-taxon matrix that includes novel dental, cranial, and postcranial characters, and samples extensively from the global record of the group. The phylogenetic analysis includes the first application of Bayesian methods to hyaenodont relationships. B. ephalmos is consistently placed within Teratodontinae, an Afro-Arabian clade with several generalist and hypercarnivorous forms, and Akhnatenavus is consistently recovered in Hyainailourinae as part of an Afro-Arabian radiation. The phylogenetic results suggest that hypercarnivory evolved independently three times within Hyaenodonta: in Teratodontinae, in Hyainailourinae, and in Hyaenodontinae. Teratodontines are consistently placed in a close relationship with Hyainailouridae (Hyainailourinae + Apterodontinae) to the exclusion of “proviverrines,” hyaenodontines, and several North American clades, and we propose that the superfamily Hyainailouroidea be used to describe this relationship. Using the topologies recovered from each phylogenetic method, we reconstructed the biogeographic history of Hyaenodonta using parsimony optimization (PO), likelihood optimization (LO), and Bayesian Binary Markov chain Monte Carlo (MCMC) to examine support for the Afro-Arabian origin of Hyaenodonta. Across all analyses, we found that Hyaenodonta most likely originated in Europe, rather than Afro-Arabia. The clade is estimated by tip-dating analysis to have undergone a rapid radiation in the Late Cretaceous and Paleocene; a radiation currently not documented by fossil evidence. During the Paleocene, lineages are reconstructed as dispersing to Asia, Afro-Arabia, and North America. The place of origin of Hyainailouroidea is likely Afro-Arabia according to the Bayesian topologies but it is ambiguous using parsimony. All topologies support the constituent clades–Hyainailourinae, Apterodontinae, and Teratodontinae–as Afro-Arabian and tip-dating estimates that each clade is established in Afro-Arabia by the middle Eocene. creator: Matthew R. Borths creator: Patricia A. Holroyd creator: Erik R. Seiffert uri: https://doi.org/10.7717/peerj.2639 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Borths et al. title: Pico and nanoplankton abundance and carbon stocks along the Brazilian Bight link: https://peerj.com/articles/2587 last-modified: 2016-11-10 description: Pico and nanoplankton communities from the Southwest Atlantic Ocean along the Brazilian Bight are poorly described. The hydrography in this region is dominated by a complex system of layered water masses, which includes the warm and oligotrophic Tropical Water (TW), the cold and nutrient rich South Atlantic Central Water (SACW) and the Coastal Water (CW), which have highly variable properties. In order to assess how pico- and nanoplankton communities are distributed in these different water masses, we determined by flow cytometry the abundance of heterotrophic bacteria, Prochlorococcus, Synechococcus and autotrophic pico and nanoeukaryotes along three transects, extending from 23°S to 31°S and 39°W to 49°W. Heterotrophic bacteria (including archaea, maximum of 1.5 × 106 cells mL−1) were most abundant in Coastal and Tropical Water whereas Prochlorococcus was most abundant in open-ocean oligotrophic waters (maximum of 300 × 103 cells mL−1). Synechococcus(up to 81 × 103 cells mL−1), as well as autotrophic pico and nanoeukaryotes seemed to benefit from the influx of nutrient-rich waters near the continental slope. Autotrophic pico and nanoeukaryotes were also abundant in deep chlorophyll maximum (DCM) layers from offshore waters, and their highest abundances were 20 × 103 cells mL−1 and 5 × 103 cells mL−1, respectively. These data are consistent with previous observations in other marine areas where Synechococcus and autotrophic eukaryotes dominate mesotrophic waters, whereas Prochlorococcus dominate in more oligotrophic areas. Regardless of the microbial community structure near the surface, the carbon stock dominance by autotrophic picoeukaryotes near the DCM is possibly linked to vertical mixing of oligotrophic surface waters with the nutrient-rich SACW and their tolerance to lower light levels. creator: Catherine Gérikas Ribeiro creator: Adriana Lopes dos Santos creator: Dominique Marie creator: Vivian Helena Pellizari creator: Frederico Pereira Brandini creator: Daniel Vaulot uri: https://doi.org/10.7717/peerj.2587 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Gérikas Ribeiro et al. title: Factors influencing infection and transmission of Anopheles gambiae densovirus (AgDNV) in mosquitoes link: https://peerj.com/articles/2691 last-modified: 2016-11-09 description: Anopheles gambiae densovirus (AgDNV) is a potential microbial agent for paratransgenesis and gene transduction in An. gambiae, the major vector of human malaria in sub-Saharan Africa. Understanding the interaction between AgDNV and An. gambiae is critical for using AgDNV in a basic and applied manner for Anopheles gene manipulation. Here, we tested the effects of mosquito age, sex, blood feeding status, and potential for horizontal transmission using an enhanced green fluorescent protein (EGFP) reporter AgDNV system. Neither mosquito age at infection nor feeding regime affected viral titers. Female mosquitoes were more permissive to viral infection than males. Despite low viral titers, infected males were able to venereally transmit virus to females during mating, where the virus was localized with the transferred sperm in the spermathecae. These findings will be useful for designing AgDNV-based strategies to manipulate Anopheles gambiae. creator: Tapan K. Barik creator: Yasutsugu Suzuki creator: Jason L. Rasgon uri: https://doi.org/10.7717/peerj.2691 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Barik et al. title: Charge neutralization as the major factor for the assembly of nucleocapsid-like particles from C-terminal truncated hepatitis C virus core protein link: https://peerj.com/articles/2670 last-modified: 2016-11-09 description: BackgroundHepatitis C virus (HCV) core protein, in addition to its structural role to form the nucleocapsid assembly, plays a critical role in HCV pathogenesis by interfering in several cellular processes, including microRNA and mRNA homeostasis. The C-terminal truncated HCV core protein (C124) is intrinsically unstructured in solution and is able to interact with unspecific nucleic acids, in the micromolar range, and to assemble into nucleocapsid-like particles (NLPs) in vitro. The specificity and propensity of C124 to the assembly and its implications on HCV pathogenesis are not well understood.MethodsSpectroscopic techniques, transmission electron microscopy and calorimetry were used to better understand the propensity of C124 to fold or to multimerize into NLPs when subjected to different conditions or in the presence of unspecific nucleic acids of equivalent size to cellular microRNAs.ResultsThe structural analysis indicated that C124 has low propensity to self-folding. On the other hand, for the first time, we show that C124, in the absence of nucleic acids, multimerizes into empty NLPs when subjected to a pH close to its isoelectric point (pH ≈ 12), indicating that assembly is mainly driven by charge neutralization. Isothermal calorimetry data showed that the assembly of NLPs promoted by nucleic acids is enthalpy driven. Additionally, data obtained from fluorescence correlation spectroscopy show that C124, in nanomolar range, was able to interact and to sequester a large number of short unspecific nucleic acids into NLPs.DiscussionTogether, our data showed that the charge neutralization is the major factor for the nucleocapsid-like particles assembly from C-terminal truncated HCV core protein. This finding suggests that HCV core protein may physically interact with unspecific cellular polyanions, which may correspond to microRNAs and mRNAs in a host cell infected by HCV, triggering their confinement into infectious particles. creator: Theo Luiz Ferraz de Souza creator: Sheila Maria Barbosa de Lima creator: Vanessa L. de Azevedo Braga creator: David S. Peabody creator: Davis Fernandes Ferreira creator: M. Lucia Bianconi creator: Andre Marco de Oliveira Gomes creator: Jerson Lima Silva creator: Andréa Cheble de Oliveira uri: https://doi.org/10.7717/peerj.2670 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 De Souza et al. title: Low self-concept in poor readers: prevalence, heterogeneity, and risk link: https://peerj.com/articles/2669 last-modified: 2016-11-09 description: There is evidence that poor readers are at increased risk for various types of low self-concept—particularly academic self-concept. However, this evidence ignores the heterogeneous nature of poor readers, and hence the likelihood that not all poor readers have low self-concept. The aim of this study was to better understand which types of poor readers have low self-concept. We tested 77 children with poor reading for their age for four types of self-concept, four types of reading, three types of spoken language, and two types of attention. We found that poor readers with poor attention had low academic self-concept, while poor readers with poor spoken language had low general self-concept in addition to low academic self-concept. In contrast, poor readers with typical spoken language and attention did not have low self-concept of any type. We also discovered that academic self-concept was reliably associated with reading and receptive spoken vocabulary, and that general self-concept was reliably associated with spoken vocabulary. These outcomes suggest that poor readers with multiple impairments in reading, language, and attention are at higher risk for low academic and general self-concept, and hence need to be assessed for self-concept in clinical practice. Our results also highlight the need for further investigation into the heterogeneous nature of self-concept in poor readers. creator: Genevieve McArthur creator: Anne Castles creator: Saskia Kohnen creator: Erin Banales uri: https://doi.org/10.7717/peerj.2669 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 McArthur et al. title: A global analysis of bird plumage patterns reveals no association between habitat and camouflage link: https://peerj.com/articles/2658 last-modified: 2016-11-09 description: Evidence suggests that animal patterns (motifs) function in camouflage. Irregular mottled patterns can facilitate concealment when stationary in cluttered habitats, whereas regular patterns typically prevent capture during movement in open habitats. Bird plumage patterns have predominantly converged on just four types—mottled (irregular), scales, bars and spots (regular)—and habitat could be driving convergent evolution in avian patterning. Based on sensory ecology, we therefore predict that irregular patterns would be associated with visually noisy closed habitats and that regular patterns would be associated with open habitats. Regular patterns have also been shown to function in communication for sexually competing males to stand-out and attract females, so we predict that male breeding plumage patterns evolved in both open and closed habitats. Here, taking phylogenetic relatedness into account, we investigate ecological selection for bird plumage patterns across the class Aves. We surveyed plumage patterns in 80% of all avian species worldwide. Of these, 2,756 bird species have regular and irregular plumage patterns as well as habitat information. In this subset, we tested whether adult breeding/non-breeding plumages in each sex, and juvenile plumages, were associated with the habitat types found within the species’ geographical distributions. We found no evidence for an association between habitat and plumage patterns across the world’s birds and little phylogenetic signal. We also found that species with regular and irregular plumage patterns were distributed randomly across the world’s eco-regions without being affected by habitat type. These results indicate that at the global spatial and taxonomic scale, habitat does not predict convergent evolution in bird plumage patterns, contrary to the camouflage hypothesis. creator: Marius Somveille creator: Kate L.A. Marshall creator: Thanh-Lan Gluckman uri: https://doi.org/10.7717/peerj.2658 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Somveille et al. title: The potential for self-seeding by the coral Pocillopora spp. in Moorea, French Polynesia link: https://peerj.com/articles/2544 last-modified: 2016-11-09 description: Coral reefs in Moorea, French Polynesia, suffered catastrophic coral mortality through predation by Acanthaster planci from 2006 to 2010, and Cyclone Oli in 2010, yet by 2015 some coral populations were approaching pre-disturbance sizes. Using long-term study plots, we quantified population dynamics of spawning Pocillopora spp. along the north shore of Moorea between 2010 and 2014, and considered evidence that population recovery could be supported by self-seeding. Results scaled up from study plots and settlement tiles suggest that the number of Pocillopora spp. colonies on the outer reef increased 1,890-fold between 2010 and 2014/2015, and in the back reef, 8-fold between 2010 and 2014/2015. Assuming that spawning Pocillopora spp. in Moorea release similar numbers of eggs as con-generics in Hawaii, and fertilization success is similar to other spawning corals, the capacity of Pocillopora spp. to produce larvae was estimated. These estimates suggest that Pocillopora spp. in Moorea produced a large excess of larvae in 2010 and 2014 relative to the number required to produce the recruits found in the back reef and outer reef in 2010 and 2014, even assuming that ∼99.9% of the larvae do not recruit in Moorea. Less than a third of the recruits in one year would have to survive to produce the juvenile Pocillopora spp. found in the back and outer reefs in 2010 and 2014/2015. Our first order approximations reveal the potential for Pocillopora spp. on the north shore of Moorea to produce enough larvae to support local recruitment and population recovery following a catastrophic disturbance. creator: Georgios Tsounis creator: Peter J. Edmunds uri: https://doi.org/10.7717/peerj.2544 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Tsounis and Edmunds title: The origin of the lower fourth molar in canids, inferred by individual variation link: https://peerj.com/articles/2689 last-modified: 2016-11-08 description: BackgroundAn increase in tooth number is an exception during mammalian evolution. The acquisition of the lower fourth molar in the bat-eared fox (Otocyon megalotis, Canidae, Carnivora, Mammalia) is one example; however, its developmental origin is not clear. In some canids (Canidae), individual variation exist as supernumerary molar M4. This study focuses on the acquisition of the lower fourth molar in canids and proposes that the inhibitory cascade model can explain its origin.MethodsOcclusal view projected area of lower molars was determined from 740 mandibles obtained from Canis latrans, Nyctereutes procyonoides, and Urocyon cinereoargenteus museum specimens. For each molar, relative sizes of molars (M2/M1 and M3/M1 scores) affected by inhibition/activation dynamics during development, were compared between individuals with and without supernumerary molar (M4).ResultsPossession of a supernumerary molar was associated with significantly larger M2/M1 score in Canis latrans, M3/M1 score in Nyctereutes procyonoides, and M2/M1 and M3/M1 scores in Urocyon cinereoargenteus compared to individuals of these species that lacked supernumerary molars.DiscussionWe propose that, in canids, the supernumerary fourth molar is attributable to reduced inhibition and greater activation during molar development. In the bat-eared fox, altered inhibition and activation dynamics of dental development during omnivorous-insectivorous adaptation may be a contributing factor in the origin of the lower fourth molar. creator: Masakazu Asahara uri: https://doi.org/10.7717/peerj.2689 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Asahara title: Antioxidant nutrition in Atlantic salmon (Salmo salar) parr and post-smolt, fed diets with high inclusion of plant ingredients and graded levels of micronutrients and selected amino acids link: https://peerj.com/articles/2688 last-modified: 2016-11-08 description: The shift from marine to plant-based ingredients in fish feeds affects the dietary concentrations and bioavailability of micronutrients, amino acids and lipids and consequently warrants a re-evaluation of dietary nutrient recommendations. In the present study, an Atlantic salmon diet high in plant ingredients was supplemented with graded levels of nutrient premix (NP), containing selected amino acids, taurine, cholesterol, vitamins and minerals. This article presents the results on the antioxidant nutrients vitamin C, E and selenium (Se), and effects on tissue redox status. The feed ingredients appeared to contain sufficient levels of vitamin E and Se to cover the requirements to prevent clinical deficiency symptoms. The body levels of α-tocopherol (TOH) in parr and that of Se in parr and post-smolt showed a linear relationship with dietary concentration, while α-TOH in post-smolt seemed to be saturable with a breakpoint near 140 mg kg−1. Ascorbic acid (Asc) concentration in the basal feed was below the expected minimum requirement, but the experimental period was probably too short for the fish to develop visible deficiency symptoms. Asc was saturable in both parr and post-smolt whole body at dietary concentrations of 190 and 63–89 mg kg−1, respectively. Maximum whole body Asc concentration was approximately 40 mg kg−1 in parr and 14 mg kg−1 in post-smolt. Retention ranged from 41 to 10% in parr and from −206 to 12% in post-smolt with increasing NP supplementation. This indicates that the post-smolts had an extraordinarily high consumption of Asc. Analyses of glutathione (GSH) and glutathione disulphide (GSSG) concentrations and the calculated GSH based redox potentials in liver and muscle tissue, indicated only minor effects of diets on redox regulation. However, the post-smolt were more oxidized than the parr. This was supported by the high consumption of Asc and high expression of gpx1 and gpx3 in liver. Based on the present trials, the recommendations for supplementation of vitamin C and E in diets for Atlantic salmon are similar to current practices, e.g. 150 mg kg−1 of α-TOH and 190 mg kg−1 Asc which was the saturating concentration in parr. Higher concentrations than what would prevent clinical deficiency symptoms are necessary to protect fish against incidents of oxidative stress and to improve immune and stress responses. There were no indications that the Se requirement exceeded the current recommendation of 0.3 mg kg−1. creator: Kristin Hamre creator: Nini H. Sissener creator: Erik-Jan Lock creator: Pål A. Olsvik creator: Marit Espe creator: Bente E. Torstensen creator: Joana Silva creator: Johan Johansen creator: Rune Waagbø creator: Gro-Ingunn Hemre uri: https://doi.org/10.7717/peerj.2688 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Hamre et al. title: Proteogenomic analyses indicate bacterial methylotrophy and archaeal heterotrophy are prevalent below the grass root zone link: https://peerj.com/articles/2687 last-modified: 2016-11-08 description: Annually, half of all plant-derived carbon is added to soil where it is microbially respired to CO2. However, understanding of the microbiology of this process is limited because most culture-independent methods cannot link metabolic processes to the organisms present, and this link to causative agents is necessary to predict the results of perturbations on the system. We collected soil samples at two sub-root depths (10–20 cm and 30–40 cm) before and after a rainfall-driven nutrient perturbation event in a Northern California grassland that experiences a Mediterranean climate. From ten samples, we reconstructed 198 metagenome-assembled genomes that represent all major phylotypes. We also quantified 6,835 proteins and 175 metabolites and showed that after the rain event the concentrations of many sugars and amino acids approach zero at the base of the soil profile. Unexpectedly, the genomes of novel members of the Gemmatimonadetes and Candidate Phylum Rokubacteria phyla encode pathways for methylotrophy. We infer that these abundant organisms contribute substantially to carbon turnover in the soil, given that methylotrophy proteins were among the most abundant proteins in the proteome. Previously undescribed Bathyarchaeota and Thermoplasmatales archaea are abundant in deeper soil horizons and are inferred to contribute appreciably to aromatic amino acid degradation. Many of the other bacteria appear to breakdown other components of plant biomass, as evidenced by the prevalence of various sugar and amino acid transporters and corresponding hydrolyzing machinery in the proteome. Overall, our work provides organism-resolved insight into the spatial distribution of bacteria and archaea whose activities combine to degrade plant-derived organics, limiting the transport of methanol, amino acids and sugars into underlying weathered rock. The new insights into the soil carbon cycle during an intense period of carbon turnover, including biogeochemical roles to previously little known soil microbes, were made possible via the combination of metagenomics, proteomics, and metabolomics. creator: Cristina N. Butterfield creator: Zhou Li creator: Peter F. Andeer creator: Susan Spaulding creator: Brian C. Thomas creator: Andrea Singh creator: Robert L. Hettich creator: Kenwyn B. Suttle creator: Alexander J. Probst creator: Susannah G. Tringe creator: Trent Northen creator: Chongle Pan creator: Jillian F. Banfield uri: https://doi.org/10.7717/peerj.2687 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Butterfield et al. title: Cranial bone histology of Metoposaurus krasiejowensis (Amphibia, Temnospondyli) from the Late Triassic of Poland link: https://peerj.com/articles/2685 last-modified: 2016-11-08 description: In this study, 21 skull bones of Metoposaurus krasiejowensis from the Late Triassic of Poland were investigated histologically. Dermal bones show a diploë structure, with an ornamented external surface. The ridges consist of mostly well vascularized parallel-fibered bone; the valleys are built of an avascular layer of lamellar bone. The thick middle region consists of cancellous bone, with varying porosity. The thin and less vascularized internal cortex consists of parallel-fibered bone. The numerous Sharpey’s fibers and ISF are present in all bones. The cyclicity of growth is manifested as an alternation of thick, avascular annuli and high vascularized zones as well as a sequence of resting lines. The detailed histological framework of dermal bones varies even within a single bone; this seems to be related to the local biomechanical loading of the particular part of the skull. The dynamic processes observed during the ornamentation creation indicate that the positions of the ridges and grooves change during growth and could be a specific adaptation to changing biomechanical conditions and stress distribution during bone development. In the supratemporal, the cementing lines show that the remodeling process could be involved in the creations of sculpture. The common occurrence of ISF suggests that metaplastic ossification plays an important role during cranial development. Endochondral bones preserved the numerous remains of calcified cartilage. This indicates that ossification follows a pattern known for stereospondyl intercentra, with relatively slow ossification of the trabecular part and late development of the periosteal cortex. The large accumulation of Sharpey’s fibers in the occipital condyles indicates the presence of strong muscles and ligaments connecting the skull to the vertebral column. creator: Kamil Gruntmejer creator: Dorota Konietzko-Meier creator: Adam Bodzioch uri: https://doi.org/10.7717/peerj.2685 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Gruntmejer et al. title: Comparisons of forecasting for hepatitis in Guangxi Province, China by using three neural networks models link: https://peerj.com/articles/2684 last-modified: 2016-11-08 description: This study compares and evaluates the prediction of hepatitis in Guangxi Province, China by using back propagation neural networks based genetic algorithm (BPNN-GA), generalized regression neural networks (GRNN), and wavelet neural networks (WNN). In order to compare the results of forecasting, the data obtained from 2004 to 2013 and 2014 were used as modeling and forecasting samples, respectively. The results show that when the small data set of hepatitis has seasonal fluctuation, the prediction result by BPNN-GA will be better than the two other methods. The WNN method is suitable for predicting the large data set of hepatitis that has seasonal fluctuation and the same for the GRNN method when the data increases steadily. creator: Ruijing Gan creator: Ni Chen creator: Daizheng Huang uri: https://doi.org/10.7717/peerj.2684 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Gan et al. title: RGFA: powerful and convenient handling of assembly graphs link: https://peerj.com/articles/2681 last-modified: 2016-11-08 description: The “Graphical Fragment Assembly” (GFA) is an emerging format for the representation of sequence assembly graphs, which can be adopted by both de Bruijn graph- and string graph-based assemblers. Here we present RGFA, an implementation of the proposed GFA specification in Ruby. It allows the user to conveniently parse, edit and write GFA files. Complex operations such as the separation of the implicit instances of repeats and the merging of linear paths can be performed. A typical application of RGFA is the editing of a graph, to finish the assembly of a sequence, using information not available to the assembler. We illustrate a use case, in which the assembly of a repetitive metagenomic fosmid insert was completed using a script based on RGFA. Furthermore, we show how the API provided by RGFA can be employed to design complex graph editing algorithms. As an example, we developed a detection algorithm for CRISPRs in a de Bruijn graph. Finally, RGFA can be used for comparing assembly graphs, e.g., to document the changes in a graph after applying a GUI editor. A program, GFAdiff is provided, which compares the information in two graphs, and generate a report or a Ruby script documenting the transformation steps between the graphs. creator: Giorgio Gonnella creator: Stefan Kurtz uri: https://doi.org/10.7717/peerj.2681 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Gonnella and Kurtz title: Community assembly and functional leaf traits mediate precipitation use efficiency of alpine grasslands along environmental gradients on the Tibetan Plateau link: https://peerj.com/articles/2680 last-modified: 2016-11-08 description: The alpine grasslands on the Tibetan Plateau are sensitive and vulnerable to climate change. However, it is still unknown how precipitation use efficiency (PUE), the ratio of aboveground net primary productivity (ANPP) to precipitation, is related to community assembly of plant species, functional groups or traits for the Tibetan alpine grasslands along actual environmental gradients. We conducted a multi-site field survey at grazing-excluded pastures across meadow, steppe and desert-steppe to measure aboveground biomass (AGB) in August, 2010. We used species richness (SR), the Shannon diversity index, and cover-weighted functional group composition (FGC) of 1-xerophytes, 2-mesophytes, and 3-hygrophytes to describe community assembly at the species level; and chose community-level leaf area index (LAIc), specific leaf area (SLAc), and species-mixed foliar δ13C to quantify community assembly at the functional trait level. Our results showed that PUE decreased with increasing accumulated active temperatures (AccT) when daily temperature average is higher than 5 °C, but increased with increasing climatic moisture index (CMI), which was demined as the ratio of growing season precipitation (GSP) to AccT. We also found that PUE increased with increasing SR, the Shannon diversity index, FGC and LAIc, decreased with increasing foliar δ13C, and had no relation with SLAc at the regional scale. Neither soil total nitrogen (STN) nor organic carbon has no influence on PUE at the regional scale. The community assembly of the Shannon index, LAIc and SLAc together accounted for 46.3% of variance in PUE, whilst CMI accounted for 47.9% of variance in PUE at the regional scale. This implies that community structural properties and plant functional traits can mediate the sensitivity of alpine grassland productivity in response to climate change. Thus, a long-term observation on community structural and functional changes is recommended for better understanding the response of alpine ecosystems to regional climate change on the Tibetan Plateau. creator: Shaowei Li creator: Jianshuang Wu uri: https://doi.org/10.7717/peerj.2680 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Li and Wu title: Cloning, expression and characterization of a cold-adapted endo-1, 4-β-glucanase from Citrobacter farmeri A1, a symbiotic bacterium of Reticulitermes labralis link: https://peerj.com/articles/2679 last-modified: 2016-11-08 description: BackgroundMany biotechnological and industrial applications can benefit from cold-adapted EglCs through increased efficiency of catalytic processes at low temperature. In our previous study, Citrobacter farmeri A1 which was isolated from a wood-inhabiting termite Reticulitermes labralis could secrete a cold-adapted EglC. However, its EglC was difficult to purify for enzymatic properties detection because of its low activity (0.8 U/ml). The objective of the present study was to clone and express the C. farmeri EglC gene in Escherichia coli to improve production level and determine the enzymatic properties of the recombinant enzyme.MethodsThe EglC gene was cloned from C. farmeri A1 by thermal asymmetric interlaced PCR. EglC was transformed into vector pET22b and functionally expressed in E. coli. The recombination protein EglC22b was purified for properties detection.ResultsSDS-PAGE revealed that the molecular mass of the recombinant endoglucanase was approximately 42 kDa. The activity of the E. coli pET22b-EglC crude extract was 9.5 U/ml. Additionally, it was active at pH 6.5–8.0 with an optimum pH of 7.0. The recombinant enzyme had an optimal temperature of 30–40 °C and exhibited >50% relative activity even at 5 °C, whereas it lost approximately 90% of its activity after incubation at 60 °C for 30 min. Its activity was enhanced by Co2+ and Fe3+, but inhibited by Cd2+, Zn2+, Li+, Triton X-100, DMSO, acetonitrile, Tween 80, SDS, and EDTA.ConclusionThese biochemical properties indicate that the recombinant enzyme is a cold-adapted endoglucanase that can be used for various industrial applications. creator: Xi Bai creator: Xianjun Yuan creator: Aiyou Wen creator: Junfeng Li creator: Yunfeng Bai creator: Tao Shao uri: https://doi.org/10.7717/peerj.2679 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Bai et al. title: Planning horizon affects prophylactic decision-making and epidemic dynamics link: https://peerj.com/articles/2678 last-modified: 2016-11-08 description: The spread of infectious diseases can be impacted by human behavior, and behavioral decisions often depend implicitly on a planning horizon—the time in the future over which options are weighed. We investigate the effects of planning horizons on epidemic dynamics. We developed an epidemiological agent-based model (along with an ODE analog) to explore the decision-making of self-interested individuals on adopting prophylactic behavior. The decision-making process incorporates prophylaxis efficacy and disease prevalence with the individuals’ payoffs and planning horizon. Our results show that for short and long planning horizons individuals do not consider engaging in prophylactic behavior. In contrast, individuals adopt prophylactic behavior when considering intermediate planning horizons. Such adoption, however, is not always monotonically associated with the prevalence of the disease, depending on the perceived protection efficacy and the disease parameters. Adoption of prophylactic behavior reduces the epidemic peak size while prolonging the epidemic and potentially generates secondary waves of infection. These effects can be made stronger by increasing the behavioral decision frequency or distorting an individual’s perceived risk of infection. creator: Luis G. Nardin creator: Craig R. Miller creator: Benjamin J. Ridenhour creator: Stephen M. Krone creator: Paul Joyce creator: Bert O. Baumgaertner uri: https://doi.org/10.7717/peerj.2678 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Nardin et al. title: Deconvoluting simulated metagenomes: the performance of hard- and soft- clustering algorithms applied to metagenomic chromosome conformation capture (3C) link: https://peerj.com/articles/2676 last-modified: 2016-11-08 description: BackgroundChromosome conformation capture, coupled with high throughput DNA sequencing in protocols like Hi-C and 3C-seq, has been proposed as a viable means of generating data to resolve the genomes of microorganisms living in naturally occuring environments. Metagenomic Hi-C and 3C-seq datasets have begun to emerge, but the feasibility of resolving genomes when closely related organisms (strain-level diversity) are present in the sample has not yet been systematically characterised.MethodsWe developed a computational simulation pipeline for metagenomic 3C and Hi-C sequencing to evaluate the accuracy of genomic reconstructions at, above, and below an operationally defined species boundary. We simulated datasets and measured accuracy over a wide range of parameters. Five clustering algorithms were evaluated (2 hard, 3 soft) using an adaptation of the extended B-cubed validation measure.ResultsWhen all genomes in a sample are below 95% sequence identity, all of the tested clustering algorithms performed well. When sequence data contains genomes above 95% identity (our operational definition of strain-level diversity), a naive soft-clustering extension of the Louvain method achieves the highest performance.DiscussionPreviously, only hard-clustering algorithms have been applied to metagenomic 3C and Hi-C data, yet none of these perform well when strain-level diversity exists in a metagenomic sample. Our simple extension of the Louvain method performed the best in these scenarios, however, accuracy remained well below the levels observed for samples without strain-level diversity. Strain resolution is also highly dependent on the amount of available 3C sequence data, suggesting that depth of sequencing must be carefully considered during experimental design. Finally, there appears to be great scope to improve the accuracy of strain resolution through further algorithm development. creator: Matthew Z. DeMaere creator: Aaron E. Darling uri: https://doi.org/10.7717/peerj.2676 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 DeMaere and Darling title: Yangtze River, an insignificant genetic boundary in tufted deer (Elaphodus cephalophus): the evidence from a first population genetics study link: https://peerj.com/articles/2654 last-modified: 2016-11-08 description: Great rivers were generally looked at as the geographical barrier to gene flow for many taxonomic groups. The Yangtze River is the third largest river in the world, and flows across South China and into the East China Sea. Up until now, few studies have been carried out to evaluate its effect as a geographical barrier. In this study, we attempted to determine the barrier effect of the Yangtze River on the tufted deer (Elaphodus cephalophus) using the molecular ecology approach. Using mitochondrial DNA control region (CR) sequences and 13 nuclear microsatellite loci, we explored the genetic structure and gene flow in two adjacent tufted deer populations (Dabashan and Wulingshan populations), which are separated by the Yangtze River. Results indicated that there are high genetic diversity levels in the two populations, but no distinguishable haplotype group or potential genetic cluster was detected which corresponded to specific geographical population. At the same time, high gene flow was observed between Wulingshan and Dabashan populations. The tufted deer populations experienced population decrease from 0.3 to 0.09 Ma BP, then followed by a distinct population increase. A strong signal of recent population decline (T = 4,396 years) was detected in the Wulingshan population by a Markov-Switching Vector Autoregressions(MSVAR) process population demography analysis. The results indicated that the Yangtze River may not act as an effective barrier to gene flow in the tufted deer. Finally, we surmised that the population demography of the tufted deer was likely affected by Pleistocene climate fluctuations and ancient human activities. creator: Zhonglou Sun creator: Tao Pan creator: Hui Wang creator: Mujia Pang creator: Baowei Zhang uri: https://doi.org/10.7717/peerj.2654 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Sun et al. title: Iron overload induced death of osteoblasts in vitro: involvement of the mitochondrial apoptotic pathway link: https://peerj.com/articles/2611 last-modified: 2016-11-08 description: BackgroundIron overload is recognized as a new pathogenfor osteoporosis. Various studies demonstrated that iron overload could induce apoptosis in osteoblasts and osteoporosis in vivo. However, the exact molecular mechanisms involved in the iron overload-mediated induction of apoptosis in osteoblasts has not been explored.PurposeIn this study, we attempted to determine whether the mitochondrial apoptotic pathway is involved in iron-induced osteoblastic cell death and to investigate the beneficial effect of N-acetyl-cysteine (NAC) in iron-induced cytotoxicity.MethodsThe MC3T3-E1 osteoblastic cell line was treated with various concentrations of ferric ion in the absence or presence of NAC, and intracellular iron, cell viability, reactive oxygen species, functionand morphology changes of mitochondria and mitochondrial apoptosis related key indicators were detected by commercial kits. In addition, to further explain potential mechanisms underlying iron overload-related osteoporosis, we also assessed cell viability, apoptosis, and osteogenic differentiation potential in bone marrow-derived mesenchymal stemcells(MSCs) by commercial kits.ResultsFerric ion demonstrated concentration-dependent cytotoxic effects on osteoblasts. After incubation with iron, an elevation of intracelluar labile iron levels and a concomitant over-generation of reactive oxygen species (ROS) were detected by flow cytometry in osteoblasts. Nox4 (NADPH oxidase 4), an important ROS producer, was also evaluated by western blot. Apoptosis, which was evaluated by Annexin V/propidium iodide staining, Hoechst 33258 staining, and the activation of caspase-3, was detected after exposure to iron. Iron contributed to the permeabilizatio of mitochondria, leading to the release of cytochrome C (cyto C), which, in turn, induced mitochondrial apoptosis in osteoblasts via activation of Caspase-3, up-regulation of Bax, and down-regulation of Bcl-2. NAC could reverse iron-mediated mitochondrial dysfunction and blocked the apoptotic events through inhibit the generation of ROS. In addition, iron could significantly promote apoptosis and suppress osteogenic differentiation and mineralization in bone marrow-derived MSCs.ConclusionsThese findings firstly demonstrate that the mitochondrial apoptotic pathway involved in iron-induced osteoblast apoptosis. NAC could relieved the oxidative stress and shielded osteoblasts from apoptosis casused by iron-overload. We also reveal that iron overload in bone marrow-derived MSCs results in increased apoptosis and the impairment of osteogenesis and mineralization. creator: Qing Tian creator: Shilei Wu creator: Zhipeng Dai creator: Jingjing Yang creator: Jin Zheng creator: Qixin Zheng creator: Yong Liu uri: https://doi.org/10.7717/peerj.2611 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Tian et al. title: Herbarium-based studies on taxonomy, biogeography and ecology of Psilochilus (Orchidaceae) link: https://peerj.com/articles/2600 last-modified: 2016-11-08 description: BackgroundPsilochilus is a poorly studied orchid genus distributed from southern Mexico to south-eastern Brazil. A taxonomic revision of this Neotropical endemic based on morphological data is presented.Material and MethodsOver 170 dried herbarium specimens and flowers preserved in liquid of Psilochilus were analyzed. Morphological variation among examined taxa was described based on multivariate analysis. To evaluate the similarity between niches occupied by various Psilochilus species ecological niche modeling (ENM) was applied. Species richness and the distribution patterns of Psilochilus representatives were analyzed based on squares of 5° latitude and longitude while similarities among floras between biogeographical units were measured using the Bray-Curtis index for presence/absence data.Results and DiscussionA new species of the P. physurifolius-complex is described based on Central American material. Psilochilus crenatifolius is reduced to the rank of variety as P. macrophyllus var. crenatifolius. A key to 18 accepted Psilochilus species is provided. The illustrations of perianth segments of all recognized taxa are presented. The climatic niches preferred by the particular Psilochilus representatives are well separated based on ecological niche modeling analysis. Their distribution is limited mainly by the isothermality and temperature seasonality. The highest Psilochilus species richness is observed in the North Andean, Panamanian, Brazilian Planalto and Central American biogeographical provinces. A high level of endemism is observed in all those regions as well as Yungas biogeographical province. Most Psilochilus species occur in areas above 800 m of elevation. The populations were most often reported from the tropical rain forest and tropical moist deciduous forest. creator: Marta Kolanowska creator: Aleksandra M. Naczk creator: Radomir Jaskuła uri: https://doi.org/10.7717/peerj.2600 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Kolanowska et al. title: Supply-side barriers to maternal health care utilization at health sub-centers in India link: https://peerj.com/articles/2675 last-modified: 2016-11-03 description: IntroductionThere exist several barriers to maternal health service utilization in developing countries. Most of the previous studies conducted in India have focused on demand-side barriers, while only a few have touched upon supply-side barriers. None of the previous studies in India have investigated the factors that affect maternal health care utilization at health sub-centers (HSCs) in India, despite the fact that these institutions, which are the geographically closest available public health care facilities in rural areas, play a significant role in providing affordable maternal health care. Therefore, this study aims to examine the supply-side determinants of maternal service utilization at HSCs in rural India.Data and MethodsThis study uses health facility data from the nationally representative District-Level Household Survey, which was administered in 2007–2008 to examine the effect of supply-side variables on the utilization of maternal health care services across HSCs in rural India. Since the dependent variables (the number of antenatal registrations, in-facility deliveries, and postnatal care services) are count variables and exhibit considerable variability, the data were analyzed using negative binomial regression instead of Poisson regression.ResultsThe results show that those HSCs run by a contractual auxiliary nurse midwife (ANM) are likely to offer a lower volume of services when compared to those run by a permanent ANM. The availability of obstetric drugs, weighing scales, and blood pressure equipment is associated with the increased utilization of antenatal and postnatal services. The unavailability of a labor/examination table and bed screen is associated with a reduction in the number of deliveries and postnatal services. The utilization of services is expected to increase if essential facilities, such as water, telephones, toilets, and electricity, are available at the HSCs. Monitoring of ANM’s work by Village Health and Sanitation Committee (VHSC) and providing in-service training to ANM appear to have positive impacts on service utilization. The distance of ANM’s actual residence from the sub-center village where she works is negatively associated with the utilization of delivery and postnatal services. These findings are robust to the inclusion of several demand-side factors.ConclusionTo improve maternal health care utilization at HSCs, the government shouldensure the availability of basic infrastructure, drugs, and equipment at all locations. Monitoring of the ANMs’ work by VHSCs could play an important role in improving health care utilization at the HSCs; therefore, it is important to establish VHSCs in each sub-center village. The relatively low utilization of maternity services in those HSCs that are run solely by contractual ANMs requires further investigation. creator: Aditya Singh uri: https://doi.org/10.7717/peerj.2675 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Singh title: Hydrogen peroxide induced loss of heterozygosity correlates with replicative lifespan and mitotic asymmetry in Saccharomyces cerevisiae link: https://peerj.com/articles/2671 last-modified: 2016-11-03 description: Cellular aging in Saccharomyces cerevisiae can lead to genomic instability and impaired mitotic asymmetry. To investigate the role of oxidative stress in cellular aging, we examined the effect of exogenous hydrogen peroxide on genomic instability and mitotic asymmetry in a collection of yeast strains with diverse backgrounds. We treated yeast cells with hydrogen peroxide and monitored the changes of viability and the frequencies of loss of heterozygosity (LOH) in response to hydrogen peroxide doses. The mid-transition points of viability and LOH were quantified using sigmoid mathematical functions. We found that the increase of hydrogen peroxide dependent genomic instability often occurs before a drop in viability. We previously observed that elevation of genomic instability generally lags behind the drop in viability during chronological aging. Hence, onset of genomic instability induced by exogenous hydrogen peroxide treatment is opposite to that induced by endogenous oxidative stress during chronological aging, with regards to the midpoint of viability. This contrast argues that the effect of endogenous oxidative stress on genome integrity is well suppressed up to the dying-off phase during chronological aging. We found that the leadoff of exogenous hydrogen peroxide induced genomic instability to viability significantly correlated with replicative lifespan (RLS), indicating that yeast cells’ ability to counter oxidative stress contributes to their replicative longevity. Surprisingly, this leadoff is positively correlated with an inverse measure of endogenous mitotic asymmetry, indicating a trade-off between mitotic asymmetry and cell’s ability to fend off hydrogen peroxide induced oxidative stress. Overall, our results demonstrate strong associations of oxidative stress to genomic instability and mitotic asymmetry at the population level of budding yeast. creator: Emine Güven creator: Lindsay A. Parnell creator: Erin D. Jackson creator: Meighan C. Parker creator: Nilin Gupta creator: Jenny Rodrigues creator: Hong Qin uri: https://doi.org/10.7717/peerj.2671 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Guven et al. title: Antibiotic misuse among children with diarrhea in China: results from a national survey link: https://peerj.com/articles/2668 last-modified: 2016-11-03 description: BackgroundAntibiotic resistance is one of the world’s biggest public health issues, and the situation in China is particularly grave. The objective of this study is to investigate the antibiotics usage pattern among Chinese children and provide further insight in developing strategies for promoting public health education.MethodsThis is a cross-sectional study, in the study, participants are from 53,665 guardians of children aged 0–6 years, who were recruited with multistage stratified random cluster sampling in 2013/2014 from 46 community health centers in 14 provinces across China Mainland. Children’s guardians completed surveys on their previous experience on using antibiotics in treating diarrhea of their children without a prescription from any pediatrician. Odds ratios (ORs) and 95% confidential intervals (CIs) for the association between antibiotic use and its predictors were estimated using multilevel logistic regression models, with antibiotic rational use group as a reference group.ResultsThe prevalence of antibiotic misuse among children with diarrhea in the eastern, middle and western areas of China and associations between antibiotic misuse and its predictors were studied. The average rate of antibiotic misuse is 35.12%. Multilevel logistic regression revealed that living in urban areas (OR = 0.79 (0.76, 0.83)), female children (OR = 0.92 (0.88, 0.96)), guardians having higher education (OR = 0.60 (0.55, 0.66)), being raised by parents (OR = 0.90 (0.85, 0.94)), guardians having basic health knowledge (OR = 0.82 (0.79, 0.86)) are protective factors and children’s age (1–3 years OR = 1.62 (1.54, 1.71)); 4–6 years OR = 1.90 (1.77, 2.03)) is a risk factor of antibiotic misuse among children aged 0–6 years with diarrhea in China.ConclusionsOur findings confirmed that there has been a high rate of antibiotic misuse without a prescription in children with diarrhea in China, which requires considerable attention. Suitable regulations and interventions are needed to solve this problem. creator: Ruili Li creator: Feng Xiao creator: Xiaoguo Zheng creator: Huimin Yang creator: Lihong Wang creator: Delu Yin creator: Tao Yin creator: Qianqian Xin creator: Bowen Chen uri: https://doi.org/10.7717/peerj.2668 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Li et al. title: Hypergravity hinders axonal development of motor neurons in Caenorhabditis elegans link: https://peerj.com/articles/2666 last-modified: 2016-11-03 description: As space flight becomes more accessible in the future, humans will be exposed to gravity conditions other than our 1G environment on Earth. Our bodies and physiology, however, are adapted for life at 1G gravity. Altering gravity can have profound effects on the body, particularly the development of muscles, but the reasons and biology behind gravity’s effect are not fully known. We asked whether increasing gravity had effects on the development of motor neurons that innervate and control muscle, a relatively unexplored area of gravity biology. Using the nematode model organism Caenorhabditis elegans, we examined changes in response to hypergravity in the development of the 19 GABAergic DD/VD motor neurons that innervate body muscle. We found that a high gravity force above 10G significantly increases the number of animals with defects in the development of axonal projections from the DD/VD neurons. We showed that a critical period of hypergravity exposure during the embryonic/early larval stage was sufficient to induce defects. While characterizing the nature of the axonal defects, we found that in normal 1G gravity conditions, DD/VD axonal defects occasionally occurred, with the majority of defects occurring on the dorsal side of the animal and in the mid-body region, and a significantly higher rate of error in the 13 VD axons than the 6 DD axons. Hypergravity exposure increased the rate of DD/VD axonal defects, but did not change the distribution or the characteristics of the defects. Our study demonstrates that altering gravity can impact motor neuron development. creator: Saraswathi Subbammal Kalichamy creator: Tong Young Lee creator: Kyoung-hye Yoon creator: Jin Il Lee uri: https://doi.org/10.7717/peerj.2666 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Kalichamy et al. title: Host plant affects morphometric variation of Diaphorina citri (Hemiptera: Liviidae) link: https://peerj.com/articles/2663 last-modified: 2016-11-03 description: The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama, is one of the most serious citrus pests worldwide due to its role as vector of huanglongbing or citrus greening disease. While some optimal plant species for ACP oviposition and development have been identified, little is known of the influence of host plants on ACP size and shape. Our goal was to determine how size and shape of ACP wing and body size varies when development occurs on different host plants in a controlled rearing environment. ACP were reared on six different rutaceous species; Bergera koenigii, Citrus aurantifolia, Citrus macrophylla, Citrus maxima, Citrus taiwanica and Murraya paniculata. Adults were examined for morphometric variation using traditional and geometric analysis based on 12 traits or landmarks. ACP reared on C. taiwanica were consistently smaller than those reared on the other plant species. Wing aspect ratio also differed between C. maxima and C. taiwanica. Significant differences in shape were detected with those reared on M. paniculata having narrower wings than those reared on C. macrophylla. This study provides evidence of wing size and shape differences of ACP based on host plant species which potentially may impact dispersal. Further study is needed to determine if behavioral and physiological differences are associated with the observed phenotypic differences. creator: Thomson M. Paris creator: Sandra A. Allan creator: David G. Hall creator: Matthew G. Hentz creator: Gabriella Hetesy creator: Philip A. Stansly uri: https://doi.org/10.7717/peerj.2663 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Paris et al. title: Salivary microbiomes of indigenous Tsimane mothers and infants are distinct despite frequent premastication link: https://peerj.com/articles/2660 last-modified: 2016-11-03 description: BackgroundPremastication, the transfer of pre-chewed food, is a common infant and young child feeding practice among the Tsimane, forager-horticulturalists living in the Bolivian Amazon. Research conducted primarily with Western populations has shown that infants harbor distinct oral microbiota from their mothers. Premastication, which is less common in these populations, may influence the colonization and maturation of infant oral microbiota, including via transmission of oral pathogens. We collected premasticated food and saliva samples from Tsimane mothers and infants (9–24 months of age) to test for evidence of bacterial transmission in premasticated foods and overlap in maternal and infant salivary microbiota. We extracted bacterial DNA from two premasticated food samples and 12 matched salivary samples from maternal-infant pairs. DNA sequencing was performed with MiSeq (Illumina). We evaluated maternal and infant microbial composition in terms of relative abundance of specific taxa, alpha and beta diversity, and dissimilarity distances.ResultsThe bacteria in saliva and premasticated food were mapped to 19 phyla and 400 genera and were dominated by Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes. The oral microbial communities of Tsimane mothers and infants who frequently share premasticated food were well-separated in a non-metric multi-dimensional scaling ordination (NMDS) plot. Infant microbiotas clustered together, with weighted Unifrac distances significantly differing between mothers and infants. Infant saliva contained more Firmicutes (p < 0.01) and fewer Proteobacteria (p < 0.05) than did maternal saliva. Many genera previously associated with dental and periodontal infections, e.g. Neisseria, Gemella, Rothia, Actinomyces, Fusobacterium, and Leptotrichia, were more abundant in mothers than in infants.ConclusionsSalivary microbiota of Tsimane infants and young children up to two years of age do not appear closely related to those of their mothers, despite frequent premastication and preliminary evidence that maternal bacteria is transmitted to premasticated foods. Infant physiology and diet may constrain colonization by maternal bacteria, including several oral pathogens. creator: Cliff S. Han creator: Melanie Ann Martin creator: Armand E.K. Dichosa creator: Ashlynn R. Daughton creator: Seth Frietze creator: Hillard Kaplan creator: Michael D. Gurven creator: Joe Alcock uri: https://doi.org/10.7717/peerj.2660 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Han et al. title: Sevoflurane postconditioning attenuates cardiomyocyte hypoxia/reoxygenation injury via restoring mitochondrial morphology link: https://peerj.com/articles/2659 last-modified: 2016-11-03 description: BackgroundAnesthetic postconditioning is a cellular protective approach whereby exposure to a volatile anesthetic renders a tissue more resistant to subsequent ischemic/reperfusion event. Sevoflurane postconditioning (SPostC) has been shown to exert cardioprotection against ischemia/reperfusion injury, but the underlying mechanism is unclear. We hypothesized that SPostC protects cardiomyocytes against hypoxia/reoxygenation (H/R) injury by maintaining/restoring mitochondrial morphological integrity, a critical determinant of cell fate.MethodsPrimary cultures of neonatal rat cardiomyocytes (NCMs) were subjected to H/R injury (3 h of hypoxia followed by 3 h reoxygenation). Intervention with SPostC (2.4% sevoflurane) was administered for 15 min upon the onset of reoxygenation. Cell viability, Lactate dehydrogenase (LDH) level, cell death, mitochondrial morphology, mitochondrial membrane potential and mitochondrial permeability transition pore (mPTP) opening were assessed after intervention. Mitochondrial fusion and fission regulating proteins (Drp1, Fis1, Mfn1, Mfn2 and Opa1) were assessed by immunofluorescence staining and western blotting was performed to determine the level of protein expression.ResultsCardiomyocyte H/R injury resulted in significant increases in LDH release and cell death that were concomitant with reduced cell viability and reduced mitochondrial interconnectivity (mean area/perimeter ratio) and mitochondrial elongation, and with reduced mitochondrial membrane potential and increased mPTP opening. All the above changes were significantly attenuated by SPostC. Furthermore, H/R resulted in significant reductions in mitochondrial fusion proteins Mfn1, Mfn2 and Opa1 and significant enhancement of fission proteins Drp1 and Fis1. SPostC significantly enhanced Mfn2 and Opa1 and reduced Drp1, without significant impact on Mfn1 and Fis1.ConclusionsSevoflurane postconditioning attenuates cardiomyocytes hypoxia/reoxygenation injury (HRI) by restoring mitochondrial fusion/fission balance and morphology. creator: Jin Yu creator: Jianjiang Wu creator: Peng Xie creator: Yiliyaer Maimaitili creator: Jiang Wang creator: Zhengyuan Xia creator: Feng Gao creator: Xing Zhang creator: Hong Zheng uri: https://doi.org/10.7717/peerj.2659 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Yu et al. title: Foliar fungal communities strongly differ between habitat patches in a landscape mosaic link: https://peerj.com/articles/2656 last-modified: 2016-11-03 description: BackgroundDispersal events between habitat patches in a landscape mosaic can structure ecological communities and influence the functioning of agrosystems. Here we investigated whether short-distance dispersal events between vineyard and forest patches shape foliar fungal communities. We hypothesized that these communities homogenize between habitats over the course of the growing season, particularly along habitat edges, because of aerial dispersal of spores.MethodsWe monitored the richness and composition of foliar and airborne fungal communities over the season, along transects perpendicular to edges between vineyard and forest patches, using Illumina sequencing of the Internal Transcribed Spacer 2 (ITS2) region.ResultsIn contrast to our expectation, foliar fungal communities in vineyards and forest patches increasingly differentiate over the growing season, even along habitat edges. Moreover, the richness of foliar fungal communities in grapevine drastically decreased over the growing season, in contrast to that of forest trees. The composition of airborne communities did not differ between habitats. The composition of oak foliar fungal communities change between forest edge and centre.DiscussionThese results suggest that dispersal events between habitat patches are not major drivers of foliar fungal communities at the landscape scale. Selective pressures exerted in each habitat by the host plant, the microclimate and the agricultural practices play a greater role, and might account for the differentiation of foliar fugal communities between habitats. creator: Thomas Fort creator: Cécile Robin creator: Xavier Capdevielle creator: Laurent Delière creator: Corinne Vacher uri: https://doi.org/10.7717/peerj.2656 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Fort et al. title: Phytogeographic and genetic variation in Sorbus, a traditional antidiabetic medicine—adaptation in action in both a plant and a discipline link: https://peerj.com/articles/2645 last-modified: 2016-11-03 description: Mountain ash (Sorbus decora and S. americana) is used by the Cree Nation of the James Bay region of Quebec (Eeyou Istchee) as traditional medicine. Its potential as an antidiabetic medicine is thought to vary across its geographical range, yet little is known about the factors that affect its antioxidant capacity. Here, we examined metabolite gene expression in relation to antioxidant activity, linking phytochemistry and medicinal potential. Samples of leaf and bark from S. decora and S. americana were collected from 20 populations at four different latitudes. Two genes known to produce antidiabetic substances, flavonol synthase and squalene synthase, were analyzed using quantitative real time PCR. Gene expression was significantly higher for flavonol synthase compared to squalene synthase and increased in the most Northern latitude. Corresponding differences observed in the antioxidant capacity of ethanolic extracts from the bark of Sorbus spp. confirm that plants at higher latitudes increase production of stress-induced secondary metabolites and support Aboriginal perceptions of their higher medicinal potential. Modern genetic techniques such as quantitative real time PCR offer unprecedented resolution to substantiate and scrutinise Aboriginal medicinal plant perception. Furthermore, it offers valuable insights into how environmental stress can trigger an adaptive response resulting in the accumulation of secondary metabolites with human medicinal properties. creator: Anna Bailie creator: Sebastien Renaut creator: Eliane Ubalijoro creator: José A. Guerrero-Analco creator: Ammar Saleem creator: Pierre Haddad creator: John T. Arnason creator: Timothy Johns creator: Alain Cuerrier uri: https://doi.org/10.7717/peerj.2645 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Bailie et al. title: Mathematical models are a powerful method to understand and control the spread of Huanglongbing link: https://peerj.com/articles/2642 last-modified: 2016-11-03 description: Huanglongbing (HLB), or citrus greening, is a global citrus disease occurring in almost all citrus growing regions. It causes substantial economic burdens to individual growers, citrus industries and governments. Successful management strategies to reduce disease burden are desperately needed but with so many possible interventions and combinations thereof it is difficult to know which are worthwhile or cost-effective. We review how mathematical models have yielded useful insights into controlling disease spread for other vector-borne plant diseases, and the small number of mathematical models of HLB. We adapt a malaria model to HLB, by including temperature-dependent psyllid traits, “flushing” of trees, and economic costs, to show how models can be used to highlight the parameters that require more data collection or that should be targeted for intervention. We analyze the most common intervention strategy, insecticide spraying, to determine the most cost-effective spraying strategy. We find that fecundity and feeding rate of the vector require more experimental data collection, for wider temperatures ranges. Also, the best strategy for insecticide intervention is to spray for more days rather than pay extra for a more efficient spray. We conclude that mathematical models are able to provide useful recommendations for managing HLB spread. creator: Rachel A. Taylor creator: Erin A. Mordecai creator: Christopher A. Gilligan creator: Jason R. Rohr creator: Leah R. Johnson uri: https://doi.org/10.7717/peerj.2642 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Taylor et al. title: Germination pretreatments to break hard-seed dormancy in Astragalus cicer L. (Fabaceae) link: https://peerj.com/articles/2621 last-modified: 2016-11-03 description: Conservationists often propagate rare species to improve their long-term population viability. However, seed dormancy can make propagation efforts challenging by substantially lowering seed germination. Here I statistically compare several pretreatment options for seeds of Astragalus cicer L.: unscarified controls and scarification via physical damage, hot water, acid, and hydrogen peroxide. Although only 30% of unscarified seeds germinated, just physical scarification significantly improved germination, whereas one treatment, hot water, resulted in no germination at all. I recommend that rare species of Astragalus, as well as other hard-seeded legumes, be pretreated using physical scarification. Other methods may require considerable optimization, wasting precious time and seeds. creator: Joseph M. Statwick uri: https://doi.org/10.7717/peerj.2621 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Statwick title: Phylogeography of Arenaria balearica L. (Caryophyllaceae): evolutionary history of a disjunct endemic from the Western Mediterranean continental islands link: https://peerj.com/articles/2618 last-modified: 2016-11-03 description: Although it has been traditionally accepted that Arenaria balearica (Caryophyllaceae) could be a relict Tertiary plant species, this has never been experimentally tested. Nor have the palaeohistorical reasons underlying the highly fragmented distribution of the species in the Western Mediterranean region been investigated. We have analysed AFLP data (213) and plastid DNA sequences (226) from a total of 250 plants from 29 populations sampled throughout the entire distribution range of the species in Majorca, Corsica, Sardinia, and the Tuscan Archipelago. The AFLP data analyses indicate very low geographic structure and population differentiation. Based on plastid DNA data, six alternative phylogeographic hypotheses were tested using Approximate Bayesian Computation (ABC). These analyses revealed ancient area fragmentation as the most probable scenario, which is in accordance with the star-like topology of the parsimony network that suggests a pattern of long term survival and subsequent in situ differentiation. Overall low levels of genetic diversity and plastid DNA variation were found, reflecting evolutionary stasis of a species preserved in locally long-term stable habitats. creator: Javier Bobo-Pinilla creator: Sara B. Barrios de León creator: Jaume Seguí Colomar creator: Giuseppe Fenu creator: Gianluigi Bacchetta creator: Julio Peñas de Giles creator: María Montserrat Martínez-Ortega uri: https://doi.org/10.7717/peerj.2618 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Bobo-Pinilla et al. title: Analyzing pathogen suppressiveness in bioassays with natural soils using integrative maximum likelihood methods in R link: https://peerj.com/articles/2615 last-modified: 2016-11-03 description: The potential of soils to naturally suppress inherent plant pathogens is an important ecosystem function. Usually, pathogen infection assays are used for estimating the suppressive potential of soils. In natural soils, however, co-occurring pathogens might simultaneously infect plants complicating the estimation of a focal pathogen’s infection rate (initial slope of the infection-curve) as a measure of soil suppressiveness. Here, we present a method in R correcting for these unwanted effects by developing a two pathogen mono-molecular infection model. We fit the two pathogen mono-molecular infection model to data by using an integrative approach combining a numerical simulation of the model with an iterative maximum likelihood fit. We show that in presence of co-occurring pathogens using uncorrected data leads to a critical under- or overestimation of soil suppressiveness measures. In contrast, our new approach enables to precisely estimate soil suppressiveness measures such as plant infection rate and plant resistance time. Our method allows a correction of measured infection parameters that is necessary in case different pathogens are present. Moreover, our model can be (1) adapted to use other models such as the logistic or the Gompertz model; and (2) it could be extended by a facilitation parameter if infections in plants increase the susceptibility to new infections. We propose our method to be particularly useful for exploring soil suppressiveness of natural soils from different sites (e.g., in biodiversity experiments). creator: Björn C. Rall creator: Ellen Latz uri: https://doi.org/10.7717/peerj.2615 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Rall and Latz title: Alterations to the middle cerebral artery of the hypertensive-arthritic rat model potentiates intracerebral hemorrhage link: https://peerj.com/articles/2608 last-modified: 2016-11-03 description: AimsWe have recently created an age-dependent hypertensive-mono-arthritic animal model from the stroke-resistant spontaneously hypertensive rat to model populations with autoimmune disease who are hypertensive and are prone to stroke. The model exhibits signs of hemorrhagic stroke (HS) subsequent to chronic inflammation and hypertension. HS is also associated with the inability of middle cerebral arteries to undergo pressure dependent constriction (PDC). We investigated alterations in the cerebrovasculature of our hypertensive mono-arthritic animals that develop stroke.Main MethodsAnimals were fed either a high salt diet (HSD) (4% NaCl) or Purina chow (0.58% NaCl) from weaning. Complete Freund’s Adjuvant (CFA) was injected into the left hind paw at 21–28 weeks; controls received saline and histological and functional studies were performed.ResultsBrain damage was more prominent with the high salt, with inflammation exacerbating the damage. High salt alone significantly decreased middle cerebral artery’s (MCA’s) ability to undergo PDC. Inflammation significantly decreased the ability of cerebrovasculature to respond to pressure step in the regular salt diet. The responses to vasoactive peptides were also significantly attenuated in both inflamed groups regardless of diet.ConclusionInduction of chronic systemic inflammation increases brain damage, and affect the MCA’s vasogenic function, decreasing its ability to respond to intraluminal pressure. HSD further exacerbates organ damage associated with chronic inflammation, further compromising cerebrovascular function, and likely increasing the incidence of intracerebral hemorrhage and injury. creator: Amy Randell creator: Killol Chokshi creator: Brittany Kane creator: Hilary Chang creator: Safaa Naiel creator: Jeffrey G. Dickhout creator: Noriko Daneshtalab uri: https://doi.org/10.7717/peerj.2608 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Randell et al. title: Environmental, land cover and land use constraints on the distributional patterns of anurans: Leptodacylus species (Anura, Leptodactylidae) from Dry Chaco link: https://peerj.com/articles/2605 last-modified: 2016-11-03 description: Subtropical dry forests are among the most vulnerable biomes to land transformation at a global scale. Among them, the Dry Chaco suffers an accelerated change due to agriculture expansion and intensification. The Dry Chaco ecoregion is characterized by high levels of endemisms and species diversity, which are the result of a variety of climates and reliefs, allowing a wide variety of environments. The amphibian group exhibits a high richness in the Dry Chaco, which has been barely studied in relation to land cover changes. We used ecological niche models (ENMs) to assess the potential geographic distribution of 10 Leptodactylus species (Anura, Leptodactylidae), which are mainly distributed within the Dry Chaco. We characterized these distributions environmentally, analyzed their overlap with land cover classes, and assessed their diversity of ecoregions. Also, we evaluated how these species potential distribution is affected by the transformation of land, and quantified the proportional area of the potential distribution in protected areas. We found that temperature seasonality is the main constraint to the occurrence of the species studied, whose main habitats are savannas, grasslands and croplands. The main threats to these species are the effects of climate change over spatial patterns of seasonality, which could affect their breeding and reproduction mode; the loss of their natural habitat; the exposure to contaminants used by intensive agriculture and their underrepresentation in protected areas. creator: Regina Gabriela Medina creator: Maria Laura Ponssa creator: Ezequiel Aráoz uri: https://doi.org/10.7717/peerj.2605 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Medina et al. title: Regional variation in fire weather controls the reported occurrence of Scottish wildfires link: https://peerj.com/articles/2649 last-modified: 2016-11-02 description: Fire is widely used as a traditional habitat management tool in Scotland, but wildfires pose a significant and growing threat. The financial costs of fighting wildfires are significant and severe wildfires can have substantial environmental impacts. Due to the intermittent occurrence of severe fire seasons, Scotland, and the UK as a whole, remain somewhat unprepared. Scotland currently lacks any form of Fire Danger Rating system that could inform managers and the Fire and Rescue Services (FRS) of periods when there is a risk of increased of fire activity. We aimed evaluate the potential to use outputs from the Canadian Fire Weather Index system (FWI system) to forecast periods of increased fire risk and the potential for ignitions to turn into large wildfires. We collated four and a half years of wildfire data from the Scottish FRS and examined patterns in wildfire occurrence within different regions, seasons, between urban and rural locations and according to FWI system outputs. We used a variety of techniques, including Mahalanobis distances, percentile analysis and Thiel-Sen regression, to scope the best performing FWI system codes and indices. Logistic regression showed significant differences in fire activity between regions, seasons and between urban and rural locations. The Fine Fuel Moisture Code and the Initial Spread Index did a tolerable job of modelling the probability of fire occurrence but further research on fuel moisture dynamics may provide substantial improvements. Overall our results suggest it would be prudent to ready resources and avoid managed burning when FFMC > 75 and/or ISI > 2. creator: G. Matt Davies creator: Colin J. Legg uri: https://doi.org/10.7717/peerj.2649 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Davies and Legg title: A public data set of human balance evaluations link: https://peerj.com/articles/2648 last-modified: 2016-11-02 description: The goal of this study was to create a public data set with results of qualitative and quantitative evaluations related to human balance. Subject’s balance was evaluated by posturography using a force platform and by the Mini Balance Evaluation Systems Tests. In the posturography test, we evaluated subjects standing still for 60 s in four different conditions where vision and the standing surface were manipulated: on a rigid surface with eyes open; on a rigid surface with eyes closed; on an unstable surface with eyes open; on an unstable surface with eyes closed. Each condition was performed three times and the order of the conditions was randomized. In addition, the following tests were employed in order to better characterize each subject: Short Falls Efficacy Scale International; International Physical Activity Questionnaire Short Version; and Trail Making Test. The subjects were also interviewed to collect information about their socio-cultural, demographic, and health characteristics. The data set comprises signals from the force platform (raw data for the force, moments of forces, and centers of pressure) of 163 subjects plus one file with information about the subjects and balance conditions and the results of the other evaluations. All the data is available at PhysioNet and at Figshare. creator: Damiana A. Santos creator: Marcos Duarte uri: https://doi.org/10.7717/peerj.2648 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Santos and Duarte title: Seasonal regulation of herbivory and nutrient effects on macroalgal recruitment and succession in a Florida coral reef link: https://peerj.com/articles/2643 last-modified: 2016-11-02 description: Herbivory and nutrient enrichment are drivers of benthic dynamics of coral reef macroalgae; however, their impact may vary seasonally. In this study we evaluated the effects of herbivore pressure, nutrient availability and potential propagule supply on seasonal recruitment and succession of macroalgal communities on a Florida coral reef. Recruitment tiles, replaced every three months, and succession tiles, kept in the field for nine months, were established in an ongoing factorial nutrient enrichment-herbivore exclusion experiment. The ongoing experiment had already created very different algal communities across the different herbivory and nutrient treatments. We tracked algal recruitment, species richness, and species abundance through time. Our results show seasonal variation in the effect of herbivory and nutrient availability on recruitment of coral reef macroalgae. In the spring, when there was higher macroalgal species richness and abundance of recruits, herbivory appeared to have more control on macroalgal community structure than did nutrients. In contrast, there was no effect of either herbivory or nutrient enrichment on macroalgal communities on recruitment tiles in cooler seasons. The abundance of recruits on tiles was positively correlated with the abundance of algal in the ongoing, established experiment, suggesting that propagule abundance is likely a strong influence on algal recruitment and early succession. Results of the present study suggest that abundant herbivorous fishes control recruitment and succession of macroalgae, particularly in the warm season when macroalgal growth is higher. However, herbivory appears less impactful on algal recruitment and community dynamics in cooler seasons. Ultimately, our data suggest that the timing of coral mortality (e.g., summer vs. winter mortality) and freeing of benthic space may strongly influence the dynamics of algae that colonize open space. creator: Alain Duran creator: Ligia Collado-Vides creator: Deron E. Burkepile uri: https://doi.org/10.7717/peerj.2643 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Duran et al. title: Region-of-interest analyses of one-dimensional biomechanical trajectories: bridging 0D and 1D theory, augmenting statistical power link: https://peerj.com/articles/2652 last-modified: 2016-11-02 description: One-dimensional (1D) kinematic, force, and EMG trajectories are often analyzed using zero-dimensional (0D) metrics like local extrema. Recently whole-trajectory 1D methods have emerged in the literature as alternatives. Since 0D and 1D methods can yield qualitatively different results, the two approaches may appear to be theoretically distinct. The purposes of this paper were (a) to clarify that 0D and 1D approaches are actually just special cases of a more general region-of-interest (ROI) analysis framework, and (b) to demonstrate how ROIs can augment statistical power. We first simulated millions of smooth, random 1D datasets to validate theoretical predictions of the 0D, 1D and ROI approaches and to emphasize how ROIs provide a continuous bridge between 0D and 1D results. We then analyzed a variety of public datasets to demonstrate potential effects of ROIs on biomechanical conclusions. Results showed, first, that a priori ROI particulars can qualitatively affect the biomechanical conclusions that emerge from analyses and, second, that ROIs derived from exploratory/pilot analyses can detect smaller biomechanical effects than are detectable using full 1D methods. We recommend regarding ROIs, like data filtering particulars and Type I error rate, as parameters which can affect hypothesis testing results, and thus as sensitivity analysis tools to ensure arbitrary decisions do not influence scientific interpretations. Last, we describe open-source Python and MATLAB implementations of 1D ROI analysis for arbitrary experimental designs ranging from one-sample t tests to MANOVA. creator: Todd C. Pataky creator: Mark A. Robinson creator: Jos Vanrenterghem uri: https://doi.org/10.7717/peerj.2652 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Pataky et al. title: The Ecology of Stress: linking life-history traits with physiological control mechanisms in free-living guanacos link: https://peerj.com/articles/2640 last-modified: 2016-11-02 description: BackgroundProviding the context for the evolution of life-history traits, habitat features constrain successful ecological and physiological strategies. In vertebrates, a key response to life’s challenges is the activation of the Stress (HPA) and Gonadal (HPG) axes. Much of the interest in stress ecology is motivated by the desire to understand the physiological mechanisms in which the environment affects fitness. As reported in the literature, several intrinsic and extrinsic factors affect variability in hormone levels. In both social and non-social animals, the frequency and type of interaction with conspecifics, as well as the status in social species, can affect HPA axis activity, resulting in changes in the reproductive success of animals. We predicted that a social environment can affect both guanaco axes by increasing the secretion of testosterone (T) and Glucocorticoid (GCs) in response to individual social interactions and the energetic demands of breeding. Assuming that prolonged elevated levels of GCs over time can be harmful to individuals, it is predicted that the HPA axis suppresses the HPG axis and causes T levels to decrease, as GCs increase.MethodsAll of the data for individuals were collected by non-invasive methods (fecal samples) to address hormonal activities. This is a novel approach in physiological ecology because feces are easily obtained through non-invasive sampling in animal populations.ResultsAs expected, there was a marked adrenal (p-value = .3.4e−12) and gonadal (p-value = 0.002656) response due to seasonal variation in Lama guanicoe. No significant differences were found in fecal GCs metabolites between males/females*season for the entire study period (p-value = 0.2839). Despite the seasonal activity variation in the hormonal profiles, our results show a positive correlation (p-value = 1.952e−11, COR = 0.50) between the adrenal and gonadal system. The marked endocrine (r2 = 0.806) and gonad (r2 = 0.7231) response due to seasonal variation in male guanaco individuals highlights the individual’s energetic demands according to life-history strategies. This is a remarkable result because no inhibition was found between the axes as theory suggests. Finally, the dataset was used to build a reactive scope model for guanacos.DiscussionGuanacos cope with the trade-off between sociability and reproductive benefits and costs, by regulating their GCs and T levels on a seasonal basis, suggesting an adaptive role of both axes to different habitat pressures. The results presented here highlight the functional role of stress and gonad axes on a critical phase of a male mammal’s life—the mating period—when all of the resources are at the disposal of the male and must be used to maximize the chances for reproductive success. creator: Ramiro J.A. Ovejero Aguilar creator: Graciela A. Jahn creator: Mauricio Soto-Gamboa creator: Andrés J. Novaro creator: Pablo Carmanchahi uri: https://doi.org/10.7717/peerj.2640 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Ovejero Aguilar et al. title: Elevational pattern of bird species richness and its causes along a central Himalaya gradient, China link: https://peerj.com/articles/2636 last-modified: 2016-11-02 description: This study examines the relative importance of six variables: area, the mid-domain effect, temperature, precipitation, productivity, and habitat heterogeneity on elevational patterns of species richness for breeding birds along a central Himalaya gradient in the Gyirong Valley, the longest of five canyons in the Mount Qomolangma National Nature Reserve. We conducted field surveys in each of twelve elevational bands of 300 m between 1,800 and 5,400 m asl four times throughout the entire wet season. A total of 169 breeding bird species were recorded and most of the species (74%) were small-ranged. The species richness patterns of overall, large-ranged and small-ranged birds were all hump-shaped, but with peaks at different elevations. Large-ranged species and small-ranged species contributed equally to the overall richness pattern.Based on the bivariate and multiple regression analyses, area and precipitation were not crucial factors in determining the species richness along this gradient. The mid-domain effect played an important role in shaping the richness pattern of large-ranged species. Temperature was negatively correlated with overall and large-ranged species but positively correlated with small-ranged species. Productivity was a strong explanatory factor among all the bird groups, and habitat heterogeneity played an important role in shaping the elevational richness patterns of overall and small-ranged species. Our results highlight the need to conserve primary forest and intact habitat in this area. Furthermore, we need to increase conservation efforts in this montane biodiversity hotspot in light of increasing anthropogenic activities and land use pressure. creator: Xinyuan Pan creator: Zhifeng Ding creator: Yiming Hu creator: Jianchao Liang creator: Yongjie Wu creator: Xingfeng Si creator: Mingfang Guo creator: Huijian Hu creator: Kun Jin uri: https://doi.org/10.7717/peerj.2636 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Pan et al. title: Three-dimensional paleohistology of the scale and median fin spine of Lophosteus superbus (Pander 1856) link: https://peerj.com/articles/2521 last-modified: 2016-11-02 description: Lophosteus superbus is one of only a handful of probable stem-group osteichthyans known from the fossil record. First collected and described in the late 19th century from the upper Silurian Saaremaa Cliff locality in Estonia, it is known from a wealth of disarticulated scales, fin spines, and bone fragments. In this study we provide the first description of the morphology and paleohistology of a fin spine and scale from Lophosteus using virtual thin sections and 3D reconstructions that were segmented using phase-contrast synchrotron X-ray microtomography. These data reveal that both structures have fully or partially buried odontodes, which retain fine morphological details in older generations, including sharp nodes and serrated ridgelets. The vascular architecture of the fin spine tip, which is composed of several layers of longitudinally directed bone vascular canals, is much more complex compared to the bulbous horizontal canals within the scale, but they both have distinctive networks of ascending canals within each individual odontode. Other histological characteristics that can be observed from the data are cell spaces and Sharpey’s fibers that, when combined with the vascularization, could help to provide insights into the growth of the structure. The 3D data of the scales from Lophosteus superbus is similar to comparable data from other fossil osteichthyans, and the morphology of the reconstructed buried odontodes from this species is identical to scale material of Lophosteus ohesaarensis, casting doubt on the validity of that species. The 3D data presented in this paper is the first for fossil fin spines and so comparable data is not yet available. However, the overall morphology and histology seems to be similar to the structure of placoderm dermal plates. The 3D datasets presented here provide show that microtomography is a powerful tool for investigating the three-dimensional microstructure of fossils, which is difficult to study using traditional histological methods. These results also increase the utility of fin spines and scales suggest that these data are a potentially rich source of morphological data that could be used for studying questions relating to early vertebrate growth and evolution. creator: Anna Jerve creator: Qingming Qu creator: Sophie Sanchez creator: Henning Blom creator: Per Erik Ahlberg uri: https://doi.org/10.7717/peerj.2521 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Jerve et al. title: Comparison of the effectiveness of ISJ and SSR markers and detection of outlier loci in conservation genetics of Pulsatilla patens populations link: https://peerj.com/articles/2504 last-modified: 2016-11-02 description: BackgroundResearch into the protection of rare and endangered plant species involves genetic analyses to determine their genetic variation and genetic structure. Various categories of genetic markers are used for this purpose. Microsatellites, also known as simple sequence repeats (SSR), are the most popular category of markers in population genetics research. In most cases, microsatellites account for a large part of the noncoding DNA and exert a neutral effect on the genome. Neutrality is a desirable feature in evaluations of genetic differences between populations, but it does not support analyses of a population’s ability to adapt to a given environment or its evolutionary potential. Despite the numerous advantages of microsatellites, non-neutral markers may supply important information in conservation genetics research. They are used to evaluate adaptation to specific environmental conditions and a population’s adaptive potential. The aim of this study was to compare the level of genetic variation in Pulsatilla patens populations revealed by neutral SSR markers and putatively adaptive ISJ markers (intron-exon splice junction).MethodsThe experiment was conducted on 14 Polish populations of P. patens and three P. patens populations from the nearby region of Vitebsk in Belarus. A total of 345 individuals were examined. Analyses were performed with the use of eight SSR primers specific to P. patens and three ISJ primers.ResultsSSR markers revealed a higher level of genetic variation than ISJ markers (He = 0.609, He = 0.145, respectively). An analysis of molecular variance (AMOVA) revealed that, the overall genetic diversity between the analyzed populations defined by parameters FST and ΦPT for SSR (20%) and ΦPT for ISJ (21%) markers was similar. Analysis conducted in the Structure program divided analyzed populations into two groups (SSR loci) and three groups (ISJ markers). Mantel test revealed correlations between the geographic distance and genetic diversity of Polish populations of P. patens for ISJ markers, but not for SSR markers.ConclusionsThe results of the present study suggest that ISJ markers can complement the analyses based on SSRs. However, neutral and adaptive markers should not be alternatively applied. Neutral microsatellite markers cannot depict the full range of genetic variation in a population because they do not enable to analyze functional variation. Although ISJ markers are less polymorphic, they can contribute to the reliability of analyses based on SSRs. creator: Katarzyna Bilska creator: Monika Szczecińska uri: https://doi.org/10.7717/peerj.2504 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Bilska and Szczecińska title: Performance, usability and comparison of two versions of a new macular vision test: the handheld Radial Shape Discrimination test link: https://peerj.com/articles/2650 last-modified: 2016-11-01 description: BackgroundCentral vision, critical for everyday tasks such as reading and driving, is impacted by age-related changes in the eye and by diseases such as age-related macular degeneration. The detection of changes in macular function is therefore important. The Radial Shape Discrimination (RSD) test measures the threshold at which distortions in a radial frequency pattern can be detected and there is evidence that it is more sensitive to macular pathology than visual acuity (VA). It also provides a more quantitative measure of macular function than the commonly available Amsler grid. Recently, handheld versions of the test (hRSD) in which stimuli are presented on mobile devices (e.g., Apple iPod Touch, iPhone) have been developed. We investigated the characteristics of the hRSD test in healthy participants.MethodsData were collected using both three-alternative forced choice (3AFC) and 4AFC versions of the hRSD test, presented on an Apple iPod Touch. For the 3AFC version, data from a single test session were available for 186 (72 male; mean ± SD age 42 ± 17y; range 16–90y) healthy participants. Test-retest data were available for subgroups of participants (intra-session: N = 74; tests approximately 2 months apart: N = 30; tests 39 months apart: N = 15). The 3AFC and 4AFC versions were directly compared in 106 participants who also completed a usability questionnaire. Distance and near VA and Pelli Robson Contrast Sensitivity (CS) data were collected and undilated fundoscopy performed on the majority of participants.ResultsMean (±SD) 3AFC hRSD threshold was −0.77 ± 0.14 logMAR, and was statistically significantly correlated with age (Pearson r = 0.35; p < 0.001). The linear regression of hRSD threshold on age had a slope of +0.0026 compared to +0.0051 for near VA (which also correlated with age: r = 0.51; p < 0.001). There were no statistically significant differences in hRSD thresholds for any of the test-retest subgroups. We also observed no statistically significant difference between 3AFC (−0.82 ± 0.11 logMAR) and 4AFC (−0.80 ± 0.12 logMAR) hRSD thresholds (t = 1.85, p = 0.067) and participants reported excellent test usability with no strong preference expressed between the 3AFC and 4AFC versions of the test.DiscussionThe 3AFC hRSD thresholds we report are consistent with a number of previous studies, as is its greater stability in ageing compared to VA. We have also shown that in the absence of pathology, thresholds are stable over short and long timescales. The 4AFC thresholds we have reported provide a baseline for future investigations, and we have confirmed that 3AFC and 4AFC thresholds are similar, providing a basis of comparisons between studies using the different versions. As the hRSD test is easy to use and relatively inexpensive, clinical studies are now required to establish its ability to detect and monitor macular pathologies. creator: Jae Y. Ku creator: Ashli F. Milling creator: Noelia Pitrelli Vazquez creator: Paul C. Knox uri: https://doi.org/10.7717/peerj.2650 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Ku et al. title: The abundance of large, piscivorous Ferox Trout (Salmo trutta) in Loch Rannoch, Scotland link: https://peerj.com/articles/2646 last-modified: 2016-11-01 description: BackgroundFerox Trout are large, long-lived piscivorous Brown Trout (Salmo trutta). Due to their exceptionally large size, Ferox Trout are highly sought after by anglers while their life-history strategy, which includes delayed maturation, multiphasic growth and extended longevity, is of interest to ecological and evolutionary modelers. However, despite their recreational and theoretical importance, little is known about the typical abundance of Ferox Trout.MethodsTo rectify this situation a 16 year angling-based mark-recapture study was conducted on Loch Rannoch, which at 19 km2 is one of the largest lakes in the United Kingdom.ResultsA hierarchical Bayesian Jolly-Seber analysis of the data suggest that if individual differences in catchability are negligible the population of Ferox Trout in Loch Rannoch in 2009 was approximately 71 fish. The results also suggest that a single, often unaccompanied, highly-experienced angler was able to catch roughly 8% of the available fish on an annual basis.DiscussionIt is recommended that anglers adopt a precautionary approach and release all trout with a fork length ≥400 mm caught by trolling in Loch Rannoch. There is an urgent need to assess the status of Ferox Trout in other lakes. creator: Alastair Thorne creator: Alisdair I. MacDonald creator: Joseph L. Thorley uri: https://doi.org/10.7717/peerj.2646 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Thorne et al. title: Non-target effects of a glyphosate-based herbicide on Common toad larvae (Bufo bufo, Amphibia) and associated algae are altered by temperature link: https://peerj.com/articles/2641 last-modified: 2016-11-01 description: BackgroundGlyphosate-based herbicides are the most widely used pesticides in agriculture, horticulture, municipalities and private gardens that can potentially contaminate nearby water bodies inhabited by amphibians and algae. Moreover, the development and diversity of these aquatic organisms could also be affected by human-induced climate change that might lead to more periods with extreme temperatures. However, to what extent non-target effects of these herbicides on amphibians or algae are altered by varying temperature is not well known.MethodsWe studied effects of five concentrations of the glyphosate-based herbicide formulation Roundup PowerFlex (0, 1.5, 3, 4 mg acid equivalent glyphosate L−1 as a one time addition and a pulse treatment of totally 4 mg a.e. glyphosate L−1) on larval development of Common toads (Bufo bufo, L.; Amphibia: Anura) and associated algae communities under two temperature regimes (15 vs. 20 °C).ResultsHerbicide contamination reduced tail growth (−8%), induced the occurrence of tail deformations (i.e. lacerated or crooked tails) and reduced algae diversity (−6%). Higher water temperature increased tadpole growth (tail and body length (tl/bl) +66%, length-to-width ratio +4%) and decreased algae diversity (−21%). No clear relation between herbicide concentrations and tadpole growth or algae density or diversity was observed. Interactive effects of herbicides and temperature affected growth parameters, tail deformation and tadpole mortality indicating that the herbicide effects are temperature-dependent. Remarkably, herbicide-temperature interactions resulted in deformed tails in 34% of all herbicide treated tadpoles at 15 °C whereas no tail deformations were observed for the herbicide-free control at 15 °C or any tadpole at 20 °C; herbicide-induced mortality was higher at 15 °C but lower at 20 °C.DiscussionThese herbicide- and temperature-induced changes may have decided effects on ecological interactions in freshwater ecosystems. Although no clear dose-response effect was seen, the presence of glyphosate was decisive for an effect, suggesting that the lowest observed effect concentration (LOEC) in our study was 1.5 mg a.e. glyphosate L−1 water. Overall, our findings also question the relevance of pesticide risk assessments conducted at standard temperatures. creator: Fabian Baier creator: Edith Gruber creator: Thomas Hein creator: Elisabeth Bondar-Kunze creator: Marina Ivanković creator: Axel Mentler creator: Carsten A. Brühl creator: Bernhard Spangl creator: Johann G. Zaller uri: https://doi.org/10.7717/peerj.2641 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Baier et al. title: Whole-brain ex-vivo quantitative MRI of the cuprizone mouse model link: https://peerj.com/articles/2632 last-modified: 2016-11-01 description: Myelin is a critical component of the nervous system and a major contributor to contrast in Magnetic Resonance (MR) images. However, the precise contribution of myelination to multiple MR modalities is still under debate. The cuprizone mouse is a well-established model of demyelination that has been used in several MR studies, but these have often imaged only a single slice and analysed a small region of interest in the corpus callosum. We imaged and analyzed the whole brain of the cuprizone mouse ex-vivo using high-resolution quantitative MR methods (multi-component relaxometry, Diffusion Tensor Imaging (DTI) and morphometry) and found changes in multiple regions, including the corpus callosum, cerebellum, thalamus and hippocampus. The presence of inflammation, confirmed with histology, presents difficulties in isolating the sensitivity and specificity of these MR methods to demyelination using this model. creator: Tobias C. Wood creator: Camilla Simmons creator: Samuel A. Hurley creator: Anthony C. Vernon creator: Joel Torres creator: Flavio Dell’Acqua creator: Steve C.R. Williams creator: Diana Cash uri: https://doi.org/10.7717/peerj.2632 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Wood et al. title: Therapeutic role of human hepatocyte growth factor (HGF) in treating hair loss link: https://peerj.com/articles/2624 last-modified: 2016-11-01 description: Hepatocyte growth factor (HGF) is a paracrine hormone that plays an important role in epithelial-mesenchymal transition. HGF secreted by mesenchymal cells affects many properties of epithelial cells, such as proliferation, motility, and morphology. HGF has been reported to promote follicular growth. The purpose of the present study is to investigate the therapeutic role of HGF in hair loss treatment. A recombinant vector containing the human HGF (hHGF) gene (pTARGET-hHGF) was constructed, and the expression of hHGF in vitro was quantitatively and qualitatively evaluated. The effect of hHGF on hair growth was tested in mice, and results demonstrated that pTARGET-hHGF was successfully delivered into fibroblasts in vitro leading to a high expression of hHGF. Local injections of the pTARGET-hHGF recombinant vector into mice resulted in multiple beneficial effects compared to placebo, including faster hair regeneration, improved follicle development, and significantly increased HGF receptor (HGF-R). In conclusion, we have established a nonviral vector of hHGF which could be utilized to manipulate the sheath fibroblasts surrounding hair follicles (HF), thereby stimulating hair regeneration. creator: Yonghao Qi creator: Miao Li creator: Lian Xu creator: Zhijing Chang creator: Xiong Shu creator: Lijun Zhou uri: https://doi.org/10.7717/peerj.2624 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Qi et al. title: Production of cytotoxic compounds in dedifferentiated cells of Jatropha curcas L. (Euphorbiaceae) link: https://peerj.com/articles/2616 last-modified: 2016-11-01 description: This study addresses the in vitro culture as an alternative to obtain compounds with cytotoxic activity from the medicinal plant Jatropha curcas (Euphorbiaceae). We determined the presence of cytotoxic compounds in both whole plants and dedifferentiated cells. We evaluated the effect of auxin, cytokinins and light on callus induction in cotyledon explants. We found that the most effective combination to induce callus was the auxin 2,4-D (5 mM) with the cytokinin 6-BAP (2.5 mM), on Murashige-Skoog medium in darkness. We compared the callogenic potential among accessions from different geographic origins, finding that ARR-251107-MFG7 is most prone to form callus. The roots of J. curcas grown in field produced a compound chromatographically similar to the cytotoxic diterpene jatrophone. The profile of compounds extracted from the dedifferentiated cells was similar to that of the whole plant, including a relatively abundant stilbene-like compound. This study contributes to the future establishment of protocols to produce anti-cancer compounds from J. curcas cultivated in vitro. creator: Isidro Ovando-Medina creator: Leny P. Pérez-Díaz creator: Sonia Ruiz-González creator: Miguel Salvador-Figueroa creator: Marcos E. Urbina-Reyes creator: Lourdes Adriano-Anaya uri: https://doi.org/10.7717/peerj.2616 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Ovando-Medina et al. title: Ischemic postconditioning and pinacidil suppress calcium overload in anoxia-reoxygenation cardiomyocytes via down-regulation of the calcium-sensing receptor link: https://peerj.com/articles/2612 last-modified: 2016-11-01 description: Ischemic postconditioning (IPC) and ATP sensitive potassium channel (KATP) agonists (e.g. pinacidil and diazoxide) postconditioning are effective methods to defeat myocardial ischemia-reperfusion (I/R) injury, but their specific mechanisms of reducing I/R injury are not fully understood. We observed an intracellular free calcium ([Ca2+]i) overload in Anoxia/reoxygenation (A/R) cardiomyocytes, which can be reversed by KATP agonists diazoxide or pinacidil. The calcium-sensing receptor (CaSR) regulates intracellular calcium homeostasis. CaSR was reported to be involved in the I/R-induced apoptosis in rat cardiomyocytes. We therefore hypothesize that IPC and pinacidil postconditioning (PPC) reduce calcium overload in I/R cardiomyocytes by the down-regulation of CaSR. A/R model was established with adult rat caridomyocyte. mRNA and protein expression of CaSR were detected, IPC, PPC and KATP’s effects on [Ca2+]i concentration was assayed too. IPC and PPC ameliorated A/R insult induced [Ca2+]i overload in cardiomyocytes. In addition, they down-regulated the mRNA and protein level of CaSR as we expected. CaSR agonist spermine and KATP blocker glibenclamide offset IPC’s effects on CaSR expression and [Ca2+]i modulation. Our data indicate that CaSR down-regulation contributes to the mitigation of calcium overload in A/R cardiomyocytes, which may partially represents IPC and KATP’s myocardial protective mechanism under I/R circumstances. creator: Lin Zhang creator: Song Cao creator: Shengli Deng creator: Gang Yao creator: Tian Yu uri: https://doi.org/10.7717/peerj.2612 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Zhang et al. title: Root bacterial endophytes alter plant phenotype, but not physiology link: https://peerj.com/articles/2606 last-modified: 2016-11-01 description: Plant traits, such as root and leaf area, influence how plants interact with their environment and the diverse microbiota living within plants can influence plant morphology and physiology. Here, we explored how three bacterial strains isolated from the Populus root microbiome, influenced plant phenotype. We chose three bacterial strains that differed in predicted metabolic capabilities, plant hormone production and metabolism, and secondary metabolite synthesis. We inoculated each bacterial strain on a single genotype of Populus trichocarpa and measured the response of plant growth related traits (root:shoot, biomass production, root and leaf growth rates) and physiological traits (chlorophyll content, net photosynthesis, net photosynthesis at saturating light–Asat, and saturating CO2–Amax). Overall, we found that bacterial root endophyte infection increased root growth rate up to 184% and leaf growth rate up to 137% relative to non-inoculated control plants, evidence that plants respond to bacteria by modifying morphology. However, endophyte inoculation had no influence on total plant biomass and photosynthetic traits (net photosynthesis, chlorophyll content). In sum, bacterial inoculation did not significantly increase plant carbon fixation and biomass, but their presence altered where and how carbon was being allocated in the plant host. creator: Jeremiah A. Henning creator: David J. Weston creator: Dale A. Pelletier creator: Collin M. Timm creator: Sara S. Jawdy creator: Aimée T. Classen uri: https://doi.org/10.7717/peerj.2606 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Henning et al. title: A potential third Manta Ray species near the Yucatán Peninsula? Evidence for a recently diverged and novel genetic Manta group from the Gulf of Mexico link: https://peerj.com/articles/2586 last-modified: 2016-11-01 description: We present genetic and morphometric support for a third, distinct, and recently diverged group of Manta ray that appears resident to the Yucatán coastal waters of the Gulf of Mexico. Individuals of the genus Manta from Isla Holbox are markedly different from the other described manta rays in their morphology, habitat preference, and genetic makeup. Herein referred to as the Yucatán Manta Ray, these individuals form two genetically distinct groups: (1) a group of mtDNA haplotypes divergent (0.78%) from the currently recognized Manta birostris and M. alfredi species, and (2) a group possessing mtDNA haplotypes of M. birostris and highly similar haplotypes. The latter suggests the potential for either introgressive hybridization between Yucatán Manta Rays and M. birostris, or the retention of ancestral M. birostris signatures among Yucatán Manta Rays. Divergence of the genetically distinct Yucatán Manta Ray from M. birostris appears quite recent (<100,000 YBP) following fit to an Isolation-with-Migration model, with additional support for asymmetrical gene flow from M. birostris into the Yucatán Manta Ray. Formal naming of the Yucatán Manta Ray cannot yet be assigned until an in-depth taxonomic study and further confirmation of the genetic identity of existing type specimens has been performed. creator: Silvia Hinojosa-Alvarez creator: Ryan P. Walter creator: Pindaro Diaz-Jaimes creator: Felipe Galván-Magaña creator: E. Misty Paig-Tran uri: https://doi.org/10.7717/peerj.2586 license: http://creativecommons.org/licenses/by/4.0/ rights: ©2016 Hinojosa-Alvarez et al. title: Effects of CO2 laser irradiation on matrix-rich biofilm development formation–an in vitro study link: https://peerj.com/articles/2458 last-modified: 2016-11-01 description: BackgroundA carbon dioxide (CO2) laser has been used to morphologically and chemically modify the dental enamel surface as well as to make it more resistant to demineralization. Despite a variety of experiments demonstrating the inhibitory effect of a CO2 laser in reduce enamel demineralization, little is known about the effect of surface irradiated on bacterial growth. Thus, this in vitro study was preformed to evaluate the biofilm formation on enamel previously irradiated with a CO2 laser (λ = 10.6 µM).MethodsFor this in vitro study, 96 specimens of bovine enamel were employed, which were divided into two groups (n = 48): 1) Control-non-irradiated surface and 2) Irradiated enamel surface. Biofilms were grown on the enamel specimens by one, three and five days under intermittent cariogenic condition in the irradiated and non-irradiated surface. In each assessment time, the biofilm were evaluated by dry weigh, counting the number of viable colonies and, in fifth day, were evaluated by polysaccharides analysis, quantitative real time Polymerase Chain Reaction (PCR) as well as by contact angle. In addition, the morphology of biofilms was characterized by fluorescence microscopy and field emission scanning electron microscopy (FESEM). Initially, the assumptions of equal variances and normal distribution of errors were conferred and the results are analyzed statistically by t-test and Mann Whitney test.ResultsThe mean of log CFU/mL obtained for the one-day biofilm evaluation showed that there is statistical difference between the experimental groups. When biofilms were exposed to the CO2 laser, CFU/mL and CFU/dry weight in three day was reduced significantly compared with control group. The difference in the genes expression (Glucosyltransferases (gtfB) and Glucan-binding protein (gbpB)) and polysaccharides was not statically significant. Contact angle was increased relative to control when the surface was irradiated with the CO2 laser. Similar morphology was also visible with both treatments; however, the irradiated group revealed evidence of melting and fusion in the specimens.ConclusionIn conclusion, CO2 laser irradiation modifies the energy surface and disrupts the initial biofilm formation. creator: Bruna Raquel Zancopé creator: Vanessa B. Dainezi creator: Marinês Nobre-dos-Santos creator: Sillas Duarte creator: Vanessa Pardi creator: Ramiro M. Murata uri: https://doi.org/10.7717/peerj.2458 license: http://creativecommons.org/licenses/by/4.0/ rights: © 2016 Zancope et al.