Online ISSN: 2167-8359
Regional association plots for three HLA-DQB1*06:02 negative EHS risk loci. (A) The region 2q21.2 Cochran-Armitage trend test P-value, the SNP rs16826003 is located in an intron of NCKAP5 gene (B) The region 15q14 Cochran-Armitage trend test, SNP rs11854769 is located 42 kb upstream of the nearest gene, SPRED1 (C) The 9q34 region based on P-minimum, rs10988217 is located in an intron of CRAT gene. Each of the top markers is indicated by purple diamonds. SNPs that were genotyped using Affymetrix 6.0 are marked by triangles. Imputed SNPs are plotted as circles. The color intensity represents the extent of the LD with the marker SNP, red (r 2 ≥ 0.8), orange (0.6 ≤ 0.8), green (0.4 ≤ 0.6), light blue (0.2 ≤ 0.4), and dark blue (r 2 ≤ 0.2). Light blue in the background indicates local recombination rate.
List of SNPs of interest for HLA-DQB1*06:02 negative hypersomnias samples in Caucasian population.
(A) The region 2q21.2 Cochran–Armitage trend test P-value, the SNP rs16826003 is located in an intron of NCKAP5 gene (B) The region 15q14 Cochran–Armitage trend test, SNP rs11854769 is located 42 kb upstream of the nearest gene, SPRED1 (C) The 9q34 region based on P-minimum, rs10988217 is located in an intron of CRAT gene. Each of the top markers is indicated by purple diamonds. SNPs that were genotyped using Affymetrix 6.0 are marked by triangles. Imputed SNPs are plotted as circles. The color intensity represents the extent of the LD with the marker SNP, red (r2 ≥ 0.8), orange (0.6 ≤ 0.8), green (0.4 ≤ 0.6), light blue (0.2 ≤ 0.4), and dark blue (r2 ≤ 0.2). Light blue in the background indicates local recombination rate.
Essential hypersomnia (EHS), a sleep disorder characterized by excessive daytime sleepiness, can be divided into two broad classes based on the presence or absence of the HLA-DQB1*06:02 allele. HLA-DQB1*06:02-positive EHS and narcolepsy with cataplexy are associated with the same susceptibility genes. In contrast, there are fewer studies of HLA-DQB1*06:02 negative EHS which, we hypothesized, involves a different pathophysiological pathway than does narcolepsy with cataplexy. In order to identify susceptibility genes associated with HLA-DQB1*06:02 negative EHS, we conducted a genome-wide association study (GWAS) of 125 unrelated Japanese EHS patients lacking the HLA-DQB1*06:02 allele and 562 Japanese healthy controls. A comparative study was also performed on 268 HLA-DQB1*06:02 negative Caucasian hypersomnia patients and 1761 HLA-DQB1*06:02 negative Caucasian healthy controls. We identified three SNPs that each represented a unique locus- rs16826005 (P = 1.02E-07; NCKAP5), rs11854769 (P = 6.69E-07; SPRED1), and rs10988217 (P = 3.43E-06; CRAT) that were associated with an increased risk of EHS in this Japanese population. Interestingly, rs10988217 showed a similar tendency in its association with both HLA-DQB1*06:02 negative EHS and narcolepsy with cataplexy in both Japanese and Caucasian populations. This is the first GWAS of HLA-DQB1*06:02 negative EHS, and the identification of these three new susceptibility loci should provide additional insights to the pathophysiological pathway of this condition.
Activity of thiazole and thiadiazole analogs of the BTD series against M. tuberculosis and Vero cell line. 
We demonstrated that the 3-substituted benzothiophene-1,1-dioxide class of compounds are effective inhibitors of Mycobacterium tuberculosis growth under aerobic conditions. We examined substitution at the C-3 position of the benzothiophene-1,1-dioxide series systematically to delineate structure-activity relationships influencing potency and cytotoxicity. Compounds were tested for inhibitory activity against virulent M. tuberculosis and eukaryotic cells. The tetrazole substituent was most potent, with a minimum inhibitory concentration (MIC) of 2.6 µM. However, cytotoxicity was noted with even more potency (Vero cell TC50 = 0.1 µM). Oxadiazoles had good anti-tubercular activity (MICs of 3-8 µM), but imidazoles, thiadiazoles and thiazoles had little activity. Cytotoxicity did not track with anti-tubercular activity, suggesting different targets or mode of action between bacterial and eukaryotic cells. However, we were unable to derive analogs without cytotoxicity; all compounds synthesized were cytotoxic (TC50 of 0.1-5 µM). We conclude that cytotoxicity is a liability in this series precluding it from further development. However, the series has potent anti-tubercular activity and future efforts towards identifying the mode of action could result in the identification of novel drug targets.
Cat impoundments were increasing at the municipal San Jose animal shelter in 2009, despite long-term successful low cost sterilization programs and attempts to lower the euthanasia rate of treatable-rehabilitatable impounds beginning in 2008. San Jose Animal Care and Services implemented a new strategy designed to control overall feral cat reproduction by altering and returning feral cats entering the shelter system, rather than euthanizing the cats. The purpose of this case study was to determine how the program affected the shelter cat intakes over time. In just over four years, 10,080 individual healthy adult feral cats, out of 11,423 impounded at the shelter during this time frame, were altered and returned to their site of capture. Included in the 11,423 cats were 862 cats impounded from one to four additional times for a total of 958 (9.5%) recaptures of the previously altered 10,080 cats. The remaining 385 healthy feral cats were euthanized at the shelter from March 2010 to June 2014. Four years into the program, researchers observed cat and kitten impounds decreased 29.1%; euthanasia decreased from over 70% of intakes in 2009, to 23% in 2014. Euthanasia in the shelter for Upper Respiratory Disease decreased 99%; dead cat pick up off the streets declined 20%. Dog impounds did not similarly decline over the four years. No other laws or program changes were implemented since the beginning of the program.
Suppressor of cytokine signaling 3 (SOCS3) is a key regulator of cytokine signaling in macrophages and T cells. Although SOCS3 seems to contribute to the balance between the pro-inflammatory actions of IL-6 family of cytokines and anti-inflammatory signaling of IL-10 by negatively regulating gp130/Jak/Stat3 signal transduction, how and the molecular mechanisms whereby SOCS3 controls the downstream impact of TLR4 are largely unknown and current data are controversial. Furthermore, very little is known regarding SOCS3 function in cells other than myeloid cells and T cells. Our previous study demonstrates that SOCS3 is expressed in osteoblasts and functions as a critical inhibitor of LPS-induced IL-6 expression. However, the function of SOCS3 in osteoblasts remains largely unknown. In the current study, we report for the first time that LPS stimulation of osteoblasts induces the transcriptional activation of matrix metalloproteinase (MMP)-13, a central regulator of bone resorption. Importantly, we demonstrate that SOCS3 overexpression leads to a significant decrease of LPS-induced MMP-13 expression in both primary murine calvariae osteoblasts and a mouse osteoblast-like cell line, MC3T3-E1. Our findings implicate SOCS3 as an important regulatory mediator in bone inflammatory diseases by targeting MMP-13.
Insulin secretion from pancreatic β-cells plays an essential role in blood glucose homeostasis and type 2 diabetes. Many genes are involved in the secretion of insulin and most of these genes can be targeted by microRNAs (miRNAs). However, the role of miRNAs in insulin secretion and type 2 diabetes has not been exhaustively studied. The expression miR-184, a miRNA enriched in pancreatic islets, negatively correlates with insulin secretion, suggesting that it is a good candidate for miRNA-mediated regulation of insulin secretion. Here we report that miR-184 inhibits insulin secretion in the MIN6 pancreatic β-cell line through the repression of its target Slc25a22, a mitochondrial glutamate carrier. Our study provides new insight into the regulation of insulin secretion by glutamate transport in mitochondria.
Helminths include both parasitic nematodes (roundworms) and platyhelminths (trematode and cestode flatworms) that are abundant, and are of clinical importance. The genetic characterization of parasitic flatworms using advanced molecular tools is central to the diagnosis and control of infections. Although the nuclear genome houses suitable genetic markers (e.g., in ribosomal (r) DNA) for species identification and molecular characterization, the mitochondrial (mt) genome consistently provides a rich source of novel markers for informative systematics and epidemiological studies. In the last decade, there have been some important advances in mtDNA genomics of helminths, especially lung flukes, liver flukes and intestinal flukes. Fasciolopsis buski, often called the giant intestinal fluke, is one of the largest digenean trematodes infecting humans and found primarily in Asia, in particular the Indian subcontinent. Next-generation sequencing (NGS) technologies now provide opportunities for high throughput sequencing, assembly and annotation within a short span of time. Herein, we describe a high-throughput sequencing and bioinformatics pipeline for mt genomics for F. buski that emphasizes the utility of short read NGS platforms such as Ion Torrent and Illumina in successfully sequencing and assembling the mt genome using innovative approaches for PCR primer design as well as assembly. We took advantage of our NGS whole genome sequence data (unpublished so far) for F. buski and its comparison with available data for the Fasciola hepatica mtDNA as the reference genome for design of precise and specific primers for amplification of mt genome sequences from F. buski. A long-range PCR was carried out to create an NGS library enriched in mt DNA sequences. Two different NGS platforms were employed for complete sequencing, assembly and annotation of the F. buski mt genome. The complete mt genome sequences of the intestinal fluke comprise 14,118 bp and is thus the shortest trematode mitochondrial genome sequenced to date. The noncoding control regions are separated into two parts by the tRNA-Gly gene and don't contain either tandem repeats or secondary structures, which are typical for trematode control regions. The gene content and arrangement are identical to that of F. hepatica. The F. buski mtDNA genome has a close resemblance with F. hepatica and has a similar gene order tallying with that of other trematodes. The mtDNA for the intestinal fluke is reported herein for the first time by our group that would help investigate Fasciolidae taxonomy and systematics with the aid of mtDNA NGS data. More so, it would serve as a resource for comparative mitochondrial genomics and systematic studies of trematode parasites.
We show for the first time the effects of heavy-hydrogen water ((2)H2O) and heavy-oxygen water (H2 (18)O) on the gliding speed of microtubules on kinesin-1 coated surfaces. Increased fractions of isotopic waters used in the motility solution decreased the gliding speed of microtubules by a maximum of 21% for heavy-hydrogen and 5% for heavy-oxygen water. We also show that gliding microtubule speed returns to its original speed after being treated with heavy-hydrogen water. We discuss possible interpretations of these results and the importance for future studies of water effects on kinesin and microtubules. We also discuss the implication for using heavy waters in biomolecular devices incorporating molecular motors.
Cattle density and distribution 1960s and 2000s. Estimated cattle (heads per square kilometre) density around 1960 (upper left), 2000 (upper right) and difference (2000-1960, bottom) relative to the mean.
Effect of climate variability on cattle holdings. Upper panel (left to right): Sign of slope for precipitation (P), wet season temperature (T w ), and dry season temperature (T d ). Positive values means increased precipitation/temperature is associated with increased number of cattle and vice versa. Middle panel (left to right): Percent variance explained by precipitation, wet season temperature and dry season temperature. Lower panel (left to right): Total variance explained by the model, variance explained by variability in Gompertz function, and variance explained by climate variability.  
Climate sensitivity. Left: Relationship between the variance explained by the Gompertz function and climate. In countries where the number of cattle fit well with the Gompertz curve there is little climate sensitivity. Where the carrying capacity is close to its limit the cattle populations will fluctuate around the mean and hence the Gompertz function will not explain the variance. In these countries climate variability is the major source of variation. Vertical lines are box-and-whisker diagrams plotted at quantiles 0–0.1 to 0.9–1 by 0.1. Black dot indicate median for each quantile. Right: The effect of one standard deviation increase in precipitation vs annual mean rainfall (1961–1990). Y-axis is in %, and show the change in cattle numbers relative to the mean number of cattle (1961–1990). A value of 50 on the y-axis would indicate one standard deviation of rainfall (over five years) would increase the national cattle holdings by 50% relative to the mean. A value of −50 would indicate a reduction of 50%. Vertical lines are box-and-whisker diagrams plotted at quantiles 0–0.25 to 0.75–1 by 0.25. Black dot indicate median for each quantile.  
Future precipitation patterns. The mean expected changes from 1961-2000 in precipitation under three different climate scenarios. Shading indicate standard deviations, black contours show the presence of cattle in the 1960s. Only values where more than 66% of the models agree are shown. Black dots indicate more than 90% of the models agree on the sign of change.
The role of cattle in developing countries is as a source of high-quality food, as draft animals, and as a source of manure and fuel. Cattle represent important contribution to household incomes, and in drought prone areas they can act as an insurance against weather risk. So far, no studies have addressed how historical variations in temperature and rainfall have influenced cattle populations in Africa. The focus of this study is to assess the historical impact of climate variability on national cattle holdings. We reconstruct the cattle density and distribution for two time periods; 1955-1960 and 2000-2005. Based on estimates from FAO and official numbers, we generated a time series of cattle densities from 1961-2008, and compared these data with precipitation and temperature anomalies for the same period. We show that from 1961-2008 rainfall and temperature have been modulating, and occasionally controlling, the number of cattle in Africa.
The measurable benefits of animal control programs are unknown and the aim of this study was to determine the impact of these programs on pet population changes. A prospective cross-sectional study of 1000 households was implemented in 2005 to evaluate characteristics of the owned and unowned population of dogs and cats in Santa Clara County, California. The same population was previously studied 12 years earlier. During this time period, the county instituted in 1994 and then subsequently disestablished a municipal spay/neuter voucher program for cats. Dog intakes declined from 1992-2005, as they similarly did for an adjacent county (San Mateo). However, cat intakes declined significantly more in Santa Clara County than San Mateo, with an average annual decline of approximately 700 cats for the 12 year period. Time series analysis showed a greater than expected decline in the number of cats surrendered to shelters in Santa Clara County during the years the voucher program was in effect (1994-2005). The net savings to the county by reducing the number of cat shelter intakes was estimated at approximately $1.5 million. The measurable benefits of animal control programs are unknown and the aim of this study was to determine the impact of these programs on pet population changes.
Originally identified as an ornithischian dinosaur, Crosbysaurus harrisae has been found in New Mexico, Arizona, and its type locality in Texas, as well as in North Carolina. The genus has been reassessed by other workers in light of reinterpretations about the postcrania of another putative Triassic ornithischian, Revueltosaurus. The understanding of Triassic dental faunas has become more complicated by the extreme convergence between pseudosuchian archosaurs and ornithischian dinosaur dental morphologies. We report here on a new specimen of Crosbysaurus (MNA V10666) from the Chinle Formation at Comb Ridge in southeastern Utah. This new specimen is assigned to Crosbysaurus sp. on the basis of the unique compound posterior denticles, labiolingual width, and curvature. While MNA V10666 does not help resolve the affinities of Crosbysaurus, it does represent the extension of the geographic range of this taxon for approximately 250 kilometers. This is the first record of the genus Crosbysaurus in Utah and as such it represents the northernmost known record of this taxon. This indicates that Crosbysaurus was not limited to the southern area of the Chinle/Dockum deposition but instead was widespread across the Late Triassic paleoriver systems of western Pangea. The reported specimen was found in close association with a typical Late Triassic Chinle fauna, including phytosaurs, metoposaurs, and dinosauromorphs.
Background. Despite the growing share of neonatal mortality in under-5 mortality in the recent decades in India, most studies have focused on infant and child mortality putting neonatal mortality on the back seat. The development of focused and evidence-based health interventions to reduce neonatal mortality warrants an examination of factors affecting it. Therefore, this study attempt to examine individual, household, and community level factors affecting neonatal mortality in rural India.Data and methods. We analysed information on 171,529 singleton live births using the data from the most recent round of the District Level Household Survey conducted in 2007–08. Principal component analysis was used to create an asset index. Two-level logistic regression was performed to analyse the factors associated with neonatal deaths in rural India.Results. The odds of neonatal death were lower for neonates born to mothers with secondary level education (O R = 0.60, p = 0.01) compared to those born to illiterate mothers. A progressive reduction in the odds occurred as the level of fathers’ education increased. The odds of neonatal death were lower for infants born to unemployed mothers (O R = 0.89, p = 0.00) compared to those who worked as agricultural worker/farmer/laborer. The odds decreased if neonates belonged to Scheduled Tribes (O R = 0.72, p = 0.00) or ‘Others’ caste group (O R = 0.87, p = 0.04) and to the households with access to improved sanitation (O R = 0.87, p = 0.02), pucca house (O R = 0.87, p = 0.03) and electricity (O R = 0.84, p = 0.00). The odds were higher for male infants (O R = 1.21, p = 0.00) and whose mother experienced delivery complications (O R = 1.20, p = 0.00). Infants whose mothers received two tetanus toxoid injections (O R = 0.65, p = 0.00) were less likely to die in the neonatal period. Children of higher birth order were less likely to die compared to first birth order.Conclusion. Ensuring the consumption of an adequate quantity of Tetanus Toxoid (TT) injections by pregnant mothers, targeting vulnerable groups like young, first time and Scheduled Caste mothers, and improving overall household environment by increasing access to improved toilets, electricity, and pucca houses could also contribute to further reductions in neonatal mortality in rural India. Any public health interventions aimed at reducing neonatal death in rural India should consider these factors.
The causes of bee declines remain hotly debated, particularly the contribution of neonicotinoid insecticides. In 2013 the UK's Food & Environment Research Agency made public a study of the impacts of exposure of bumblebee colonies to neonicotinoids. The study concluded that there was no clear relationship between colony performance and pesticide exposure, and the study was subsequently cited by the UK government in a policy paper in support of their vote against a proposed moratorium on some uses of neonicotinoids. Here I present a simple re-analysis of this data set. It demonstrates that these data in fact do show a negative relationship between both colony growth and queen production and the levels of neonicotinoids in the food stores collected by the bees. Indeed, this is the first study describing substantial negative impacts of neonicotinoids on colony performance of any bee species with free-flying bees in a field realistic situation where pesticide exposure is provided only as part of normal farming practices. It strongly suggests that wild bumblebee colonies in farmland can be expected to be adversely affected by exposure to neonicotinoids.
Various Drosophila species and source databases used for the analysis. The GC% for each genome was calculated using infoseq from the EMBOSS package. 
The consensus phylogenetic tree obtained by combining the trees obtained for each of the 13 enzymes. The phylogenetic tree for each enzyme was calculated by extracting the corresponding fragments and then counting the number of shared fragment between every pair of species. The upper branch support values represent the percentage agreement over 13 enzymes and the bottom values indicate number of enzymes out of total 13 enzymes supporting the branch.
Total number of fragments generated using 13 different Type IIB restriction enzymes for each of the 21 Drosophila genomes. 
Type IIB restriction endonucleases are site-specific endonucleases that cut both strands of double-stranded DNA upstream and downstream of their recognition sequences. These restriction enzymes have recognition sequences that are generally interrupted and range from 5 to 7 bases long. They produce DNA fragments which are uniformly small, ranging from 21 to 33 base pairs in length (without cohesive ends). The fragments are generated from throughout the entire length of a genomic DNA providing an excellent fractional representation of the genome. In this study we simulated restriction enzyme digestions on 21 sequenced genomes of various Drosophila species using the predicted targets of 16 Type IIB restriction enzymes to effectively produce a large and arbitrary selection of loci from these genomes. The fragments were then used to compare organisms and to calculate the distance between genomes in pair-wise combination by counting the number of shared fragments between the two genomes. Phylogenetic trees were then generated for each enzyme using this distance measure and the consensus was calculated. The consensus tree obtained agrees well with the currently accepted tree for the Drosophila species. We conclude that multi-locus sub-genomic representation combined with next generation sequencing, especially for individuals and species without previous genome characterization, can accelerate studies of comparative genomics and the building of accurate phylogenetic trees.
Longitudinal KSA 2011 THC residents comparison to baseline. Longitudinal KSA comparisons for THC residents at the pre-defined time points compared to their baseline. 
KSA compared between THC residents (n = 12) and TR Residents (n = 12). KSA comparisons between TR and THC residents at pre-defined time points. 
KSA2011 TR residents comparison to baseline. Longitudinal KSA comparisons for TR residents at pre-defined time points compared to their baseline. 
Purpose. The effect of patient centered medical home (PCMH) curriculum interventions on residents’ self-reported and demonstrated knowledge, skills and attitudes in PCMH competency arenas (KSA) is lacking in the literature. This study aimed to assess the impact of PCMH curricular innovations on the KSA of Internal Medicine residents. Methods. Twenty four (24) Internal Medicine residents—12 Traditional (TR) track residents and 12 Teaching Health Center (THC) track residents—began training in Academic Year (AY) 2011 at the Wright Center for Graduate Medical Education (WCGME). They were followed through AY2013, covering three years of training. PCMH curricular innovations were focally applied July 2011 until May 2012 to THC residents. These curricular innovations were spread program-wide in May 2012. Semi-annual, validated PCMH Clinician Assessments assessing KSA were started in AY2011 and were completed by all residents. Results. Mean KSA scores of TR residents were similar to those of THC residents at baseline for all PCMH competencies. In May 2012, mean scores of THC residents were significantly higher than TR residents for most KSA. After program-wide implementation of PCMH innovations, mean scores of TR residents for all KSA improved and most became equalized to those of THC residents. Globally improved KSA scores of THC and TR residents were maintained through May 2014, with the majority of improvements above baseline and reaching statistical significance. Conclusions. PCMH curricular innovations inspired by Health Resources and Services Administration (HRSA’s) Teaching Health Center funded residency program expansion quickly and consistently improved the KSA of Internal Medicine residents.
Main characteristics of studies included in this meta-analysis. 
Quality assessment of studies. 
Meta-analysis of polymorphisms of rs7517847. 
Meta-analysis of polymorphisms of rs2201841. 
To comprehensively evaluate the association between rs7517847 and rs2201841 polymorphisms in the Interleukin-23 (IL-23) receptor gene and ankylosing spondylitis (AS), a meta-analysis was performed. The Pubmed, Embase, MEDLINE, Cochrane, China National Knowledge Infrastructure (CNKI), VIP, Wanfang and China Biology Medicine disc (CBMdisc) databases were searched to identify eligible studies on rs7517847 and rs2201841 polymorphisms in the IL-23 receptor gene and AS that were published through September 2014. Data of interest were extracted from each study, and the meta-analysis was performed using STATA 12.0. Four studies were eligible for the meta-analysis and included a total patient population of 2,465. With regards to rs7517847, the current study showed that the genotype GG and allele G might play a protective role during AS (OR = 0.76, 95% CI [0.59-0.99]; OR = 0.88, 95% CI [0.78-0.99] for homozygote and allelic models, respectively). However, according to the meta-analysis, there was no statistical association between the genotype or allele of rs2201841 and an individual's susceptibility to AS in all genetic models. In conclusion, it was the IL-23 rs7517847 polymorphism rather than the rs2201841 polymorphism that had a statistical association with AS. Nevertheless, more evidence is needed to confirm this result. Consequently, it is necessary to carry out more high-quality studies to confirm the associations between these two single nucleotide polymorphisms and AS.
Castration-resistant prostate cancer (CRPC) expresses high levels of the anti-apoptotic proteins Bcl-2, Bcl-xL and Mcl-1, resulting in resistance to apoptosis and association with poor prognosis. Docetaxel, an antimitotic drug that is the first-line treatment strategy for CRPC, is known to provide a small survival benefit. However, docetaxel chemotherapy alone is not enough to counteract the high levels of Bcl-2/Bcl-xL/Mcl-1 present in CRPC. ABT-737 is a small molecule that binds to Bcl-2/Bcl-xL (but not Mcl-1) with high affinity and disrupts their interaction with pro-apoptotic Bax/Bak, thus enhancing apoptosis. Our results indicate that ABT-737 can sensitize androgen-dependent LNCaP and CRPC PC3 cells to docetaxel- and to the novel antimitotic ENMD-1198-mediated caspase-dependent apoptosis. CRPC DU145 cells, however, are more resistant to ABT-737 because they are Bax null and not because they express the highest levels of anti-apoptotic Mcl-1 (associated with ABT-737 resistance). Knockdown of Bax or Bak in LNCaP indicates that ABT-737-induced antimitotic enhancement of apoptosis is more dependent on the levels of Bax than Bak. Furthermore, we find that the ability of docetaxel to increase cyclin B1/Cdk1-mediated phosphorylation of Bcl-2/Bcl-xL and decrease Mcl-1 is required for ABT-737 to enhance apoptosis in PC3 cells, as determined by addition of Cdk1 inhibitor purvalanol A and expression of shRNA specific for cyclin B1. Overall, our data suggests that the high levels of anti-apoptotic proteins in Bax-expressing CRPC cells can be overcome by targeting Bcl-2/Bcl-xL with ABT-737 and Mcl-1 with antimitotics.
We analyzed the applicability of high-resolution (2)H-HMR spectroscopy for the analysis of microbe metabolism in samples of mitochondrion isolated from rat liver and from aqueous extracts of homogenates of rat liver and other organs and tissues in the presence of high D2O contents. Such analysis is possible due to the fast microbe adaptation to life in the heavy water. It is also shown that some enzymatic processes typical for the intact cells are preserved in the homogenized tissue preparations. The microbial and cellular metabolic processes can be differentiated via the strategic use of cell poisons and antibiotics.
Species traits may provide a short-cut to predicting generalities in species turnover in response to environmental change, particularly for poorly known taxa. We ask if morphological traits of assemblages respond predictably to macrohabitats across a large scale. Ant assemblages were collected at nine paired pasture and remnant sites from within three areas along a 300 km distance. We measured ten functional morphological traits for replicate individuals of each species. We used a fourth corner model to test associations between microhabitat variables, macrohabitats (pastures and remnants) and traits. In addition, we tested the phylogenetic independence of traits, to determine if responses were likely to be due to filtering by morphology or phylogeny. Nine of ten traits were predicted by macrohabitat and the majority of these traits were independent of phylogeny. Surprisingly, microhabitat variables were not associated with morphological traits. Traits which were associated with macrohabitats were involved in locomotion, feeding behaviour and sensory ability. Ants in remnants had more maxillary palp segments, longer scapes and wider eyes, while having shorter femurs, smaller apical mandibular teeth and shorter Weber's lengths. A clear relationship between traits and macrohabitats across a large scale suggests that species are filtered by coarse environmental differences. In contrast to the findings of previous studies, fine-scale filtering of morphological traits was not apparent. If such generalities in morphological trait responses to habitat hold across even larger scales, traits may prove critical in predicting the response of species assemblages to global change.
Leishmania amazonensis is the causative agent of American cutaneous leishmaniasis, an important neglected tropical disease. Once Leishmania amazonensis is inoculated into the human host, promastigotes are exposed to the extracellular matrix (ECM) of the dermis. However, little is known about the interaction between the ECM and Leishmania promastigotes. In this study we established L. amazonensis promastigote culture in a three-dimensional (3D) environment mainly composed of Collagen I (COL I). This 3D culture recreates in vitro some aspects of the human host infection site, enabling the study of the interaction mechanisms of L. amazonensis with the host ECM. Promastigotes exhibited "freeze and run" migration in the 3D COL I matrix, which is completely different from the conventional in vitro swimming mode of migration. Moreover, L. amazonensis promastigotes were able to invade, migrate inside, and remodel the 3D COL I matrix. Promastigote trans-matrix invasion and the freeze and run migration mode were also observed when macrophages were present in the matrix. At least two classes of proteases, metallo- and cysteine proteases, are involved in the 3D COL I matrix degradation caused by Leishmania. Treatment with a mixture of protease inhibitors significantly reduced promastigote invasion and migration through this matrix. Together our results demonstrate that L. amazonensis promastigotes release proteases and actively remodel their 3D environment, facilitating their migration. This raises the possibility that promastigotes actively interact with their 3D environment during the search for their cellular "home"-macrophages. Supporting this hypothesis, promastigotes migrated faster than macrophages in a novel 3D co-culture model.
Inertial properties of body segments, such as mass, centre of mass or moments of inertia, are important parameters when studying movements of the human body. However, these quantities are not directly measurable. Current approaches include using regression models which have limited accuracy: geometric models with lengthy measuring procedures or acquiring and post-processing MRI scans of participants. We propose a geometric methodology based on 3D photogrammetry using multiple cameras to provide subject-specific body segment parameters while minimizing the interaction time with the participants. A low-cost body scanner was built using multiple cameras and 3D point cloud data generated using structure from motion photogrammetric reconstruction algorithms. The point cloud was manually separated into body segments, and convex hulling applied to each segment to produce the required geometric outlines. The accuracy of the method can be adjusted by choosing the number of subdivisions of the body segments. The body segment parameters of six participants (four male and two female) are presented using the proposed method. The multi-camera photogrammetric approach is expected to be particularly suited for studies including populations for which regression models are not available in literature and where other geometric techniques or MRI scanning are not applicable due to time or ethical constraints.
Effect of the PKA inhibitor H-89 on rates of lipolysis. 3T3-L1 adipocytes were treated for 4 h with glucose plus no further additions (Ctl) or 5 mM butyrate (But). Isoproterenol (1 µM) and H-89 (50 µM) were added as indicated and glycerol was measured after another 30 min. Data depicted are representative of three or more independent experiments. Data shown are means ± SE (n = 3). * * P < 0.01. 
Effect of butyrate on MAPK and perilipin. 3T3-L1 adipocytes were treated with or without 25 mM glucose and 50 ng/ml TNF-α or 5 mM butyrate for 24 h. Total protein extracts were prepared and Western blots were performed with antibodies raised against (A) perilipin (approx 57 kDa), (B) phosphorylated MAP Kinase or (C) total MAP Kinase (ERK 1 and 2 ran at approx. 44 and 42 kDa, respectively). Data depicted are representative of three or more independent experiments. 
Effect of glucose on the lipolytic effect of butyrate and trichostatin A. (A) 3T3-L1 adipocytes were incubated in glucose free media supplemented with pyruvate for 24 h with no additions (Ctl), 50 ng/ml TNF-α (TNF), 5 mM butyrate (But), 25 mM glucose (Glc), or in combinations as indicated. Cells were washed and glycerol release measured as in the legend for Fig. 1. (B) 3T3-L1 adipocytes were treated with 1 µM Trichostatin A (TSA) in the presence or absence of 25 mM glucose. Data depicted are representative of three or more independent experiments. Data shown are means ± SE (n = 3). * * * P < 0.001. 
Effect of butyrate on phosphorylation of AMPK. 3T3-L1 adipocytes were treated with or without 25 mM glucose and 50 ng/ml TNF-α or 5 mM butyrate for 6 h. Total protein extracts were prepared and Western blots were performed with antibodies raised against phospho-AMPK (T172) (A) or total AMPK (B). Both ran at approx. 62 kDa. Data depicted are representative of three or more independent experiments. 
Effect of iodoacetate and dichloroacetate. (A) 3T3-L1 adipocytes were treated in 25 mM glucose for 6 h with no additions (ctl), 50 ng/ml TNF-α (TNF), or 5 mM butyrate. (B) Adipocytes were treated with TNF and glucose and either 100 µM iodoacetate (Iod) or 100 µM dichloroacetate (DCA) and then glycerol release was measured as in the legend for Fig. 1. (C) Same conditions as (B), but with butyrate. Data depicted are representative of three or more independent experiments. Data shown are means ± SE (n = 3). * P < 0.05; * * P < 0.01. 
We determined the effect of butyrate and other short-chain fatty acids (SCFA) on rates of lipolysis in 3T3-L1 adipocytes. Prolonged treatment with butyrate (5 mM) increased the rate of lipolysis approximately 2-3-fold. Aminobutyric acid and acetate had little or no effect on lipolysis, however propionate stimulated lipolysis, suggesting that butyrate and propionate act through their shared activity as histone deacetylase (HDAC) inhibitors. Consistent with this, the HDAC inhibitor trichostatin A (1 µM) also stimulated lipolysis to a similar extent as did butyrate. Western blot data suggested that neither mitogen-activated protein kinase (MAPK) activation nor perilipin down-regulation are necessary for SCFA-induced lipolysis. Stimulation of lipolysis with butyrate and trichostatin A was glucose-dependent. Changes in AMP-activated protein kinase (AMPK) phosphorylation mediated by glucose were independent of changes in rates of lipolysis. The glycolytic inhibitor iodoacetate prevented both butyrate- and tumor necrosis factor-alpha-(TNF-α) mediated increases in rates of lipolysis indicating glucose metabolism is required. However, unlike TNF-α- , butyrate-stimulated lipolysis was not associated with increased lactate release or inhibited by activation of pyruvate dehydrogenase (PDH) with dichloroacetate. These data demonstrate an important relationship between lipolytic activity and reported HDAC inhibitory activity of butyrate, other short-chain fatty acids and trichostatin A. Given that HDAC inhibitors are presently being evaluated for the treatment of diabetes and other disorders, more work will be essential to determine if these effects on lipolysis are due to inhibition of HDAC.
A comparison of iSNO-AAPair with the existing prediction methods a via the independent dataset test for the four different metrics (cf. Eq. (19)). 
A comparison of iSNO-AAPair with the existing prediction methods a on the 14 independent proteins (cf. Supplemental Information S3). 
A schematic illustration to show a peptide generated from a protein sequence by the sliding window (Chou, 2001d) with cysteine (C) located at its center. Adapted from Chou (Chou, 2001b) with permission.
A schematic drawing to show the pairwise coupling between nearest residues (blue solid line) and that between the next nearest residues (red dashed line).
As one of the most important and universal posttranslational modifications (PTMs) of proteins, S-nitrosylation (SNO) plays crucial roles in a variety of biological processes, including the regulation of cellular dynamics and many signaling events. Knowledge of SNO sites in proteins is very useful for drug development and basic research as well. Unfortunately, it is both time-consuming and costly to determine the SNO sites purely based on biological experiments. Facing the explosive protein sequence data generated in the post-genomic era, we are challenged to develop automated vehicles for timely and effectively determining the SNO sites for uncharacterized proteins. To address the challenge, a new predictor called iSNO-AAPair was developed by taking into account the coupling effects for all the pairs formed by the nearest residues and the pairs by the next nearest residues along protein chains. The cross-validation results on a state-of-the-art benchmark have shown that the new predictor outperformed the existing predictors. The same was true when tested by the independent proteins whose experimental SNO sites were known. A user-friendly web-server for iSNO-AAPair was established at, by which users can easily obtain their desired results without the need to follow the mathematical equations involved during its development.
Leaf-tying caterpillars act as ecosystem engineers by building shelters between overlapping leaves, which are inhabited by other arthropods. Leaf-tiers have been observed to leave their ties and create new shelters (and thus additional microhabitats), but the ecological factors affecting shelter fidelity are poorly known. For this study, we explored the effects of resource limitation and occupant density on shelter fidelity and assessed the consequences of shelter abandonment. We first quantified the area of leaf material required for a caterpillar to fully develop for two of the most common leaf-tiers that feed on white oak, Quercus alba. On average, Psilocorsis spp. caterpillars consumed 21.65 ± 0.67 cm(2) leaf material to complete development. We also measured the area of natural leaf ties found in a Maryland forest, to determine the distribution of resources available to caterpillars in situ. Of 158 natural leaf ties examined, 47% were too small to sustain an average Psilocorsis spp. caterpillar for the entirety of its development. We also manipulated caterpillar densities within experimental ties on potted trees to determine the effects of cohabitants on the likelihood of a caterpillar to leave its tie. We placed 1, 2, or 4 caterpillars in ties of a standard size and monitored the caterpillars twice daily to track their movement. In ties with more than one occupant, caterpillars showed a significantly greater propensity to leave their tie, and left sooner and at a faster rate than those in ties as single occupants. To understand the consequences of leaf tie abandonment, we observed caterpillars searching a tree for a site to build a shelter in the field. This is a risky behavior, as 17% of the caterpillars observed died while searching for a shelter site. Caterpillars that successfully built a shelter traveled 110 ± 20 cm and took 28 ± 7 min to find a suitable site to build a shelter. In conclusion, leaf-tying caterpillars must frequently abandon their leaf tie due to food limitation and interactions with other caterpillars, but this is a costly behavior.
Following a series of experiments in which six orangutans and one gorilla discriminated photographs of different animal species in a two-choice touch screen procedure, Vonk & MacDonald (2002) and Vonk & MacDonald (2004) concluded that orangutans, but not the gorilla, seemed to learn intermediate level category discriminations, such as primates versus non-primates, more rapidly than they learned concrete level discriminations, such as orangutans versus humans. In the current experiments, four of the same orangutans and the gorilla were presented with delayed matching-to-sample tasks in which they were rewarded for matching photos of different members of the same primate species; golden lion tamarins, Japanese macaques, and proboscis monkeys, or family; gibbons, lemurs (Experiment 1), and subsequently for matching photos of different species within the following classes: birds, reptiles, insects, mammals, and fish (Experiment 2). Members of both Great Ape species were rapidly able to match the photos at levels above chance. Orangutans matched images from both category levels spontaneously whereas the gorilla showed effects of learning to match intermediate level categories. The results show that biological knowledge is not necessary to form natural categories at both concrete and intermediate levels.
Rank-abundance (A) and rank-frequency (B) plots of Alnus-associated ectomycorrhizal fungal taxa sampled in mesh bags and root tips. 
A number of recent studies suggest that interspecific competition plays a key role in determining the structure of ectomycorrhizal (ECM) fungal communities. Despite this growing consensus, there has been limited study of ECM fungal community dynamics in abiotically stressful environments, which are often dominated by positive rather than antagonistic interactions. In this study, we examined the ECM fungal communities associated with the host genus Alnus, which live in soils high in both nitrate and acidity. The nature of ECM fungal species interactions (i.e., antagonistic, neutral, or positive) was assessed using taxon co-occurrence and DNA sequence abundance correlational analyses. ECM fungal communities were sampled from root tips or mesh in-growth bags in three monodominant A. rubra plots at a site in Oregon, USA and identified using Illumina-based amplification of the ITS1 gene region. We found a total of 175 ECM fungal taxa; 16 of which were closely related to known Alnus-associated ECM fungi. Contrary to previous studies of ECM fungal communities, taxon co-occurrence analyses on both the total and Alnus-associated ECM datasets indicated that the ECM fungal communities in this system were not structured by interspecific competition. Instead, the co-occurrence patterns were consistent with either random assembly or significant positive interactions. Pair-wise correlational analyses were also more consistent with neutral or positive interactions. Taken together, our results suggest that interspecific competition does not appear to determine the structure of all ECM fungal communities and that abiotic conditions may be important in determining the specific type of interaction occurring among ECM fungi.
Clinical characteristics of study population. 
Endoscopic features and treatments given. 
Results of multiple logistic regression analysis (forward: LR). 
Aim. Helicobacter pylori (H. pylori) infection is exceptionally rare in population from the north-eastern region of Peninsular Malaysia. This provides us an opportunity to contemplate the future without H. pylori in acute non-variceal upper gastrointestinal (GI) bleeding. Methods. All cases in the GI registry with GI bleeding between 2003 and 2006 were reviewed. Cases with confirmed non-variceal aetiology were analysed. Rockall score > 5 was considered high risk for bleeding and primary outcomes studied were in-hospital mortality, recurrent bleeding and need for surgery. Results. The incidence of non-variceal upper GI bleeding was 2.2/100,000 person-years. Peptic ulcer bleeding was the most common aetiology (1.8/100,000 person-years). In-hospital mortality (3.6%), recurrent bleeding (9.6%) and need for surgery (4.0%) were uncommon in this population with a largely low risk score (85.2% with score ≤5). Elderly were at greater risk for bleeding (mean 68.5 years, P = 0.01) especially in the presence of duodenal ulcers (P = 0.04) despite gastric ulcers being more common. NSAIDs, aspirin and co-morbidities were the main risk factors. Conclusions. The absence of H. pylori infection may not reduce the risk of peptic ulcer bleeding in the presence of risk factors especially offending drugs in the elderly.
msl-2 expression variation among somatic tissues. msl-2 expression estimates were retrieved for all four FlyAtlas replicates for each tissue. Tissues are sorted by their median log 2 msl-2 intensity among the FlyAtlas replicates.
In Drosophila melanogaster, the male-specific lethal (MSL) complex has been studied extensively for its role in upregulating male X-linked genes. Recent advances in high-throughput technologies have improved our understanding of how the MSL complex mediates dosage compensation through chromosome-wide chromatin modifications. Most studies, however, have focused on cell line models that cannot reflect any potential heterogeneity of in vivo dosage compensation. Comparisons between cell line and organismal gene-level dosage compensation upregulation suggest the possibility of variation in MSL complex activity among somatic tissues. We hypothesize the degree, up to but not exceeding 2-fold, to which the MSL complex upregulates male X-linked genes varies quantitatively by tissue type. In this model, MSL complex abundance acts as a rheostat to control the extent of upregulation. Using publicly available expression data, we provide evidence for our model in Drosophila somatic tissues. Specifically, we find X-to-autosome expression correlates with the tissue-specific expression of msl-2 which encodes an essential male-specific component of the MSL complex. This result suggests MSL complex mediated dosage compensation varies quantitatively by tissue type. Furthermore, this result has consequences for models explaining the organismal-scale molecular and evolutionary consequences of MSL-mediated dosage compensation.
Far more attention has been paid to the microbes in our feces than the microbes in our food. Research efforts dedicated to the microbes that we eat have historically been focused on a fairly narrow range of species, namely those which cause disease and those which are thought to confer some “probiotic” health benefit. Little is known about the effects of ingested microbial communities that are present in typical American diets, and even the basic questions of which microbes, how many of them, and how much they vary from diet to diet and meal to meal, have not been answered. We characterized the microbiota of three different dietary patterns in order to estimate: the average total amount of daily microbes ingested via food and beverages, and their composition in three daily meal plans representing three different dietary patterns. The three dietary patterns analyzed were: (1) the Average American (AMERICAN): focused on convenience foods, (2) USDA recommended (USDA): emphasizing fruits and vegetables, lean meat, dairy, and whole grains, and (3) Vegan (VEGAN): excluding all animal products. Meals were prepared in a home kitchen or purchased at restaurants and blended, followed by microbial analysis including aerobic, anaerobic, yeast and mold plate counts as well as 16S rRNA PCR survey analysis. Based on plate counts, the USDA meal plan had the highest total amount of microbes at 1.3 × 109 CFU per day, followed by the VEGAN meal plan and the AMERICAN meal plan at 6 × 106 and 1.4 × 106 CFU per day respectively. There was no significant difference in diversity among the three dietary patterns. Individual meals clustered based on taxonomic composition independent of dietary pattern. For example, meals that were abundant in Lactic Acid Bacteria were from all three dietary patterns. Some taxonomic groups were correlated with the nutritional content of the meals. Predictive metagenome analysis using PICRUSt indicated differences in some functional KEGG categories across the three dietary patterns and for meals clustered based on whether they were raw or cooked. Further studies are needed to determine the impact of ingested microbes on the intestinal microbiota, the extent of variation across foods, meals and diets, and the extent to which dietary microbes may impact human health. The answers to these questions will reveal whether dietary microbes, beyond probiotics taken as supplements—i.e., ingested with food—are important contributors to the composition, inter-individual variation, and function of our gut microbiota.
g θ curves for various θ parameters. As θ goes to zero, the g θ converge pointwise to g, which is 1 on the interior of the unit interval and 0 on the boundaries.
ANOVA p-values for various diversity statistics applied to the skin microbiome data of Oh et al. (2012). Rows are ordered by increasing mean rank across sites. 
In microbial ecology studies, the most commonly used ways of investigating alpha (within-sample) diversity are either to apply non-phylogenetic measures such as Simpson's index to Operational Taxonomic Unit (OTU) groupings, or to use classical phylogenetic diversity (PD), which is not abundance-weighted. Although alpha diversity measures that use abundance information in a phylogenetic framework do exist, they are not widely used within the microbial ecology community. The performance of abundance-weighted phylogenetic diversity measures compared to classical discrete measures has not been explored, and the behavior of these measures under rarefaction (sub-sampling) is not yet clear. In this paper we compare the ability of various alpha diversity measures to distinguish between different community states in the human microbiome for three different datasets. We also present and compare a novel one-parameter family of alpha diversity measures, BWPDθ, that interpolates between classical phylogenetic diversity (PD) and an abundance-weighted extension of PD. Additionally, we examine the sensitivity of these phylogenetic diversity measures to sampling, via computational experiments and by deriving a closed form solution for the expectation of phylogenetic quadratic entropy under re-sampling. On the three datasets, a phylogenetic measure always performed best, and two abundance-weighted phylogenetic diversity measures were the only measures ranking in the top four across all datasets. OTU-based measures, on the other hand, are less effective in distinguishing community types. In addition, abundance-weighted phylogenetic diversity measures are less sensitive to differing sampling intensity than their unweighted counterparts. Based on these results we encourage the use of abundance-weighted phylogenetic diversity measures, especially for cases such as microbial ecology where species delimitation is difficult.
Study Location. We simulated and compared capture-mark-recapture using grid-based and targeted sampling designs for brown bears (Ursus acrtos) inhabiting the Kenai Peninsula (center, with elevation shaded from low [light gray] to high [black]), south-central Alaska, USA (inset). The grid-based design used cells with an area of 49 km 2 , 81 km 2 (pictured in gray), and 121 km 2. The targeted design sampled places where important biological resources concentrated the target species (i.e., bear's attraction to anadromous streams; black lines).
Comparison of sampling configurations. Results describing the bias, precision and accuracy of abundance estimates generated from the grid and targeted sampling configurations, with trap placement stationary or moved between capture sessions (N = 42 for each configuration). 
Comparison of capture probabilities. Mean and standard deviation of capture probabilities for 5 sampling configurations. We used a t-test to compare capture probabilities. 
Comparison of recapture probabilities. Mean and standard deviation of recapture probabilities for 5 sampling configurations. We used a t-test to compare recapture probabilities. 
Wildlife biologists often use grid-based designs to sample animals and generate abundance estimates. Although sampling in grids is theoretically sound, in application, the method can be logistically difficult and expensive when sampling elusive species inhabiting extensive areas. These factors make it challenging to sample animals and meet the statistical assumption of all individuals having an equal probability of capture. Violating this assumption biases results. Does an alternative exist? Perhaps by sampling only where resources attract animals (i.e., targeted sampling), it would provide accurate abundance estimates more efficiently and affordably. However, biases from this approach would also arise if individuals have an unequal probability of capture, especially if some failed to visit the sampling area. Since most biological programs are resource limited, and acquiring abundance data drives many conservation and management applications, it becomes imperative to identify economical and informative sampling designs. Therefore, we evaluated abundance estimates generated from grid and targeted sampling designs using simulations based on geographic positioning system (GPS) data from 42 Alaskan brown bears (Ursus arctos). Migratory salmon drew brown bears from the wider landscape, concentrating them at anadromous streams. This provided a scenario for testing the targeted approach. Grid and targeted sampling varied by trap amount, location (traps placed randomly, systematically or by expert opinion), and traps stationary or moved between capture sessions. We began by identifying when to sample, and if bears had equal probability of capture. We compared abundance estimates against seven criteria: bias, precision, accuracy, effort, plus encounter rates, and probabilities of capture and recapture. One grid (49 km(2) cells) and one targeted configuration provided the most accurate results. Both placed traps by expert opinion and moved traps between capture sessions, which raised capture probabilities. The grid design was least biased (-10.5%), but imprecise (CV 21.2%), and used most effort (16,100 trap-nights). The targeted configuration was more biased (-17.3%), but most precise (CV 12.3%), with least effort (7,000 trap-nights). Targeted sampling generated encounter rates four times higher, and capture and recapture probabilities 11% and 60% higher than grid sampling, in a sampling frame 88% smaller. Bears had unequal probability of capture with both sampling designs, partly because some bears never had traps available to sample them. Hence, grid and targeted sampling generated abundance indices, not estimates. Overall, targeted sampling provided the most accurate and affordable design to index abundance. Targeted sampling may offer an alternative method to index the abundance of other species inhabiting expansive and inaccessible landscapes elsewhere, provided their attraction to resource concentrations.
Large scale surveys in mammalian tissue culture cells suggest that the protein expressed at the median abundance is present at 8,000 - 16,000 molecules per cell and that differences in mRNA expression between genes explain only 10-40% of the differences in protein levels. We find, however, that these surveys have significantly underestimated protein abundances and the relative importance of transcription. Using individual measurements for 61 housekeeping proteins to rescale whole proteome data from Schwanhausser et al., we find that the median protein detected is expressed at 170,000 molecules per cell and that our corrected protein abundance estimates show a higher correlation with mRNA abundances than do the uncorrected protein data. In addition, we estimated the impact of further errors in mRNA and protein abundances, showing that mRNA levels explain at least 56% of the differences in protein abundance for the genes detected by Schwanhausser et al., though because one major source of error could not be estimated the true percent contribution could be higher. We also employed a second, independent strategy to determine the contribution of mRNA levels to protein expression. We show that the variance in translation rates directly measured by ribosome profiling is only 12% of that inferred by Schwanhausser et al. and that the measured and inferred translation rates correlate only poorly (R2=0.13). Based on this, our second strategy suggests that mRNA levels explain ~81% of the variance in protein levels. We also determined the percent contributions of transcription, RNA degradation, translation and protein degradation to the variance in protein abundances using both of our strategies. While the magnitudes of the two estimates vary, they both suggest that transcription plays a more important role than the earlier studies implied and translation a much smaller role.
Phylogenetic relationships of the Plasmodium cyt-b lineages found in Aburria jacutinga from two captive populations.
Phylogenetic relationships of the Haemoproteus cyt-b lineages found in Aburria jacutinga from two captive populations.
Infectious diseases can cause deleterious effects on bird species, leading to population decline and extinction. Haemosporidia can be recognized by their negative effects on host fitness, including reproductive success and immune responses. In captivity, outbreaks of haemosporidian infection have been observed in birds in zoos and aviaries. The endemic Brazilian Atlantic rainforest species Aburria jacutinga is one of the most endangered species in the Cracidae family, and wild populations of this species are currently found mainly in conservation areas in only two Brazilian states. In this study, we aimed to evaluate the effects of avian haemosporidia on hematological and biochemical parameters in two captive populations of A. jacutinga. Forty-two animals were assessed, and the haemosporidian prevalence was similar for males and females. The occurrence of haemosporidian infection in captive A. jacutinga observed in this study was similar to results found in other captive and wild birds in Brazil. We found three different lineages of haemosporidia. Two lineages were identified as Plasmodium sp., one of which was previously detected in Europe and Asia, and the other is a new lineage closely related to P. gallinaceum. A new third lineage was identified as Haemoproteus sp. We found no significant differences in hematological and biochemical values between infected and non-infected birds, and the haemosporidian lineage did not seem to have an impact on the clinical and physiological parameters of A. jacutinga. This is the first report on an evaluation of natural haemosporidian infections diagnosed by microscopic and molecular methods in A. jacutinga by hematology, blood biochemistry, and serum protein values. Determining physiological parameters, occurrence and an estimation of the impact of haemosporidia in endangered avian species may contribute to the management of species rehabilitation and conservation.
Osmotic pressure ψ o and NaCl concentration based on the electrical conductivity used in Reichman, Bellairs & Mulligan (2006). Electrical conductivity (dS m −1 ) a NaCl concentration (g kg −1 H 2 O) b Osmotic pressure ψ o (MPa) 
Germination results. (A) Germinability G and (B) time required for maximum germination ¯ t of A. harpophylla in relation to the water potential ψ. Error bars indicate the standard deviation across 3 replicates of 50 seeds.
Initial soil water conditions play a critical role when seeding is the primary approach to revegetate post-mining areas. In some semi-arid climates, such as the Brigalow Belt Bioregion in eastern Australia, extensive areas are affected by open-cut mining. Together with erratic rainfall patterns and clayey soils, the Brigalow Belt denotes a unique biome which is representative of other water-limited ecosystems worldwide. Apart from other environmental stressors, germination is governed by the water potential of the surrounding soil material. While previous studies have confirmed the high tolerance of Brigalow (Acacia harpophylla) seeds to a broad range of temperature and salinity, the question of how soil water potential triggers seed germination remains. In this study, we used three replicates of 50 seeds of Brigalow to investigate germination in relation to water potential as an environmental stressor. Solutions of Polyethylene Glycol (PEG 6000) were applied to expose seeds to nine osmotic water potentials ranging from soil water saturation (0 MPa) and field capacity (-.01 to -.03 MPa) to the permanent wilting point (-1.5 MPa). We measured germinability (number of germinated seeds relative to total number of seeds per lot) and mean germination time (mean time required for maximum germination of a seed lot) to quantify germination. Based on the empirical data of the germination we estimated the parameters of the hydrotime model which simulates timing and success of seed emergence. Our findings indicate that Brigalow seeds are remarkably tolerant to water stress, with germination being observed at a water potential as low as -1.5 MPa. Likewise, the average base water potential of a seed population (hydrotime model) was very low and ranged between -1.533 and -1.451 MPa. In general, Brigalow seeds germinate opportunistically over a broad range of abiotic conditions related to temperature, salinity, and water availability. Direct seeding and germination of native plants on post-mining land may be an effective and economically viable solution in order to re-establish plant communities. However, due to their capacity to reproduce asexually, alternative rehabilitation approaches such as transplantation of whole soil-root compartments may become attractive for restoration ecologists to achieve safe, stable, and non-polluting ecosystems.
Frequency and severity of academic stressors. 
Frequency and severity of psychosocial stressors. 
Introduction. Medicine is one of the most stressful fields of education because of its highly demanding professional and academic requirements. Psychological stress, anxiety, depression and sleep disturbances are highly prevalent in medical students. Methods. This cross-sectional study was undertaken at the Combined Military Hospital Lahore Medical College and the Institute of Dentistry in Lahore (CMH LMC), Pakistan. Students enrolled in all yearly courses for the Bachelor of Medicine and Bachelor of Surgery (MBBS) degree were included. The questionnaire consisted of four sections: (1) demographics (2) a table listing 34 potential stressors, (3) the 14-item Perceived Stress Scale (PSS-14), and (4) the Pittsburgh Quality of Sleep Index (PSQI). Logistic regression was run to identify associations between group of stressors, gender, year of study, student’s background, stress and quality of sleep. Results. Total response rate was 93.9% (263/280 respondents returned the questionnaire). The mean (SD) PSS-14 score was 30 (6.97). Logistic regression analysis showed that cases of high-level stress were associated with year of study and academic-related stressors only. Univariate analysis identified 157 cases with high stress levels (59.7%). The mean (SD) PSQI score was 8.1 (3.12). According to PSQI score, 203/263 respondents (77%) were poor sleepers. Logistic regression showed that mean PSS-14 score was a significant predictor of PSQI score (OR 1.99, P < 0.05). Conclusion. We found a very high prevalence of academic stress and poor sleep quality among medical students. Many medical students reported using sedatives more than once a week. Academic stressors contributed significantly to stress and sleep disorders in medical students.
Validation of the radiotracer method. The relationship between urea efflux rate measured by [ 14 C]urea radiotracer appearance ("hot") versus urea efflux rate measured by chemical assay of urea appearance ("cold") in the perfused head preparation of the dogfish shark (N = 6).
Urea and analogue efflux rates in the perfused head preparation of the dogfish shark. Open bars represent urea efflux rates; shaded bars represent analogue efflux rates. The treatment (perfusion saline) is noted below the bars: Control saline 1-350 mmol L −1 urea (N = 12); Control saline 2-175 mmol L −1 urea + 175 mmol L −1 mannitol (N = 5); 175 mmol L −1 urea + 175 mmol L −1 [ 14 C]thiourea (N = 6); 175 mmol L −1 urea + 175 mmol L −1 [ 14 C]acetamide (N = 6). Means ± 1 SEM. Means bearing the same letters are not significantly different (P < 0.05).
Urea concentrations in plasma or perfusate versus those measured simultaneously in gill tissue. Black bars represent concentrations in the plasma or perfusate; open bars represent concentrations in the gill tissue. In (A), concentrations are expressed on a per kg tissue or plasma/perfusate basis; in (B) concentrations are expressed on a per kg tissue H 2 O or plasma/perfusate H 2 O basis. The treatment is noted below the bars: In vivo (N = 9); Control saline 1-350 mmol L −1 urea (N = 10); Control saline 2-175 mmol L −1 urea + 175 mmol L −1 mannitol (N = 5); 175 mmol L −1 urea + 175 mmol L −1 [ 14 C]thiourea (N = 6); 175 mmol L −1 urea + 175 mmol L −1 [ 14 C]acetamide (N = 6). Means ± 1 SEM. Plasma/perfusate means bearing the same capital letters are not significantly different (P < 0.05); gill tissue means bearing the same lower case letters are not significantly different (P < 0.05); asterisks indicate gill tissue mean significantly different (P < 0.05) from corresponding plasma/perfusate mean.
Analogue concentrations in the perfusate versus those measured simultaneously in gill tissue. Black bars represent concentrations in the perfusate; open bars represent concentrations in the gill tissue. In (A), concentrations are expressed on a per kg tissue or perfusate basis; in (B) concentrations are expressed on a per kg tissue H 2 O or perfusate H 2 O basis. The treatment (perfusion saline) is noted below the bars: 175 mmol L −1 urea + 175 mmol L −1 [ 14 C]thiourea (N = 6); 175 mmol L −1 urea + 175 mmol L −1 [ 14 C]acetamide (N = 6). Means ± 1 SEM. Perfusate means bearing the same capital letters are not significantly different (P < 0.05); gill tissue means bearing the same lower case letters are not significantly different (P < 0.05); asterisks indicate gill tissue mean significantly different (P < 0.05) from corresponding perfusate mean.
The branchial mechanism of urea retention in elasmobranchs was investigated using an in vitro isolated-perfused head preparation, as well as in vivo samples, in the spiny dogfish shark. Both in vivo and in control saline perfusions containing 350 mmol L(-1) urea, calculated intracellular urea concentrations in gill epithelial cells were close to extracellular concentrations. Urea efflux to the external water fell only non-significantly, and calculated gill intracellular urea concentration did not change when perfusate urea concentration was reduced from 350 to 175 mmol L(-1) with osmotic compensation by 175 mmol L(-1) mannitol. However, when the urea analogues thiourea or acetamide were present in the perfusate at concentrations equimolar (175 mmol L(-1)) to those of urea (175 mmol L(-1)), urea efflux rates were increased 4-fold and 6.5-fold respectively, and calculated gill intracellular urea concentrations were depressed by about 55%. Analogue efflux rates were similar to urea efflux rates. Previous studies have argued that either the basolateral or apical membranes provided the limiting permeability barrier, and/or that a back-transporter on the basolateral membranes of gill cells is responsible for urea retention. The present results provide new evidence that the apical membrane is the limiting factor in maintaining gill urea impermeability, and raise the prospect that a urea back-transporter, which can be competitively inhibited by thiourea and acetamide, operates at the apical membrane.
Transmission electron micrographs of mature spermatozoon of Stephanostomum murielae in region I-IV. (A and B) longitudinal and cross-section sections of region I showing the anterior spermatozoon extremity; (C and D) consecutive cross-sections showing (C) the formation of the second axoneme, continuous layer of cortical microtubules and (D) both axonemes formed accompanied with discontinuous layer of cortical microtubules; (E-G) consecutive cross-sections of region II containing the external ornamentation of the plasma membrane and the first mitochondrion (F and G). Note the presence of spine-like body (F) and four attachment zones (arrowheads); (H-J): region III or transitional areas showing the posterior part of the first mitochondrion, the axonemes and a decreasing number of cortical microtubules; (K): proximal part of region IV showing the second mitochondrion. Scale bars: 0.3 µm. Ase, anterior spermatozoon extremity; Ax1, first axoneme; C2, centriole of the second axoneme; Cm, cortical microtubules; Eo, external ornamentation of the plasma membrane; G, granules of glycogen; M1, first mitochondrion; M2, second mitochondrion; Sb, spine-like body. 
Transmission electron micrographs of mature spermatozoon of Stephanostomum murielae in region IV and V. (A and B) Cross-section in (A) proximal area of region IV showing second mitochondrion and (B) simultaneous presence of the nucleus and the second mitochondrion; (C) cross-sections showing the second axoneme, the nucleus and few cortical microtubules; (D and E) cross-sections showing disorganization of the second axoneme (D) and a nucleus in posterior tip of the spermatozoon (E); (F) granules of glycogen evidenced according to the Thiéry's test. Scale bars: 0.3 µm. Cm, cortical microtubules; D, doublets; G, granules of glycogen; M2, second mitochondrion; N, nucleus. 
Transmission electron micrographs of mature spermatozoon of Stephanostomoides tenuis in region III-V. (A) cross-section in distal part of region III showing both axonemes and few microtubules (about 2); (B) proximal part of region IV showing appearance of the second mitochondrion. (C) Two cross-sections showing second mitochondrion associated with microtubules; (D) cross-section exhibiting simultaneous presence of the nucleus and the second mitochondrion accompanied by few cortical microtubules; (E) longitudinal sections showing the second mitochondrion (not moniliform); (F) crosssections showing nucleus and few microtubules; (G) cross-sections in posterior tip of the spermatozoon exhibiting only the nucleus. Scale bars: 0.2 µm. Cm, cortical microtubules; M, microtubule; M2, second mitochondrion; N, nucleus. 
The mature spermatozoa of Stephanostomum murielae and Stephanostomoides tenuis are described by transmission electron microscopy. They present several ultrastructural features previously reported in other digeneans. Their spermatozoa possess two axonemes of different length showing the 9 + '1' trepaxonematan pattern, four attachment zones, two mitochondria (with an anterior moniliform one in S. murielae), a nucleus, two bundles of parallel cortical microtubules, external ornamentation of the plasma membrane, spine-like bodies and granules of glycogen. The main differences between the mature spermatozoon of S. murielae and S. tenuis are the maximum number of cortical microtubules, the morphology of the anterior spermatozoon extremity and the anterior mitochondrion. This study is the first concerning members of the family Acanthocolpidae. The main ultrastructural characteristics discussed are the morphology of the anterior and posterior spermatozoon extremities, antero-lateral electron dense material, external ornamentations, spine-like bodies and number and morphology of mitochondria. In addition, the phylogenetic significance of all these ultrastructural features is discussed and compared to molecular results in order to highlight the complex relationships in the Digenea.
Cytotoxicity of harmine in MCF-7 cells. MCF-7 cells were incubated with harmine at different concentrations (0 µM, 10 µM, 20 µM and 30 µM) and multiple time periods (24 h, 48 h, 72 h and 96 h). Cell metabolic activity was determined by MTT assay. Viability of vehicle-treated samples was set at 100%: 24 h, white bars; 48 h, black bars; 72 h, hatched bars; 96 h, dotted bars. Results are derived from two independent experiments performed in quadruplicate (mean ± SD). 
Effect of harmine on telomerase activity in MCF-7 cells. (A) MCF-7 cells were incubated with harmine at two concentrations (10 µM and 20 µM) for 24 h, 48 h, 72 h and 96 h. At the end of incubation, telomerase activity was evaluated by applying TRAP assay; the TRAP products were then separated on a 12% PAGE gel and their intensity (all bands) was quantified by using ImageJ software and values were plotted in (B): ctr, vehicle control, black bar; cells treated with 10 µM of harmine, white bars; cells treated with 20 µM of harmine, dotted bars. Results derived from two independent experiments (mean ± SD). p values indicate the significant changes in relative telomerase activity for the sample treated with harmine with respect to the vehicle treated controls. Unpaired t test: * p < 0.05; * * p ≤ 0.01. 
Harmine induces a general DNA damage response byover-expressing p53/p21 and γ H 2 AX. (A) MCF-7 cells were incubated with harmine at 20 µM for 24 h, 48 h and 96 h, then 25 µg of total protein extracted from cells after treatment of harmine or vehicle only was separated by PAGE and analyzed by Western blot. (B) changes in protein level after the treatment were calculated with respect to vehicle controls (100%): p21, black bars; p53, white bars; γ H 2 AX, grey bars; c-Myc, hatched bars. 
The end replication problem, which occurs in normal somatic cells inducing replicative senescence, is solved in most cancer cells by activating telomerase. The activity of telomerase is highly associated with carcinogenesis which makes the enzyme an attractive biomarker in cancer diagnosis and treatment. The indole alkaloid harmine has multiple pharmacological properties including DNA intercalation which can lead to frame shift mutations. In this study, harmine was applied to human breast cancer MCF-7 cells. Its activity towards telomerase was analyzed by utilizing the telomeric repeat amplification protocol (TRAP). Our data indicate that harmine exhibits a pronounced cytotoxicity and induces an anti-proliferation state in MCF-7 cells which is accompanied by a significant inhibition of telomerase activity and an induction of an accelerated senescence phenotype by over-expressing elements of the p53/p21 pathway.
Throughput versus sensitivity. Speed (measured as the number of 100 nt query sequences processed per second) plotted versus sensitivity (expressed as the overall percentage of mapped pairs). Data for 10 million 100 nt paired-end reads from the YanHuang genome. Workstation hardware: 12 CPU cores (24 threads of execution), one NVidia K20c GPU. Results for unpaired reads are similar (Supplementary Figure S9).  
When computing alignments of DNA sequences to a large genome, a key element in achieving high processing throughput is to prioritize locations in the genome where high-scoring mappings might be expected. We formulated this task as a series of list-processing operations that can be efficiently performed on graphics processing unit (GPU) hardware.We followed this approach in implementing a read aligner called Arioc that uses GPU-based parallel sort and reduction techniques to identify high-priority locations where potential alignments may be found. We then carried out a read-by-read comparison of Arioc's reported alignments with the alignments found by several leading read aligners. With simulated reads, Arioc has comparable or better accuracy than the other read aligners we tested. With human sequencing reads, Arioc demonstrates significantly greater throughput than the other aligners we evaluated across a wide range of sensitivity settings. The Arioc software is available at It is released under a BSD open-source license.
Structural equation model of the six dimensions of psychological well-being and harmony in life via self-fulfilling group. All correlations (between different psychological well-being dimensions) and all paths (from the six dimensions of psychological well-being to harmony in life) and their standardized parameter estimates. Chi-square = .00; DF = 00; comparative fit index = 1.00; incremental fit index = 1.00 and normed fit index = 1.00. e = error. Red standardized parameter estimates of regression weights are significant at the p < .001 level (n = 160). 
Summary of the results showing the differences between affective profiles in the 6 dimensions of psychological well-being and harmony in life. 
Background. An important outcome from the debate on whether wellness equals happiness, is the need of research focusing on how psychological well-being might influence humans’ ability to adapt to the changing environment and live in harmony. To get a detailed picture of the influence of positive and negative affect, the current study employed the affective profiles model in which individuals are categorised into groups based on either high positive and low negative affect (self-fulfilling); high positive and high negative affect (high affective); low positive and low negative affect (low affective); and high negative and low positive affect (self-destructive). The aims were to (1) investigate differences between affective profiles in psychological well-being and harmony and (2) how psychological well-being and its dimensions relate to harmony within the four affective profiles. Method. 500 participants (mean age = 34.14 years, SD. = ±12.75 years; 187 males and 313 females) were recruited online and required to answer three self-report measures: The Positive Affect and Negative Affect Schedule; The Scales of Psychological Well-Being (short version) and The Harmony in Life Scale. We conducted a Multivariate Analysis of Variance where the affective profiles and gender were the independent factors and psychological well-being composite score, its six dimensions as well as the harmony in life score were the dependent factors. In addition, we conducted four multi-group (i.e., the four affective profiles) moderation analyses with the psychological well-being dimensions as predictors and harmony in life as the dependent variables. Results. Individuals categorised as self-fulfilling, as compared to the other profiles, tended to score higher on the psychological well-being dimensions: positive relations, environmental mastery, self-acceptance, autonomy, personal growth, and purpose in life. In addition, 47% to 66% of the variance of the harmony in life was explained by the dimensions of psychological well-being within the four affective profiles. Specifically, harmony in life was significantly predicted by environmental mastery and self-acceptance across all affective profiles. However, for the low affective group high purpose in life predicted low levels of harmony in life. Conclusions. The results demonstrated that affective profiles systematically relate to psychological well-being and harmony in life. Notably, individuals categorised as self-fulfilling tended to report higher levels of both psychological well-being and harmony in life when compared with the other profiles. Meanwhile individuals in the self-destructive group reported the lowest levels of psychological well-being and harmony when compared with the three other profiles. It is proposed that self-acceptance and environmental acceptance might enable individuals to go from self-destructive to a self-fulfilling state that also involves harmony in life.
Demographics and Awareness.
Personal and Family History of Cancer.
Sharing of BRCA Results.
Background. Inherited BRCA gene mutations convey a high risk for breast and ovarian cancer, but current guidelines limit BRCA mutation testing to women with early-onset cancer and relatives of mutation-positive cases. Benefits and risks of providing this information directly to consumers are unknown. Methods. To assess and quantify emotional and behavioral reactions of consumers to their 23andMe Personal Genome Service® report of three BRCA mutations that are common in Ashkenazi Jews, we invited all 136 BRCA1 and BRCA2 mutation-positive individuals in the 23andMe customer database who had chosen to view their BRCA reports to participate in this IRB-approved study. We also invited 160 mutation-negative customers who were matched for age, sex and ancestry. Semi-structured phone interviews were completed for 32 mutation carriers, 16 women and 16 men, and 31 non-carriers. Questions addressed personal and family history of cancer, decision and timing of viewing the BRCA report, recollection of the result, emotional responses, perception of personal cancer risk, information sharing, and actions taken or planned. Results. Eleven women and 14 men had received the unexpected result that they are carriers of a BRCA1 185delAG or 5382insC, or BRCA2 6174delT mutation. None of them reported extreme anxiety and four experienced moderate anxiety that was transitory. Remarkably, five women and six men described their response as neutral. Most carrier women sought medical advice and four underwent risk-reducing procedures after confirmatory mutation testing. Male carriers realized that their test results implied genetic risk for female relatives, and several of them felt considerably burdened by this fact. Sharing mutation information with family members led to screening of at least 30 relatives and identification of 13 additional carriers. Non-carriers did not report inappropriate actions, such as foregoing cancer screening. All but one of the 32 mutation-positive participants appreciated learning their BRCA mutation status. Conclusions. Direct access to BRCA mutation tests, considered a model for high-risk actionable genetic tests of proven clinical utility, provided clear benefits to participants. The unexpected information demonstrated a cascade effect as relatives of newly identified carriers also sought testing and more mutation carriers were identified. Given the absence of evidence for serious emotional distress or inappropriate actions in this subset of mutation-positive customers who agreed to be interviewed for this study, broader screening of Ashkenazi Jewish women for these three BRCA mutations should be considered.
Optimised locations of farmers' markets to reach 25% of total population, M¯ aori population and deprived groups.
Background. Evidence suggests that improved locational access to farmers’ markets increases fruit and vegetable (FV) consumption, particularly for low-income groups. Therefore, we modelled potential alternative distributions of farmers’ markets in one country (New Zealand) to explore the potential impact for deprived populations and an indigenous population (Māori). Methods. Data were collected on current farmers’ markets (n = 48), population distributions, area deprivation, and roads. Geographic analyses were performed to optimize market locations for the most deprived populations. Results. We found that, currently, farmers’ markets provided fairly poor access for the total population: 7% within 12.5 km (15 min driving time); 5% within 5 km; and 3% within 2 km. Modelling the optimal distribution of the 48 markets substantially improved access for the most deprived groups: 9% (vs 2% currently) within 12.5 km; 5% (vs 1%) within 5 km; and 3% (vs 1%) within 2 km. Access for Māori also improved: 22% (vs 7%) within 12.5 km; 12% (vs 4%) within 5 km; and 6% (vs 2%) within 2 km. Smaller pro-equity results arose from optimising the locations of the 18 least pro-equity markets or adding 10 new markets. Conclusion. These results highlight the potential for improving farmers’ market locations to increase accessibility for groups with low FV consumption. Given that such markets are easily established and relocated, local governments could consider these results to inform decisions, including subsidies for using government land and facilities. Such results can also inform central governments planning around voucher schemes for such markets and exempting them from taxes (e.g., VAT/GST).
Human protein kinases play fundamental roles mediating the majority of signal transduction pathways in eukaryotic cells as well as a multitude of other processes involved in metabolism, cell-cycle regulation, cellular shape, motility, differentiation and apoptosis. The human protein kinome contains 518 members. Most studies that focus on the human kinome require, at some point, the visualization of large amounts of data. The visualization of such data within the framework of a phylogenetic tree may help identify key relationships between different protein kinases in view of their evolutionary distance and the information used to annotate the kinome tree. For example, studies that focus on the promiscuity of kinase inhibitors can benefit from the annotations to depict binding affinities across kinase groups. Images involving the mapping of information into the kinome tree are common. However, producing such figures manually can be a long arduous process prone to errors. To circumvent this issue, we have developed a web-based tool called Kinome Render (KR) that produces customized annotations on the human kinome tree. KR allows the creation and automatic overlay of customizable text or shape-based annotations of different sizes and colors on the human kinome tree. The web interface can be accessed at: A stand-alone version is also available and can be run locally.
Dopamine receptor inhibitors prevent the apomorphine/MCH enhancement of firing. Enhancement of NAshell firing by MCH (1 µM) + apomorphine (3 µM) was prevented by pre-exposure to (A) the DA1 receptor blocker SCH23390 (SCH, 1 µM), (B) the DA2 receptor blocker raclopride (Racl, 1 µM), (C) the D1/D2 blocker α-flupenthixol (flupen, 2 µM), or (D) the MCH receptor blocker TPI (2 µM).
ANOVA for dopamine receptor mediation of apomorphine/MCH enhancement of firing. Significant effect (*P < 0.05) of MCH + apomorphine (3 µM) versus 3 µM apomorphine alone or versus MCH + 3 µM apomorphine after pre-exposure to antagonists for DA1 (SCH), DA2 (Racl), DA1/DA2 (flupen) or MCH (TPI) receptors.
(A, B) (A) Example traces and (B) grouped data across time showing that 1 µM MCH (MCH) and 3 µM apomorphine (Apo3) interact to enhance firing in NAshell neurons in vitro, while 3 µM apomorphine had no effect alone. (C) 10 µM apomorphine (Apo10) was sufficient to enhance NAshell firing. (D) 1 µM MCH in combination with 1 µM apomorphine did not increase firing. (E) 1 µM MCH in combination with 10 µM apomorphine significantly increased firing. (F) 3 µM apomorphine in combination with 1 µM MCH significantly enhanced firing under conditions where AMPA and GABAA receptors were not blocked. (G) 3 µM apomorphine + 1 µM MCH did not increase firing when the PKA inhibitor peptide PKI (20 µM) was included in the intracellular pipette.
Enhancement of NAshell firing by MCH (1 µM) + apomorphine (3 µM) was prevented by pre-exposure to (A) the DA1 receptor blocker SCH23390 (SCH, 1 µM), (B) the DA2 receptor blocker raclopride (Racl, 1 µM), (C) the D1/D2 blocker α-flupenthixol (flupen, 2 µM), or (D) the MCH receptor blocker TPI (2 µM).
Significant effect (*P < 0.05) of MCH +apomorphine (3 µM) versus 3 µM apomorphine alone or versus MCH + 3 µM apomorphine after pre-exposure to antagonists for DA1 (SCH), DA2 (Racl), DA1/DA2 (flupen) or MCH (TPI) receptors.
The MCH and dopamine receptor systems have been shown to modulate a number of behaviors related to reward processing, addiction, and neuropsychiatric conditions such as schizophrenia and depression. In addition, MCH and dopamine receptors can interact in a positive manner, for example in the expression of cocaine self-administration. A recent report (Chung et al., 2011a) showed that the DA1/DA2 dopamine receptor activator apomorphine suppresses pre-pulse inhibition, a preclinical model for some aspects of schizophrenia. Importantly, MCH can enhance the effects of lower doses of apomorphine, suggesting that co-modulation of dopamine and MCH receptors might alleviate some symptoms of schizophrenia with a lower dose of dopamine receptor modulator and thus fewer potential side effects. Here, we investigated whether MCH and apomorphine could enhance action potential firing in vitro in the nucleus accumbens shell (NAshell), a region which has previously been shown to mediate some behavioral effects of MCH. Using whole-cell patch-clamp electrophysiology, we found that MCH, which has no effect on firing on its own, was able to increase NAshell firing when combined with a subthreshold dose of apomorphine. Further, this MCH/apomorphine increase in firing was prevented by an antagonist of either a DA1 or a DA2 receptor, suggesting that apomorphine acts through both receptor types to enhance NAshell firing. The MCH/apomorphine-mediated firing increase was also prevented by an MCH receptor antagonist or a PKA inhibitor. Taken together, our results suggest that MCH can interact with lower doses of apomorphine to enhance NAshell firing, and thus that MCH and apomorphine might interact in vivo within the NAshell to suppress pre-pulse inhibition.
Scheme for cellulose-chip-based fluorescent antibody probing of cohesin-dockerin interactions. Cohesins of interest (Coh1, Coh2, and Coh3) fused to cellulose-binding modules (CBMs) are bound to the cellulose-coated chip. A dockerin of interest (Doc), fused to the xylanase XynT6, interacts only with specific cohesins (Coh2, in this case). Cohesin-dockerin interaction is measured by the co-localization of Cy5-conjugated anti-CBM antibodies and Cy3-conjugated anti-XynT6 antibodies. Cy5 fluorescence represents general protein attachment to the chip, whereas Cy3 fluorescence represents specific, dockerin-induced interaction with the cohesin partner. 
Example of a comprehensive cohesin library screening, using a type-II X-dockerin dyad as a probe. Fluorescent scans of cellulose-coated slides containing the cohesin library interacted with XDoc-Xyn10 (an X-dockerin modular dyad derived from an A. cellulolyticus GH10 xylanase). Scans show Cy5 anti-CBM signal (A) and Cy3 anti-xylanase signal (B). The boxes are labeled showing each A. cellulolyticus CBM-Coh library member (from the 3rd cohesin of scaffoldin A, designated A3 through the cohesin from scaffoldin P). Each box contains five four-fold serial dilutions of CBM-Coh, blotted vertically in quadruplicate. Two control boxes (Ct and Rf) were included on each slide to check for cross-species binding, and were labeled with CBM-Coh from C. thermocellum and R. flavefaciens, respectively. The CBM box contains only a CBM and serves as a negative control. To the right of each box, a rectangle indicating the serial dilutions of the CBM-Xyn positive control is shown. The bottom spot of the CBM-Xyn rectangle is a duplicate of the highest concentration. 
Representative histogram showing cohesin library screen results. Normalized interaction intensities between the A. cellulolyticus CBM-Coh library and XDoc-Xyn10. CBM-Cohs A3 though P are shown along with negative controls (CBM alone, C. thermocellum CBM-CohA2, Ruminococcus flavefaciens CBM-CohE), and the positive control (Xyn-CBM). CohF1 was found to have the highest experimental interaction intensity and was normalized to an interaction value of one. The dashed line indicates the threshold for positive interaction, determined by the CBM negative control. 
Architectural models of A. cellulolyticus cellulosome systems. (A) Cell-bound cellulosome systems. (B) Cell-free cellulosome systems. Confirmed cohesin-dockerin interactions (those interactions ∼2x above the background cutoff threshold) are indicated in the connectivity scheme. Color coding (yellow, green and red) of the modules, brackets, and arrows reflect the respective affinity profiles enumerated in the figure. Black circles, white circles, and black/white circles indicate modules studied in this work, in previous works, and in both this work and previous works, respectively. 
Cellulosic waste represents a significant and underutilized carbon source for the biofuel industry. Owing to the recalcitrance of crystalline cellulose to enzymatic degradation, it is necessary to design economical methods of liberating the fermentable sugars required for bioethanol production. One route towards unlocking the potential of cellulosic waste lies in a highly complex class of molecular machines, the cellulosomes. Secreted mainly by anaerobic bacteria, cellulosomes are structurally diverse, cell surface-bound protein assemblies that can contain dozens of catalytic components. The key feature of the cellulosome is its modularity, facilitated by the ultra-high affinity cohesin-dockerin interaction. Due to the enormous number of cohesin and dockerin modules found in a typical cellulolytic organism, a major bottleneck in understanding the biology of cellulosomics is the purification of each cohesin- and dockerin-containing component, prior to analyses of their interaction. As opposed to previous approaches, the present study utilized proteins contained in unpurified whole-cell extracts. This strategy was made possible due to an experimental design that allowed for the relevant proteins to be "purified" via targeted affinity interactions as a function of the binding assay. The approach thus represents a new strategy, appropriate for future medium- to high-throughput screening of whole genomes, to determine the interactions between cohesins and dockerins. We have selected the cellulosome of Acetivibrio cellulolyticus for this work due to its exceptionally complex cellulosome systems and intriguing diversity of its cellulosomal modular components. Containing 41 cohesins and 143 dockerins, A. cellulolyticus has one of the largest number of potential cohesin-dockerin interactions of any organism, and contains unusual and novel cellulosomal features. We have surveyed a representative library of cohesin and dockerin modules spanning the cellulosome's total cohesin and dockerin sequence diversity, emphasizing the testing of unusual and previously-unknown protein modules. The screen revealed several novel cell-bound cellulosome architectures, thus expanding on those previously known, as well as soluble cellulose systems that are not bound to the bacterial cell surface. This study sets the stage for screening the entire complement of cellulosomal components from A. cellulolyticus and other organisms with large cellulosome systems. The knowledge gained by such efforts brings us closer to understanding the exceptional catalytic abilities of cellulosomes and will allow the use of novel cellulosomal components in artificial assemblies and in enzyme cocktails for sustainable energy-related research programs.
Typical HPLC chromatogram of standard solution of NAC (0.1 mg mL −1 ) and released NAC from experimental PMMA resin containing 0.9 wt% of NAC. Identification of NAC was made based on the retention time of the NAC peaks registered for the standard solutions. 
Cytotoxicity of PMMA resin with or without NAC. Data are presented as the mean ± SD of three independent experiments performed in quintuplicates. (A) Effects of the extract of PMMA resin with or without NAC on the viability of HDPCs at day 3 and day 7. For each tested time period, values with different superscripts are significantly different from each other (One-way ANOVA, p < 0.05). (B) Effects of 0.54 mM NAC on the viability of HDPCs at day 3 and day 7. Values with different superscripts are significantly different from each other (One-way ANOVA, p < 0.05). NS, not significant between the control group and the experimental group. (C) Typical SEM pictures showing cell attachment and morphology on top of PMMA resins with or without NAC. The cells with round or collapsed appearances were observed in subgroups containing no or 0.15 wt.% NAC (arrows). 
Degree of conversion of PMMA resin with different concentrations of NAC at specific time after mixing. Results are presented as mean ± SD of three independent experiments. 
Mechanical properties of PMMA resin with or without NAC. Data are presented as the mean ± SD. Values with different superscripts are significantly different from each other (One-way ANOVA, p < 0.05). (A) The flexural strength (FS) of specimens in various subgroups after 24 h (n = 12). (B) Microhardness value (VHN) after 24 h (n = 15). (C) Surface roughness (Ra) of the specimens (n = 3). (D) The SEM image of the fractured surfaces for each subgroup. Pit-like internal defects can be observed for subgroup with NAC (arrows). 
Objectives. This study aimed to investigate the influences of N-acetyl cysteine (NAC) on cytotoxicity and mechanical properties of Poly-methylmethacrylate (PMMA) dental resins. Methods. Experimental PMMA resin was prepared by incorporating various concentrations of NAC (0, 0.15, 0.3, 0.6 and 0.9 wt.%). MTT assay was performed to investigate viability of human dental pulp cells after exposure to extract of PMMA resin with or without NAC. Cell adhesion on resin specimens was examined with scanning electron microscopy. Degree of conversion was studied with Fourier Transform Infrared Spectroscopy (FTIR). Flexural strength, microhardness and surface roughness was evaluated using a universal testing machine, microhardness tester and optical profilometer, respectively. Results. Incorporation of NAC into PMMA resin significantly reduced its cytotoxicity and enhanced cell adhesion on its surface. NAC induced negative influences on the mechanical and physical properties of PMMA resin in a dose-dependent manner. The degree of conversion for all experimental PMMA resins reached as high as 72% after 24 h of polymerization. All the tested properties were maintained when the concentration of incorporated NAC was 0.15 wt.%. Conclusion. The addition of 0.15 wt.% NAC remarkably improved biocompatibility of PMMA resin without exerting significant negative influence on its mechanical and physical properties.
Crystal violet (CV) stained capsules (grey ovals) in xenic A. minutissimum biofilm (scale bar: 20 µm). Micrograph depicts 11 days old culture and is a merge of the chlorophyll fluorescence channel (red; indicating diatom cells) and the bright-field image (grey). Some mature capsules are marked with arrows. Bright spots within diatom cells are lipid bodies. Bacteria are visible as light and dark speckles around and in between the diatom cells. 
Comparison of microstructures on A. minutissimum cell surfaces in a xenic biofilm. (A) Capsule material is sometimes stretched between cells and/or towards the substrate (arrows; scale bar: 2 µm). Culture was 11 days old at the time of fixation for SEM. Asterisks denote magnified areas B and C. (B) Non-encapsulated cells possess a fibrillar mesh of varying degrees of density. Frustule pores are only partially covered and in some cases, fibrils stick out from the frustule (scale bar: 1 µm). (C) Encapsulated cells are completely covered with a material of slightly granular structure, but lacking clearly discernible features (scale bar: 1 µm). 
Scanning electron micrographs of terminal parts of A. minutissimum cells at potentially different encapsulation stages within xenic biofilms (scale bars: 1 µm). Fibrillar meshes (A) may form capsule material (C) by denser growth and cross-linking of fibrils (B). Depicted samples were taken from stationary, 11 to 31 days old cultures. 
Fibrillar microstructures (arrow pairs) within capsule material of A. minutissimum cells in xenic biofilm are revealed by mechanical stress (scale bars = 1 µm). Micrographs depict samples from an 11 days old culture. (A) Tip of a partially encapsulated cell. Fibrillar substructures are continuous throughout the capsule material. (B) Capsule material stretched between cells frays into fibrils. 
Achnanthidium minutissimum is a benthic freshwater diatom that forms biofilms on submerged surfaces in aquatic environments. Within these biofilms, A. minutissimum cells produce extracellular structures which facilitate substrate adhesion, such as stalks and capsules. Both consist of extracellular polymeric substance (EPS), but the microstructure and development stages of the capsules are so far unknown, despite a number of hypotheses about their function, including attachment and protection. We coupled scanning electron microscopy (SEM) to bright-field microscopy (BFM) and found that A. minutissimum capsules mostly possess an unstructured surface. However, capsule material that was mechanically stressed by being stretched between or around cells displayed fibrillar substructures. Fibrils were also found on the frustules of non-encapsulated cells, implicating that A. minutissimum capsules may develop from fibrillar precursors. Energy-dispersive X-ray (EDX) spectroscopy revealed that the capsule material do not contain silicon, distinguishing it from the frustule material. We furthermore show that bacteria preferentially attach to capsules, instead of non-encapsulated A. minutissimum cells, which supports the idea that capsules mediate diatom-bacteria interactions.
Ocean acidification threatens the foundation of tropical coral reefs. This study investigated three aspects of ocean acidification: (i) the rates at which perforate and imperforate coral-colony skeletons passively dissolve when pH is 7.8, which is predicted to occur globally by 2100, (ii) the rates of passive dissolution of corals with respect to coral-colony surface areas, and (iii) the comparative rates of a vertical reef-growth model, incorporating passive dissolution rates, and predicted sea-level rise. By 2100, when the ocean pH is expected to be 7.8, perforate Montipora coral skeletons will lose on average 15 kg CaCO3 m(-2) y(-1), which is approximately -10.5 mm of vertical reduction of reef framework per year. This rate of passive dissolution is higher than the average rate of reef growth over the last several millennia and suggests that reefs composed of perforate Montipora coral skeletons will have trouble keeping up with sea-level rise under ocean acidification. Reefs composed of primarily imperforate coral skeletons will not likely dissolve as rapidly, but our model shows they will also have trouble keeping up with sea-level rise by 2050.
Prostate cancer is a prevalent age-related disease in North America, accounting for about 15% of all diagnosed cancers. We have previously identified lithocholic acid (LCA) as a potential chemotherapeutic compound that selectively kills neuroblastoma cells while sparing normal human neurons. Now, we report that LCA inhibits the proliferation of androgen-dependent (AD) LNCaP prostate cancer cells and that LCA is the most potent bile acid with respect to inducing apoptosis in LNCaP as well as androgen-independent (AI) PC-3 cells, without killing RWPE-1 immortalized normal prostate epithelial cells. In LNCaP and PC-3 cells, LCA triggered the extrinsic pathway of apoptosis and cell death induced by LCA was partially dependent on the activation of caspase-8 and -3. Moreover, LCA increased cleavage of Bid and Bax, down-regulation of Bcl-2, permeabilization of the mitochondrial outer membrane and activation of caspase-9. The cytotoxic actions of LCA occurred despite the inability of this bile acid to enter the prostate cancer cells with about 98% of the nominal test concentrations present in the extracellular culture medium. With our findings, we provide evidence to support a mechanism of action underlying the broad anticancer activity of LCA in various human tissues.
Background. Significant physiological changes occur during pregnancy and lactation. Intrahepatic cholestasis of pregnancy (ICP) is a liver disease closely related to disruption of bile acid homeostasis. The objective of this study was to examine the regulation of bile acid synthesis and transport in normal pregnant and lactating rats. Materials and Methods. Livers from timed pregnant SD rats were collected on gestational days (GD) 10, 14 and 19, and postnatal days (PND) 1, 7, 14 and 21. Total bile acids were determined by the enzymatic method, total RNA was isolated and subjected to real time RT-PCR analysis. Liver protein was extracted for western-blot analysis. Results. Under physiological conditions hepatic bile acids were not elevated during pregnancy but increased during lactation in rats. Bile acid synthesis rate-limiting enzyme Cyp7a1 was unchanged on gestational days, but increased on PND14 and 21 at mRNA and protein levels. Expression of Cyp8b1, Cyp27a1 and Cyp7b1 was also higher during lactation. The mRNA levels of small heterodimer partner (SHP) and protein levels of farnesoid X receptor (FXR) were increased during pregnancy and lactation. Bile acid transporters Ntcp, Bsep, Mrp3 and Mrp4 were lower at gestation, but increased during lactation. Hepatic Oatp transporters were decreased during pregnancy and lactation. Conclusion. Hepatic bile acid homeostasis is maintained during normal pregnancy in rats, probably through the FXR-SHP regulation. The expression of bile acid synthesis genes and liver bile acid accumulation were increased during lactation, together with increased expression of bile acid efflux transporter Bsep, Mrp3 and Mrp4.
Top-cited authors
Torbjørn Rognes
  • University of Oslo
Ben Nichols
  • University of Glasgow
Tomáš Flouri
  • University College London
Juan Nunez-Iglesias
  • Monash University (Australia)
Stéfan Johann van der Walt
  • University of California, Berkeley