title: Probing the stability of the SpCas9-DNA complex after cleavage

abstract: CRISPR-Cas9 is a ribonucleoprotein complex that sequence-specifically binds and cleaves double-stranded DNA. Wildtype Cas9 as well as its nickase and cleavage-incompetent mutants have been used in various biological techniques due to their versatility and programmable specificity. Cas9 has been shown to bind very stably to DNA even after cleavage of the individual DNA strands, inhibiting further turnovers and considerably slowing down in-vivo repair processes. This poses an obstacle in genome editing applications. Here, we employed single-molecule magnetic tweezers to investigate the binding stability of different S. pyogenes Cas9 variants after cleavage by challenging them with supercoiling. We find that different release mechanisms occur depending on which DNA strand is cleaved. After non-target strand cleavage, supercoils are immediately but slowly released by swiveling of the non-target strand around the DNA with friction. Consequently, Cas9 and its non-target strand nicking mutant stay stably bound to the DNA for many hours even at elevated torsional stress. After target-strand cleavage, supercoils are only removed after the collapse of the R-loop. We identified several states with different stabilities of the R-loop. Most importantly, we find that the post-cleavage state of Cas9 exhibits a higher stability compared to the pre-cleavage state. This suggests that Cas9 has evolved to remain tightly bound to its cut target.


title: LRET-derived HADDOCK structural models describe the conformational heterogeneity required for processivity of the Mre11-Rad50 DNA damage repair complex

abstract: The Mre11-Rad50-Nbs1 protein complex is one of the first responders to DNA double strand breaks. Studies have shown that the catalytic activities of the evolutionarily conserved Mre11-Rad50 (MR) core complex depend on an ATP-dependent global conformational change that takes the macromolecule from an open, extended structure in the absence of ATP to a closed, globular structure when ATP is bound. We have previously identified an additional  partially open conformation using Luminescence Resonance Energy Transfer (LRET) experiments. Here, a combination of LRET and the molecular docking program HADDOCK was used to further investigate this partially open state and identify three conformations of ATP-bound MR in solution: closed, partially open, and open, which are in addition to the extended, apo conformation. These models are supported with mutagenesis and SAXS data that corroborate the presence of these three states and suggest a mechanism for the processivity of the MR complex along the DNA.

category: biophysics


title: Loss of BCL-3 sensitises colorectal cancer cells to DNA damage, revealing a role for BCL-3 in double strand break repair by homologous recombination

abstract:
ObjectiveThe proto-oncogene BCL-3 is upregulated in a subset of colorectal cancers (CRC) and increased expression of the gene correlates with poor patient prognosis. The aim is to investigate whether inhibiting BCL-3 can increase the response to DNA damage in CRC.

DesignThe function of BCL-3 in DNA damage response was studied in vitro using siRNA and CRISPR-Cas9 genome editing and in vivo using Bcl3-/- mice. DNA damage induced by {gamma}-irradiation and/or cisplatin was quantified using H2AX and RAD51 foci, repair pathways investigated using HR/NHEJ assays and treatment with the PARP inhibitor olaparib.

ResultSuppression of BCL-3 increases double strand break number and decreases homologous recombination in CRC cells, supported by reduced RAD51 foci number and increased sensitivity to PARP inhibition. Importantly, a similar phenotype is seen in Bcl3-/- mice, where the intestinal crypts of these mice exhibit sensitivity to DNA damage and a greater number of double strand breaks compared to wild type mice. Furthermore Apc.Krasmutant x Bcl3-/- mice exhibit increased DNA damage and reduced RAD51+ cells compared to their wild type counterparts when treated with cisplatin.

ConclusionThis work identifies BCL-3 as a regulator of the cellular response to DNA damage and suggests that elevated BCL-3 expression could increase resistance of tumour cells to DNA damaging agents including radiotherapy. These findings offer a rationale for targeting BCL-3 in CRC as an adjuvant to conventional therapies and suggest that BCL-3 expression in tumours could be a useful biomarker in stratification of rectal cancer patients for neo-adjuvant chemoradiotherapy.

category: cancer biology


title: Pyroptosis-associated molecular classification and prognostic signature in glioma

abstract: Integrative analysis was performed in the Chinese Glioma Genome Atlas and The Cancer Genome Atlas to describe the pyroptosis-associated molecular classification and prognostic signature in glioma. Pyroptosis-related genes were used for consensus clustering and to develop a prognostic signature. The immune statuses, molecular alterations and clinical features of differentially expressed genes were analyzed among different subclasses and risk groups. A lncRNA-miRNA-mRNA network was built, and drug sensitivity analysis was used to identify small molecular drugs for the identified genes. Glioma can be divided into two subclasses using 30 pyroptosis-related genes. Cluster 1 displayed high immune signatures and poor prognosis as well as high immune-related function scores. A prognostic signature based on 15 pyroptosis-related genes of the CGGA cohort can predict the overall survival of glioma and was well validated in the TCGA cohort. Cluster 1 had higher risk scores. The high-risk group had high immune cell and function scores and low DNA methylation of pyroptosis-related genes. The differences in pyroptosis-related gene mutations and somatic copy numbers were significant between the high-risk and low-risk groups. The ceRNA regulatory network uncovered the regulatory patterns of different risk groups in glioma. Nine pairs of target genes and drugs were identified. In vitro, CASP8 promotes the progression of glioma cells. Pyroptosis-related genes can reflect the molecular biological and clinical features of glioma subclasses. The established prognostic signature can predict prognosis and distinguish molecular alterations in glioma patients. Our comprehensive analyses provide valuable guidelines for improving glioma patient management and individualized therapy.


title: Comprehensive analysis of pyroptosis-associated in molecular classification, immunity and prognostic of glioma


title: A Codon Constrained Method for Both Eliminating and Creating Intragenic Bacterial Promoters

abstract: Future applications of synthetic biology will require refactored genetic sequences devoid of internal regulatory elements within coding sequences. These regulatory elements include cryptic and intragenic promoters which may constitute up to a third of predicted Escherichia coli promoters. Promoter activity is dependent on the structural interaction of core bases with a {sigma} factor. Rational engineering can be used to alter key promoter element nucleotides interacting with {sigma} factors and eliminate downstream transcriptional activity. In this paper, we present COdon Restrained Promoter SilEncing (CORPSE), a system for removing intragenic promoters. CORPSE exploits the DNA-{sigma} factor structural relationship to disrupt {sigma}70 promoters embedded within gene coding sequences, with a minimum of synonymous codon changes. Additionally, we present an inverted CORPSE system, iCORPSE, which can create highly active promoters within a gene sequence while not perturbing the function of the modified gene.

category: synthetic biology


title: Exploring the impact of terminators on transgene expression in Chlamydomonas reinhardtii with a synthetic biology approach

abstract: Chlamydomonas reinhardtii has many attractive features for use as a model organism for both fundamental studies and as a biotechnological platform. Nonetheless, despite the many molecular tools and resources that have been developed, there are challenges for its successful engineering, in particular to obtain reproducible and high levels of transgene expression. Here we describe a synthetic biology approach to screen several hundred independent transformants using standardised parts to explore different parameters that might affect transgene expression. We focused on terminators and, using a standardised workflow and quantitative outputs, tested 9 different elements representing three different size classes of native terminators to determine their ability to support high level expression of a GFP reporter gene. We found that the optimal size reflected the median size of element found in the C. reinhardtii genome. The behaviour of the terminator parts was similar with different promoters, in different host strains and with different transgenes. This approach is applicable to the systematic testing of other genetic elements, facilitating comparison to determine optimal transgene design.


title: Helical Ordering of Envelope Associated Proteins and Glycoproteins in Respiratory Syncytial Virus Filamentous Virions

abstract: Human respiratory syncytial virus (RSV) causes severe respiratory illness in children and the elderly. Treatments for RSV disease are however limited and efforts to produce an effective vaccine have so far been unsuccessful. Understanding RSV virion structure is an important prerequisite for developing interventions to treat or prevent infection but has been challenging because of the fragility of virions propagated in cell culture. Here we show, using cryogenic electron microscopy (cryoEM) and cryogenic electron tomography (cryoET) of RSV particles cultivated directly on transmission electron microscopy (TEM) grids, that there is extensive helical symmetry in RSV filamentous virions. We have calculated a 16 [A] resolution three-dimensional reconstruction of the viral envelope, targeting the matrix protein (M) that forms an endoskeleton below the viral membrane. These data define a helical lattice of M proteins, showing how M is oriented relative to the viral envelope and that helical ordering of viral glycoproteins that stud the viral envelope is coordinated by the M layer. Moreover, the helically ordered viral glycoproteins in RSV filamentous virions cluster in pairs, which may have implications for the conformation of fusion (F) glycoprotein epitopes that are the principal target for vaccine and monoclonal antibody development. We also report the presence, in authentic virus infections, of N-RNA rings packaged within RSV filamentous virions. Overall, the structural data obtained provides molecular insight into the organization of the virion and the mechanism of its assembly.

category: microbiology


title: Preexisting mycobacterial infection modulates Candida albicans-driven pyroptosis

abstract: Active tuberculosis patients are at high risk of co-infection with opportunistic fungal pathogen C. albicans. However, the molecular mechanisms that orchestrate pathogenesis of Mycobacterium tuberculosis (Mtb)-C. albicans co-infection remains elusive. In the current study, we utilise a mouse model to demonstrate that Mtb promotes macrophage environment conducive for C. albicans survival. Mtb-dependent PKC{zeta}-WNT signalling axis induces expression of an E3 ubiquitin ligase, COP1. A secondary infection of C. albicans in such Mtb-infected macrophages causes COP1 to mediate the proteasomal degradation of IRF9, a cardinal factor that we identified to arbitrate an inflammatory programmed cell death, pyroptosis. In vivo experiments mimicking a preexisting Mtb infection demonstrate that inhibition of pyroptosis in mice results in increased C. albicans burden and aberrant lung tissue architecture, leading to increased host mortality. Together, our study reveals the crucial role of pyroptosis regulation for manifesting a successful C. albicans-Mtb co-infection.


title: Rock traits drive complex microbial communities at the edge of life

abstract: Antarctic deserts are among the driest and coldest ecosystems of the planet; there, some microbes hang on to life under these extreme conditions inside porous rocks, forming the so-called endolithic communities. Yet, the contribution of distinct rock traits to support complex microbial assemblies remains poorly determined. Here, we combined an extensive Antarctic rock survey with rock microbiome sequencing and ecological networks, and found that contrasting combinations of microclimatic and rock traits such as thermal inertia, porosity, iron concentration and quartz cement can help explain the multiple complex and independent microbial assemblies found in Antarctic rocks. Our work highlights the pivotal role of rocky substrate heterogeneity in sustaining contrasting groups of microorganisms, which is essential to understand life at the edge on Earth, and for searching life on other rocky planets such as Mars.


title: Perturbation of transmembrane 6 superfamily member 2 expression alters lipid metabolism in a human liver cell line

abstract:
BackgroundNonalcoholic fatty liver disease (NAFLD) is caused by accumulation of excess lipids in hepatocytes. Genome wide association studies have identified strong association of NAFLD with non-synonymous E167K amino acid mutation in transmembrane 6 superfamily member 2 (TM6SF2) protein. The E167K mutation affects TM6SF2 stability and its carriers display increased hepatic lipids levels and lower serum triglycerides. While similar phenotype is evident in mice with TM6SF2 knockdown, effects of TM6SF2 on hepatic lipid metabolism is not completely understood.

MethodsHere, we overexpressed wild-type or E167K variant of TM6SF2 or knocked down TM6SF2 expression in lipid-treated Huh-7 cells and used biochemical assays, untargeted lipidomic analysis, RNAseq transcriptome analysis and high-throughput fluorescent imaging to determine changes in lipid metabolism.

ResultsBoth knockdown and E167K overexpression increased acylglyceride levels which was decreased by wild-type TM6SF2 overexpression. Further, mean intensity of individual lipid droplets was increased by E167K overexpression and knockdown while wild-type TM6SF2 had no effects. We also observed lipid chain remodeling for acylglycerides by TM6SF2 knockdown leading to a relative increase in species with shorter and more saturated side chains. RNA sequencing revealed differential expression of several lipid metabolizing genes, including genes belonging to AKR1 family and lipases, primarily in cells with TM6SF2 knockdown.

ConclusionTaken together, our data shows that overexpression of TM6SF2 gene or its loss-of-function changes hepatic lipid species composition and expression of lipid metabolizing genes. Further, overexpression of E167K variant and TM6SF2 knockdown similarly increased hepatic lipid accumulation and lipid droplets profile further confirming a loss-of-function effect for variant.

category: cell biology


title: Regulators of male and female sexual development critical for transmission of a malaria parasite

abstract: The transmission of malaria parasites from vertebrate host to mosquito vector requires a developmental switch in asexually dividing blood-stage parasites to sexual reproduction. In Plasmodium berghei the transcription factor AP2-G is required and sufficient for this switch, but how a particular sex is determined in a haploid parasite remains unknown. Using a global screen of barcoded mutants, we here identify ten genes essential for the formation of either male or female sexual forms and validate their importance for transmission. High-resolution single-cell transcriptomics of wild-type and mutant parasites portrays the developmental bifurcation and reveals a regulatory cascade of putative gene functions in determination and subsequent differentiation of each sex. A male-determining gene with a LOTUS/OST-HTH domain points towards unexpected conservation of molecular mechanisms of gametogenesis in animals and a distantly related eukaryotic parasite.

category: genomics


title: GRAMD1C promotes autophagy initiation and mitochondrial bioenergetics through regulation of ER-mitochondria cholesterol transport

abstract: During autophagy, cytosolic cargo is sequestered into double-membrane vesicles called autophagosomes. The origin and identity of the membranes that form the autophagosome remain to be fully characterized. Here, we investigated the role of cholesterol in starvation- induced autophagy and identify a role for the ER-localized cholesterol transport protein GRAMD1C in the regulation of autophagy and mitochondrial function. We demonstrate that cholesterol depletion leads to a rapid induction of autophagy, possibly caused by a corresponding increased abundance of curved autophagy membranes. We further show that GRAMD1C is a negative regulator of starvation-induced autophagy. Similar to its yeast orthologue, GRAMD1C is recruited to mitochondria through its GRAM domain. Additionally, we find that GRAMD1C depletion leads to increased mitochondrial cholesterol accumulation and mitochondrial oxidative phosphorylation. Finally, we demonstrate that expression of GRAM family genes is linked to clear cell renal carcinoma survival, highlighting the pathophysiological relevance of cholesterol transport proteins.

category: biochemistry


title: IL-13 promotes recovery from C. difficile infection

abstract: Clostridioides difficile infection (CDI) is the leading hospital-acquired infection in North America. We have previously discovered that antibiotic disruption of the gut microbiota decreases intestinal IL-33 and IL-25 and increases susceptibility to CDI. We further found that IL-33 promotes protection through type 2 Innate Lymphoid Cells (ILC2s), which produce IL-13. However, the contribution of IL-13 to disease has never been explored. We found that administration of IL-13 protected, and anti-IL-13 exacerbated CDI as measured by weight loss and clinical score, particularly during disease resolution. Additionally, concordant with IL-13 being important for M2 macrophage polarization, we saw a decrease in M2 macrophages (CD11B+CD64+CD206+) cells following neutralization of IL-13. We also observed monocyte accumulation as early as day three post-infection following IL-13 neutralization, suggesting IL-13 may be directly or indirectly important for their recruitment or transition into macrophages. Neutralization of the decoy receptor IL-13R2 resulted in protection from disease, likely through increased available endogenous IL-13. Our data highlight the protective role of IL-13 in promoting recovery from CDI and the association of poor responses with a dysregulated monocytemacrophage compartment. These results increase our understanding of type 2 immunity in CDI and may have implications for treating disease in patients.

category: immunology


title: Comparative proteomic analysis of vancomycin-sensitive and vancomycin-intermediate resistant S. aureus

abstract: The extensive use of vancomycin has led to the development of MRSA (methicillin-resistant Staphylococcus aureus) strains with varying degrees of resistance to vancomycin. Accumulation of surplus cell wall material, decreased cross-linking of peptidoglycan, and/ or other cell wall alterations has been put forward to explain the VISA phenotype. To our knowledge, the protein profiles of hVISA and VISA strains are rarely analyzed via quantitative comparative proteomics. In this study, we subjected subcellular fractions isolated from two isogenic S. aureus strains (vancomycin-intermediate resistant S. aureus) to proteomic analysis, using an integrated quantitative proteomic approach assisted by bioinformatic analysis. In total,128 up-regulated proteins (mainly AhpC, Adh, ArgF et al.) were identified,21 down-regulated proteins (mainly LtaS, SdrD, MsrR, MsrB, OatA et al.) were obtained. The largest group of differentially expressed proteins is composed of enzymic proteins associated with metabolic and catalytic activity, which accounts for 50% and 51% of the total proteins, respectively.Some proteins which take an indispensable part in the regulatory networks of S. aureus with vancomycin treatment are related to Cell wall metabolism ( MurA ), Cell adhesion ( SdrC, SdrD, ClfA, ClfB ), Proteolysis ( Atl, LytM, SceD ) and Pressure response ( MsrA, MsrB, AhpC ) process.In conclusion, our proteomic study revealed regulatory proteins associated with vancomycin resistance in S. aureus, and some of these proteins are involved in the regulation of cell metabolism and function, providing protential targets for further development of drug resistance strategies.


title: Microbial composition in larval water enhances Aedes aegypti development but reduces transmissibility of Zika virus

abstract:
Arthropod-borne viral (arboviral) pathogens comprise a significant global disease burden, and outbreaks are expected to increase as vectors expand. Surveillance and mitigation of arboviruses like Zika virus (ZIKV) require accurate estimates of transmissibility by vector mosquitoes. While numerous laboratory vector competence experiments show that Aedes spp. mosquitoes are competent ZIKV vectors, differences in experimental protocols prevent direct comparisons of relative transmissibility across studies. An understudied factor complicating these comparisons is differential environmental microbiota exposures, where most vector competence studies use mosquitoes reared in laboratory tap water, which does not represent the microbial complexity of environmental water where wild larvae develop. We simulated natural larval development by rearing Californian Aedes aegypti with microbes obtained from cemetery headstone water, a common larval habitat in California, compared to conventional laboratory tap water. Ae. aegypti larvae reared in environmental cemetery water pupated 3 days faster and at higher rates. Female adult mosquitoes reared in environmental water were less competent vectors of ZIKV compared to laboratory water-reared Ae. aegypti, as evidenced by significantly reduced infection and transmission rates for two 2015 ZIKV strains and in two Ae. aegypti colonies from California. Microbiome comparisons of laboratory- and environment-water reared mosquitoes as well as their rearing water showed significantly higher bacterial diversity in environment water; despite this pattern, corresponding differences in diversity were not consistently detected in adult mosquitoes reared in different water sources. We also detected more significant associations between the microbial composition of adult mosquitoes and whether they ingested a bloodmeal than larval water type. Together, these results highlight the role of transient microbes in the larval environment in modulating vector competence. Laboratory vector competence likely overestimates true transmissibility of arboviruses like ZIKV when conventional laboratory water is used for rearing.

ImportanceWe observed that Ae. aegypti mosquitoes reared in water from cemetery headstones instead of the laboratory tap exhibited a reduced capacity to become infected with and transmit Zika virus. Water from the environment contained more bacterial species than tap water, but these bacteria were not consistently detected in adult mosquitoes. Our results suggest that rearing mosquito larvae in water collected from local environments as opposed to laboratory tap water, as is conventional, provides a more realistic assessment of vector competence since it better recapitulates the natural environment in which larvae develop. Given that laboratory vector competence is used to define the species to target for control, use of environmental water to rear larvae could better approximate the microbial exposures of wild mosquitoes, lessening the potential for overestimating transmission risk.


title: uPIC-M: efficient and scalable preparation of clonal single mutant libraries for high-throughput protein biochemistry

abstract: New high-throughput biochemistry techniques complement selection-based approaches and provide quantitative kinetic and thermodynamic data for thousands of protein variants in parallel. With these advances, library generation rather than data collection has become rate limiting. Unlike pooled selection approaches, high-throughput biochemistry requires mutant libraries in which individual sequences are rationally designed, efficiently recovered, sequence-validated, and separated from one another, but current strategies are unable to produce these libraries at the needed scale and specificity at reasonable cost. Here, we present a scalable, rapid, and inexpensive approach for creating User-designed Physically Isolated Clonal-Mutant (uPIC-M) libraries that utilizes recent advances in oligo synthesis, high-throughput sample preparation, and next-generation sequencing. To demonstrate uPIC-M, we created a scanning mutant library of SpAP, a 541 amino acid alkaline phosphatase, and recovered 94% of desired mutants in a single iteration. uPIC-M uses commonly available equipment and freely downloadable custom software and can produce a 5000 mutant library at 1/3 the cost and 1/5 the time of traditional techniques.

category: molecular biology


title: Surface expressed Plasmodium circumsporozoite protein (CSP) modulates cellular flexibility and motility

abstract: Plasmodium falciparum circumsporozoite protein (CSP) is a critically required abundant surface protein of sporozoites and a major vaccine candidate. However, neither the structure nor the role of CSP in sporozoite motility is well understood. Our recent in vitro data, from single-molecule pulling experiments suggested a mechanically pliable structure for P. falciparum CSP. By engineering vegetative cells of the cellular slime-mold Dictyostelium discoideum with regulatable CSP surface expression, we report evidence for direct involvement of CSP towards conferring elastic properties and motility of the cells. With an increase in the surface-CSP levels by 5-8-fold, the Youngs moduli of the cells, observed through atomic force microscopy, decreased around 2-fold, with a concomitant increase in motility by about 2-fold. Interestingly, only full length CSP expression conferred maximal flexibility and motility, as opposed to repeat region alone or the flanking domains of CSP. The enhanced motility of the CSP-expressing cells was abrogated with anti-CSP antibodies as well as phospholipase cleavage of CSP, indicating specific contribution of CSP towards motility. Measurements of the Youngs moduli of Plasmodium berghei midgut (MG) and salivary gland (SG) sporozoites revealed an inverse correlation with CSP levels with a decrease from 1.1 kPa to 0.3 kPa as the CSP concentration doubled from MG to SG sporozoites. We hypothesize that high CSP level lowers the stiffness of sporozoites possibly through its pliable surface-coat, leading to cellular flexibility. These findings may explain a sporozoites developmental ability to enhance its CSP levels during transition from midgut to salivary glands to suit a migratory mode in the host, needed for successful hepatocyte invasion.


title: Role of polycomb repressive complex 2 in regulation of human transcription factor gene expression

abstract: The study described here was pursued with the idea that information about control of human gene expression might result from comparing the promoter regions of a highly expressed gene population with that of a related population that is weakly expressed. Systematic differences in the two promoter populations would be candidates for a role in causing the distinct expression levels observed. The results reported here were obtained with human genes encoding transcription factors (TF). Among TF genes, those with wide tissue transcription are strongly expressed while those expressed in one or only a few tissues are weakly transcribed in most cases. The promoter regions of the two populations were compared using the results of ChIP-seq experiments. Results demonstrated that the promoter region of broadly expressed TF genes is enriched in binding sites for POLR2A, a component of RNA polymerase II while promoters of tissue targeted genes are enriched in EZH2, a subunit of polycomb repressive complex 2 (PRC2). It was rare to observe promoters with binding sites for both POLR2A and EZH2 or for neither one. The findings are interpreted to indicate that strong expression of broadly expressed TF genes is due to the presence of RNA polymerase II at the promoter while weak expression of tissue targeted promoters results from the presence of PRC2. Finally, transcription factor families were compared in the proportion of broadly expressed and tissue targeted genes. The results demonstrated that most families possess both broadly expressed and tissue targeted members. For instance, this was the case with 16 of 20 TF families. The results are interpreted to indicate that while individual TFs such as EZH2 may be specific for broadly expressed or tissue targeted genes, this is not a property of most TF families. Most have both broadly expressed and tissue targeted members.


title: Predictive Functions of H3K27me3 and H4K20me3 in Primate Hippocampal Stem and Progenitor Cells

abstract: Epigenetic regulations play important roles in cell fate determination during neurogenesis, a process by which different types of neurons are generated from neural stem and progenitor cells (NSPCs). Although some epigenetic changes are part of developmental and aging processes, the role of tri-methylation on histone 3 lysine 27 (H3K27me3) and histone 4 lysine 20 (H4K20me3) in primate hippocampal NSPCs remains elusive. This task is best assessed within a context resembling the human brain. As more studies emerge, the baboon represents an excellent model of human central nervous system in addition to their genomic similarity. With a focus on H3K27me3 and H4K20me3, the overarching goal of this work is to reveal their respective epigenetic landscapes in NSPCs of non-human primate baboon hippocampus. We identified putative targets of H3K27me3 and H4K20me3 that suggests a protective mechanism by dual H3K27me3/H4K20me3-mediated repression of specific-lineage gene activation important for differentiation processes while controlling the progression of the cell cycle.


title: The role of artificial photo backgrounds of shelter dogs on pet profile clicking and the perception of sociability

abstract: With the increasing prevalence of technology, the internet is often the first step for potential pet owners searching for an adoptable dog. However, best practices for the online portrayal of shelter and foster dogs remain unclear. Different online photo backgrounds appearing on adoption websites for shelter dogs may impact adoption speed by influencing viewer interest. Online clicking behaviour on pet profiles and human-directed sociability, broadly defined, has been previously linked to increased adoption likelihood. Therefore, the objective of this study was to determine the relationship between photo backgrounds of shelter dogs and online clicking as well as perceived human-directed sociability. In a virtual experiment, 680 participants were asked to rank the sociability and friendliness of four different adoptable dogs on a scale from 0-10. The photo background of each dog was digitally altered and randomly assigned to four experimental background conditions: 1) outdoor, 2) home indoor, 3) in-kennel indoor, and 4) plain coloured. As a proxy for adoption interest, a link to the dogs adoption profile was presented on each slide and the clicking behaviour of participants on this link was recorded. Mixed logistic regression and poisson models revealed that background did not affect participants link-clicking behaviour (chisq = 3.55, df = 3, p = .314) nor perceptions of sociability (statistic = 6.19, df = 3, p = .103). Across all backgrounds, only 4.74% of presented slides culminated in participant link-clicking. Sociability scores also did not predict link clicking. Assessment of participant-related factors and dog ID revealed that link-clicking and sociability scores of photographs were influenced by differences between dogs themselves and unaffected by participants awareness of study hypotheses. We conclude that artificial background types did not affect participant responses. The results demonstrate the importance of empirical data in making marketing decisions in animal shelters. Understanding which aspects of online marketing materials impact viewer interest will provide guidance for both animal shelter personnel and foster families to improve speed of adoption of the animals in their care.

category: animal behavior and cognition


title: Utility of Occupancy Models with Environmental DNA (eDNA) fromOlympic Coast National Marine Sanctuary

abstract:
Adjacent to the spectacular, rugged coastline bordering the Olympic Peninsula of Washington State, Olympic Coast National Marine Sanctuary (OCNMS) encompasses 8257 km2 of coastal waters that support one of North Americas most productive marine ecosystems. These rich waters support important recreational, commercial and subsistence fisheries for Washington state and four sovereign tribal governments: the Quinault Indian Nation and the Makah, Quileute and Hoh Tribes. Given its ecological, cultural and economic significance, managers from OCNMS are tasked with monitoring and management towards conserving the areas ecological integrity. The development of metabarcoding with environmental DNA (eDNA) as an effective freshwater monitoring tool has potential applications in marine systems, but the statistical analysis and the interpretation of those eDNA results are still under development. One promising strategy to analyze eDNA is through the use of occupancy models. Occupancy models enable us to calculate probabilities of detection from each taxon sequenced and are designed to work with presence-absence data. To inform our long term monitoring design of this coastal ecosystem, we collected 44 eDNA samples at nine of OCNMS long term mooring sites in 2019 and tested four different molecular markers for species reported and taxonomic richness. Additionally, we assessed occupancy models for use with eDNA to estimate the number of samples required to accurately predict the presence of a species. Occupancy models show great promise for use in eDNA studies provided there is sufficient replication; additionally, the choice of molecular marker strongly influences a taxas probability of detection.

Author SummaryWe show the utility of using occupancy models with eDNA in Olympic Coast National Marine Sanctuary for monitoring near-shore marine biological communities. Based on probabilities of detection, long term coastal monitoring projects using eDNA would benefit from designs prioritizing sample number (10-20 samples per site) over sites sampled. With proper study design, occupancy models provide a statistical framework for comparisons between sites and over time. By accounting for simple non-detection vs true absence, occupancy models help to eliminate noise from eDNA studies, increasing detection of true shifts in community composition. We also demonstrate that marker choice is an important study design consideration not only for the types of taxa recovered, but also for consistency between samples and timepoints.

category: bioinformatics


title: The Evaluation of Acute Myeloid Leukaemia (AML) Blood Cell Detection Models Using Different YOLO Approaches

abstract: This study proposes to evaluate the performance of Acute Myeloid Leukaemia (AML) blast cell detection models in microscopic examination images for faster diagnosis and disease monitoring. One of the popular deep learning algorithms such as You Only Look Once (YOLO) developed for object detection is the successful state-of-the-art algorithms in real-time object detection systems. We employ four versions of the YOLO algorithm: YOLOv3, YOLOv3-Tiny, YOLOv2 and YOLOv2-Tiny for detection of 15-class of AML blood cells in examination images. We also acquired the publicly available dataset from The Cancer Imaging Archive (TCIA), which consists of 18,365 expert-labelled single-cell images. Data augmentation techniques are additionally applied to enhance and balance the training images in the dataset. The overall results indicated that four types of YOLO approach have outstanding performances of more than 92% in precision and sensitivity. In comparison, YOLOv3 has more reliable performance than the other three approaches. Consistently, the AUC values for the four YOLO models are 0.969 (YOLOv3), 0.967 (YOLOv3-Tiny), 0.963 (YOLOv2), and 0.948 (YOLOv2-Tiny). Furthermore, we compare the best models performance between approaches that use the entire training dataset without using data augmentation techniques and image division with data augmentation techniques. Remarkably, by using 33.51 percent of the training data in model training, the prediction outcomes from the model that used image partitioning with data augmentation were similar to those obtained using the complete training dataset. This work potentially provides a beneficial digital rapid tool in the screening and evaluation of numerous haematological disorders.


title: BET inhibitors synergize with anti-PD1 by rescuing TCF1+ progenitor exhausted CD8+ T cells in Acute Myeloid Leukemia

abstract: Many acute myeloid leukemia (AML) patients exhibit hallmarks of immune exhaustion, such as increased myeloid derived suppressor cells (MDSCs), suppressive regulatory T cells (Tregs) and dysfunctional T cells. We have developed a mouse model of AML driven by Flt3-ITD and Tet2 deficiency displays these immune-related features, including CD8+ T cells exhibiting a terminally exhausted phenotype (TEx). This T cell subset has been shown to be refractory to immune checkpoint blockade (ICB) monotherapy. Here we show that small molecule inhibitors which target bromodomain and extra-terminal domain (BET) proteins affect both tumor-intrinsic factors but also rescue T cell exhaustion and ICB resistance. Ex vivo treatment of cells from AML mice and AML patients with BET inhibitors (BETi) reversed CD8+ T cell exhaustion by restoring proliferative capacity and expansion of the more functional precursor exhausted T cells (TPEx). This reversal is enhanced by combined BETi and anti-PD1 treatment. Finally, we show that BETi synergizes with anti-PD1 in vivo, resulting in the reduction of circulating leukemia cells, enrichment of CD8+ T cells in the bone marrow, and increased expression of Tcf7, Slamf6, and Cxcr5 in CD8+ T cells. In total, we show the potential efficacy of combining BETi and ICB therapy in the treatment of AML.


title: The role of antifreeze glycopeptides (AFGP) and polyvinyl alcohol/polyglycerol (X/Z-1000) cocktails as ice modulators during partial freezing of rat livers.

abstract: The current liver organ shortage has pushed the field of liver transplantation to develop new methods to prolong the preservation time of livers from the current clinical standard of static cold storage. Our approach, termed partial freezing, aims to induce a thermodynamically stable frozen state at deeper storage temperatures (-10{degrees}C to -15{degrees}C) than can be achieved with supercooling, while simultaneously maintaining a sufficient unfrozen fraction to limit dehydration and ice damage. This research first demonstrated that partially frozen glycerol treated rat livers were functionally similar after thawing from either -10 or -15{degrees}C with respect to subnormothermic machine perfusion metrics and histology. Next, we assessed the effect of adding either of two ice modulators, antifreeze glycoprotein (AFGP) and a polyvinyl alcohol/polyglycerol combination (X/Z-1000), on the viability and structural integrity of partially frozen rat livers compared to glycerol-only control livers. Results showed that AFGP livers had high levels of ATP and the least edema but suffered from significant endothelial cell damage. X/Z-1000 livers had the highest levels of ATP and energy charge (EC) but also demonstrated endothelial damage and post-thaw edema. Glycerol-only control livers exhibited the least DNA damage on Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining but also had the lowest levels of ATP and EC. Further research is necessary to optimize the ideal ice modulator cocktail for our partial-freezing protocol. Modifications to cryoprotective agent (CPA) combinations, as well as improvements to machine perfusion CPA loading and unloading, can help improve the viability of these partially frozen organs.

category: physiology


title: The creation of functioning gastrojejunostomy following magnetic compression

abstract:
BackgroundMagnetic compression for creating gastrojejunostomy has many advantages according to previous studies. However, following mechanical device release after healing, the anastomotic stenosis becomes the pivotal point.

MethodsRectangle-shaped magnets were used for magnetic compression in rabbits. Both paclitaxel-loaded magnets and a strategy of pyloric ligation were chosen to improve the gastrojejunostomy. Based on these choices, the half-capsule was applied to occlude the pylorus after anastomotic formation. The size and patency of the anastomoses were analyzed to evaluate the efficacy of these approaches. A histological examination was also performed.

ResultsThe positive effect of ligating the pylorus on gastrojejunostomy was significantly greater than that achieved using paclitaxel-loaded magnets during either short- or long-term follow-up. There were fewer scar tissue and collagen fibers at the anastomotic site in the treatment group than in the control group. The anastomotic aperture was of great interest at 9 months after the ligation of the pylorus following magnetic compression. In the view of the jejunum, although the aperture was barely visible, gastric juice was continuously spilling through it like a spring, and the aperture was clearly visible from the stomach side. All half-capsules failed to block the pylorus.

ConclusionThe effect of paclitaxel on maintaining gastrojejunostomy patency was temporary. The ligation of the pylorus ensured the long-term patency of gastrojejunostomy, and the aperture was comparable to the pylorus which could play an anti-reflux role. Further studies for the sort of gastrointestinal aperture are being planned.


title: Exploring of bacterial blight resistance in landraces, mining of resistant gene(s) and genetic divergence of resistant germplasm through molecular markers and quantitative traits.

abstract: Bacterial blight, one of the oldest and severe diseases of rice poses a major threat towards global rice production and food security. Thereafter, sustainable management of this disease has given paramount importance globally. In the current study, we explored 792 landraces to evaluate their disease reaction status against three highly virulent strains (BXo69, BXo87 and BXo93) of Xanthomonas oryzae pv. oryzae (Xoo). Subsequently, we intended to identify the possible candidate resistant (R) genes responsible for the resistant reaction using six STS markers correspond to Xa4, xa5, Xa7, xa13, Xa21 and Xa23 genes and finally, we evaluated morphological and molecular diversity of the potential bacterial blight resistant germplasm using quantitative traits and ISSR markers. Based on pathogenicity test, a single germplasm was found as highly resistant while, 33 germplasm were resistant and 40 were moderately resistant. Further molecular study on these 74 germplasm divulged that 41 germplasm carried Xa4 gene, 15 carried xa5 gene, 62 carried Xa7 gene, 33 carried xa13 gene, and 19 carried Xa23 gene. Only a single germplasm consisted of Xa21 gene. Interestingly, we found a wide range of gene combinations ranged from 2 to 4 genes among the germplasm resistant to bacterial blight and G3 genotype (Acc. No. 4216; highly resistant) having Xa4, Xa7, xa13, Xa21 and G43 genotype (Acc. No. 1523; resistant) having Xa4, xa5, xa13 and Xa23 gene combination being the most effective against all the Xoo strains. Nonetheless, UPGMA dendrogram and heatmap analysis based on quantitative traits identified two important clusters viz. cluster-III and cluster-VIII with multiple desired traits However, genetic similarity based on ISSR marker data pointed out 3 germplasm, namely, G20 (Acc. No. 4004; contained Xa4, Xa7, xa13 genes), G17 (Acc. No. 3981; contained Xa7, xa13 genes) and G6 (Acc. No. 991; contained Xa4, Xa7, xa13, Xa23 genes) having comparatively lower genetic similarity (0.30, 0.37 and 0.38, respectively) with elite variety BRRI dhan28. Notably, Mantle test of molecular and morphological data indicated that there was a positive correlation (r = 0.113) between them. The outcome of this study would enrich and diversify the rice gene pool and would be useful for the development of durable bacterial blight resistant varieties.

category: plant biology


title: Development and evaluation of a decision prediction tool for the reduction of fungicide applications for the control of strawberry powdery mildew epidemics

abstract: Strawberry powdery mildew (Podosphaera aphanis) causes serious losses in UK crops, potentially reducing yields by as much as 70%. Consequently, conventional fungicide application programmes tend to recommend a prophylactic approach using insurance sprays, risking the development of fungicide insensitivity and requiring careful management relative to harvest periods to avoid residual fungicides on harvested fruit. This paper describes the development of a prediction system to guide the control of P. aphanis by the application of fungicides only when pathogen infection and disease progression are likely. The system was developed over a 15-year period on commercial farms starting with its establishment, validation and then deployment to strawberry growers. This involved three stages: 1. Identification and validation of parameters for inclusion in the prediction system (2004-2008). 2. Development of the prediction system in compact disc format (2009-2015). 3. Development and validation of the prediction system in a web-based format and cost-benefit analysis (2016-2020). The prediction system was based on the temporal accumulation of conditions (temperature and relative humidity) conducive to the development of P. aphanis, which sporulates at 144 accumulated disease-conducive hours. Sensitivity analysis was performed to refine the prediction system parameters. Field validation of the results demonstrated that to effectively control disease, the application of fungicides was best done between 125 and 144 accumulated hours of disease-conducive conditions. A cost-benefit analysis indicated that, by comparison with the number and timing of fungicide applications in conventional insurance disease control programmes, the prediction system enabled good disease control with significantly fewer fungicide applications (between one and four sprays less) (df=7, t=7.6, p=0.001) and reduced costs (savings between {pound}35-{pound}493/hectare) (df=7, t=4.0, p=0.01) for the growers.


title: Retinoic acid treatment recruits macrophages and increases axonal regeneration after optic nerve injury in the frog Rana pipiens

abstract: Retinoic acid (RA) plays major roles during nervous system development, and during regeneration of the adult nervous system. We have previously shown that components of the RA signaling pathway are upregulated after optic nerve injury, and that exogenous application of all-trans retinoic acid (ATRA) greatly increases the survival of axotomized retinal ganglion cells (RGCs). The objective of the present study is to investigate the effects of ATRA application on the macrophages in the optic nerve after injury, and to determine whether this affects axonal regeneration. The optic nerve was crushed and treated with PBS, ATRA and/or clodronate-loaded liposomes. Nerves were examined at one and two weeks after axotomy with light microscopy, immunocytochemistry and electron microscopy. ATRA application to the optic nerve caused transient increases in the number of macrophages and microglia one week after injury. The macrophages are consistently labeled with M2-type markers, and have considerable phagocytic activity. ATRA increased ultrastructural features of ongoing phagocytic activity in macrophages at one and two weeks. ATRA treatment also significantly increased the numbers of regenerating GAP-43-labeled axons. Clodronate liposome treatment depleted macrophage numbers by 80%, completely eliminated the ATRA-mediated increase in axonal regeneration, and clodronate treatment alone decreased axonal numbers by 30%. These results suggest that the success of axon regeneration is partially dependent on the presence of debris-phagocytosing macrophages, and that the increases in regeneration caused by ATRA are in part due to their increased numbers. Further studies will examine whether macrophage depletion affects RGC survival.

category: neuroscience


title: Test-retest reproducibility of in vivo oscillating gradient and microscopic anisotropy diffusion MRI in mice at 9.4 Tesla

abstract:
Background and PurposeMicrostructure imaging with advanced diffusion MRI (dMRI) techniques have shown increased sensitivity and specificity to microstructural changes in various disease and injury models. Oscillating gradient spin echo (OGSE) dMRI, implemented by varying the oscillating gradient frequency, and microscopic anisotropy ({micro}A) dMRI, implemented via tensor valued diffusion encoding, may provide additional insight by increasing sensitivity to smaller spatial scales and disentangling fiber orientation dispersion from true microstructural changes, respectively. The aims of this study were to characterize the test-retest reproducibility of in vivo OGSE and {micro}A dMRI metrics in the mouse brain at 9.4 Tesla and provide estimates of required sample sizes for future investigations.

MethodsEight adult C57Bl/6 mice were scanned twice (5 days apart). Each imaging session consisted of multifrequency OGSE and {micro}A dMRI protocols. Metrics investigated included {micro}A, isotropic and anisotropic kurtosis, and the diffusion dispersion rate ({Lambda}), which explores the power-law frequency dependence of mean diffusivity. The dMRI metric maps were analyzed with mean region-of-interest (ROI) and whole brain voxel-wise analysis. Bland-Altman plots and coefficients of variation (CV) were used to assess the reproducibility of OGSE and {micro}A metrics. Furthermore, we estimated sample sizes required to detect a variety of effect sizes.

ResultsBland-Altman plots showed negligible biases between test and retest sessions. ROI-based CVs revealed high reproducibility for both {micro}A (CVs < 8 %) and {Lambda} (CVs < 15 %). Voxel-wise CV maps revealed high reproducibility for {micro}A (CVs [~] 10 %), but low reproducibility for OGSE metrics (CVs [~] 50 %).

ConclusionMost of the {micro}A dMRI metrics are reproducible in both ROI-based and voxel-wise analysis, while the OGSE dMRI metrics are only reproducible in ROI-based analysis. {micro}A and {Lambda} may provide sensitivity to subtle microstructural changes (4 - 8 %) with feasible sample sizes (10 - 15).


title: Reaching articular limits can negatively impact embodiment in virtual reality.

abstract: Providing Virtual Reality(VR) users with a 3D representation of their body complements the experience of immersion and presence in the virtual world with the experience of being physically located and more personally involved. A full-body avatar representation is known to induce a Sense of Embodiment (SoE) for this virtual body, which is associated with improvements in task performance, motivation and motor learning. Recent experimental research on embodiment provides useful guidelines, indicating the extent of discrepancy tolerated by users and, conversely, the limits and disruptive events that lead to a break in embodiment (BiE). Based on previous works on the limit of agency under movement distortion, this paper describes, studies and analyses the impact of a very common yet overlooked embodiment limitation linked to articular limits when performing a reaching movement. We demonstrate that perceiving the articular limit when fully extending the arm provides users with an additional internal proprioceptive feedback which, if not matched in the avatars movement, leads to the disruptive realization of an incorrect posture mapping. This study complements previous works on self-contact and visuo-haptic conflicts and emphasizes the risk of disrupting the SoE when distorting users movements or using a poorly-calibrated avatar.


title: IMMMUNE MEDIATORS IN IDIOPATHIC UVEITIS

abstract: To investigate which cytokines, chemokines and growth factors are involved in the immunopathogenesis of idiopathic uveitis, and whether cytokine profiles are associated with. Serum and aqueous humor (AH) samples of 75 patients with idiopathic uveitis were analyzed by multiplex immunoassay. Infectious controls consisted of 16 patients with ocular toxoplasmosis all confirmed by intraocular fluid analyses. Noninfectious controls consisted of 7 patients with Behcet disease related uveitis and 15 patients with sarcoidosis related uveitis. The control group consisted of AH and serum samples from 47 noninflammatory control patients with age-related cataract. In each sample, 27 immune mediators {+/-} IL-21 and IL-23 were measured. In idiopathic uveitis, 13 of the 29 mediators, including most proinflammatory and vascular mediators such as IL-6, IL-8, IL-12, G-CSF, GM-CSF, MCP-1, IP-10, TNF- and VEGF, were significantly elevated in the aqueous humor when compared to all controls. Moreover, IL-17, IP-10, and IL-21, were significantly elevated in the serum when compared to all controls. We clustered 4 subgroups of idiopathic uveitis using a statistical analysis of hierarchical unsupervised classification, characterized by the order of magnitude of concentrations of intraocular cytokines. The pathogenesis of idiopathic uveitis is characterized by the presence of predominantly proinflammatory cytokines and chemokines and vascular endothelial growth factor with high expression levels as compared to other causes of uveitis. There are indications for obvious Th-1/ IL21-Th17 pathways but also IL9-Th9 and increased IFN-{gamma}-inducing cytokine (IL12) and IFN-{gamma}-inducible CXC chemokine (IP-10). The combined data suggest that immune mediator expression is different among idiopathic uveitis. This study suggests various clusters among the idiopathic uveitis group rather than one specific uveitis entity.


title: Genomic diversity of bacteriophages infecting Rhodobacter capsulatus and their relatedness to its gene transfer agent RcGTA

abstract: The diversity of bacteriophages is likely unparalleled in the biome due to the immense variety of hosts and the multitude of viruses that infect them. Recent efforts have led to description at the genomic level of numerous bacteriophages that infect the Actinobacteria, but relatively little is known about those infecting other prokaryotic phyla, such as the purple non-sulfur photosynthetic -proteobacterium Rhodobacter capsulatus. This species is a common inhabitant of freshwater ecosystems and has been an important model system for the study of photosynthesis. Additionally, it is notable for its utilization of a unique form of horizontal gene transfer via a bacteriophage-like element known as the gene transfer agent (RcGTA). Only three bacteriophages of R. capsulatus had been sequenced prior to this report. Isolation and characterization at the genomic level of 26 new bacteriophages infecting this host advances the understanding of bacteriophage diversity and the origins of RcGTA. These newly discovered isolates can be grouped along with three that were previously sequenced to form six clusters with four remaining as single representatives. These bacteriophages share genes with RcGTA that seem to be related to host recognition. One isolate was found to cause lysis of a marine bacterium when exposed to high titer lysate. Although some clusters are more highly represented in the sequenced genomes, it is evident that many more bacteriophage types that infect R. capsulatus are likely to be found in the future.


title: Preliminary analysis of a neotropical plant extract antiviral potential against Chikungunya and Mayaro viruses

abstract: Chikungunya and Mayaro fevers are viral infectious diseases, without vaccine or treatment, that causes fever and arthralgia. Establishing novel antiviral tools capable of preventing or treating Chikungunya virus (CHIKV), and Mayaro virus (MAYV) infections are needed. The use of plant-based compounds that affect the replication cycles of these viruses has been proposed as a promising strategy. Chiococca alba (L.) Hitchc. is a neotropical plant used by Yucatec Mayas traditional healers, mainly, as antipyretic and antirheumatic. To evaluate the potential of C. alba methanolic extracts against CHIKV and MAYV through preliminary analysis in vitro and in silico. The cytotoxicity profile of two C. alba roots methanolic extracts in Vero cells was performed by lysosomal viability using the neutral red assay, and the antiviral potential was determined by plaque assay. We further assessed, through in silico computational predictions, the possible interactions between the active site of the nsP2 proteases of these viruses with some secondary metabolites present in C. alba extracts, identified by High-Performance Liquid Chromatography (HPLC). Our partial phytochemical analysis revealed the presence of flavonoids, and phenolic acids in the C. alba extracts. Our in vitro assays showed that both C. alba extracts inhibited more than 70% of CHIKV and MAYV activities at 60 {micro}g/mL concentration. Based on our in silico computational predictions, the flavonoids naringin and vitexin showed the greatest affinity energies with the CHIKV and MAYV nsP2 proteases, revealing the great potential of these compounds as viral inhibitors. The findings described here indicates that C. alba extracts, or their secondary metabolites, as a potential source of novel antiviral compounds.


title: Larval ecology and bionomics of Anopheles funestus in highland and lowland sites in western Kenya

abstract:
BackgroundAn. funestus is a major Afrotropical vector of human malaria. This study sought to investigate the larval ecology, sporozoite infection rates and blood meal sources of An. funestus in western Kenya.

MethodsLarval surveys were carried out in Bungoma (Highland) and Kombewa (lowland) of western Kenya. Aquatic habitats were identified, characterized, georeferenced and carefully examined for mosquito larvae and predators. Indoor resting mosquitoes were sampled using pyrethrum spray catches. Adults and larvae were morphologically and molecularly identified to species. Sporozoite infections and blood meal sources were detected using real-time PCR and ELISA respectively.

ResultsOf the 151 aquatic habitats assessed, 62/80 (78%) in Bungoma and 58/71(82%) in Kombewa were positive for mosquito larvae. Of the 3,193 larvae sampled, An. funestus larvae constitute 38% (1224/3193). Bungoma recorded a higher number of An. funestus larvae (85%, 95%, CI, 8.722-17.15) than Kombewa (15%, 95%, CI, 1.33-3.91). Molecular identification of larvae showed that 89% (n=80) were An. funestus. Approximately 59%, 35% and 5% of An. funestus larvae co-existed with An. gambiae s.l, Culex spp and An. coustani in the same habitats respectively. Of 1,221 An. funestus s.l adults sampled, molecular identifications revealed that An. funestus constituted 87% (n=201) and 88% (n=179) in Bungoma and Kombewa, respectively. The Plasmodium falciparum sporozoite rate of An. funestus in Bungoma and Kombewa was 2% (3/174) and 1% (2/157), respectively, and the human blood index of An. funestus was 84% (48/57) and 89% (39/44) and for Bungoma and Kombewa, respectively.

ConclusionMan-made ponds had the highest abundance of An. funestus larvae. Multiple regression and principal component analyses identified the distance to the nearest house as the key environmental factor associated with the abundance of An. funestus larvae in aquatic habitats. This study serves as a guide for the control of An. funestus and other mosquito species to complement existing vector control strategies.

category: ecology


title: Height-diameter allometry for tropical forest in northern Amazonia

abstract: Height measurements are essential to manage and monitor forest biomass and carbon stocks. However, accurate estimation of this variable in tropical ecosystems is still difficult due to species heterogeneity and environmental variability. In this article, we compare and discuss six nonlinear allometric models parameterized at different scales (local, regional and pantropical). We also evaluate the height measurements obtained in the field by the hypsometer when compared with the true tree height. We used a dataset composed of 180 harvested trees in two distinct areas located in the Amapa State. The functional form of the Weibull model was the best local model, showing similar performance to the pantropical model. The inaccuracy detected in the hypsometer estimates reinforces the importance of incorporating new technologies in measuring individual tree heights. Establishing accurate allometric models requires knowledge of ecophysiological and environmental processes that govern vegetation dynamics and tree height growth. It is essential to investigate the influence of different species and ecological gradients on the diameter/height ratio.


title: Population structure and genetic diversity of chickpea germplasms

abstract: In various leguminous crops, chickpea is the fourth most important legume contributing 3.1% to the total legume production. Grains of chickpea are rich source of proteins, minerals and vitamins which makes them suitable for both food and feed. For any crop to be improved, the knowledge of genetic diversity of wild and elite cultivar is very important. Therefore among various available marker systems, molecular markers are more reliable and accurate, therefore are very commonly used for genetic diversity analysis, phylogenetic studies and cultivar identification. Due to several advantages of Inter Simple Sequence Repeat (ISSR) markers in present study we analyzed the genetic diversity, structure, cross-species transferability and allelic richness in 50 chickpea collection using 23 ISSR markers. The observed parameters such as allele number varied from 3 to 16, and PIC varied from 0.15 to 0.4988 respectively. Further, range of allele size varied from 150 to 1600 bp which shows the significance of ISSR markers for chickpea germplasms characterization. On the basis of ISSR marker genotypic data dendrogram were constructed which divides these 50 chickpea in group I and II showing the reliability of ISSR markers. Among 50 chickpea, the accession P 74-1 is in group I and rest are in group II. Further we made mini-core collection of 15 diverse chickpea and subgrouped them. Dendrogram, PCA, Dissimilarity matrix and Bayesian model based genetic clustering of 50 chickpea germplasms revealed that P 74-1,P 1883, P 1260 very diverse chickpea accession. Characterization of these diverse chickpea would help in maintenance breeding, conservation and in future could be used to develop climate resilient elite cultivar of chickpea. Utilization of these novel ISSRs markers in diversity analysis and population structure characterization of 50 chickpea germplasm suggests their wider efficacy in superior scale for molecular breeding studies in chickpea.


title: Intercontinental comparison of ferns community assemblages in Malaysian and Nigerian tropical forests

abstract: Research on ferns ecology has gained attention in the last decade, yet there is paucity of information on the intercontinental comparison of ferns community across two continents. This study focused on comparing the ferns community assemblages in tropical forests of Malaysia and Nigeria, thereby assessing the patterns of the species richness and diversity across the two continents. The diversity and taxonomic compositions of ferns were assessed using 180 plots of size 10 m x 10 m in each country. The species richness and other diversity indices were determined using the combined forests data for each country and for the individual forests. The observed and rarefied-extrapolated fern species richness are significantly higher in Malaysian forests than Nigerian forests. Also, the other diversity indices (Simpson index, Margalef index, and Fishers alpha) are significantly higher in Malaysian forests except Shannon index which showed no significant difference between the two biogeographic regions. There is a very low similarity in the taxonomic composition of ferns between the two biogeographic areas, although the similarity in composition increased with increasing taxonomic levels (genus and family levels). Terrestrial and epiphytic ferns are more dominant than the other life forms in the two countries. Since the two countries receive varying degrees of environmental factors, we then hypothesize that these observed differences are due to climatic differences as well as historical and evolutionary processes.


title: Gender identity and sexual experience affect mating behaviour and chemical profile in the lesser mealworm, Alphitobius diaperinus (Coleoptera: Tenebrionidae)

abstract: Alphitobius diaperinus (Coleoptera: Tenebrionidae), the lesser mealworm, is one of the most significant pests of the poultry industry worldwide. These insects cause structural damage in poultry houses and transmit several diseases, impacting chickens productivity and rearing costs. Although semiochemicals may offer alternative insect pest management strategies, basic information regarding pheromone identity and their role on the behavioural ecology according to their circadian pattern of sexual behaviour of A. diaperinus is essentially lacking. This study is aimed to analyse the relation of gender identity and sexual experience of adults of A. diaperinus on their mating behaviour and whether this response is related to their CHC profiles secreted. The following steps were taken to achieve the studys goal. First, the circadian pattern of their sexual activity was observed in newly emerged pairs for at least twenty-one days (virgin adults) and experienced adults collected from the field to identify a difference based on their sexual experience and achieve the optimal mating season to develop the following assays. Subsequently, Y-tube olfactometer bioassays were conducted to evaluate their odour bouquet attraction based on gender and sexual experience. Additionally, mating behaviour bioassays were conducted to evaluate the two factor effects. Finally, cuticular analysis was performed using gas chromatography-mass spectrometry to evaluate possible chemical differences based on the two factors. With statistical and multivariate analysis, we found that behavioural, mating and chemical responses are different based on their sexual experience. The mating sequences were described into precopulatory, copulatory and postcopulatory phases. This finding gives us a deeper understanding of the sexual communication during mating. In summary, our findings provide new insights into the mating system and chemical ecology of A. diaperinus. The results presented here may serve as a base for further studies to develop strategies for managing this pest.


title: Low-level overexpression of wild type TDP-43 causes late-onset, progressive neurodegeneration and paralysis in mice

abstract: Modestly increased expression of transactive response DNA binding protein (TDP-43) gene have been reported in amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and other neuromuscular diseases. However, whether this modest elevation triggers neurodegeneration is not known. Although high levels of TDP-43 overexpression have been modeled in mice and shown to cause early death, models with low-level overexpression that mimic the human condition have not been established. In this study, transgenic mice overexpressing wild type TDP-43 at less than 60% above the endogenous CNS levels were constructed, and their phenotypes analyzed by a variety of techniques, including biochemical, molecular, histological, behavioral techniques and electromyography. The TDP-43 transgene was expressed in neurons, astrocytes, and oligodendrocytes in the cortex and predominantly in astrocytes and oligodendrocytes in the spinal cord. The mice developed a reproducible progressive weakness ending in paralysis in mid-life. Detailed analysis showed [~]30% loss of large pyramidal neurons in the layer V motor cortex; in the spinal cord, severe demyelination was accompanied by oligodendrocyte injury, protein aggregation, astrogliosis and microgliosis, and elevation of neuroinflammation. Surprisingly, there was no loss of lower motor neurons in the lumbar spinal cord despite the complete paralysis of the hindlimbs. However, denervation was detected at the neuromuscular junction. These results demonstrate that low-level TDP-43 overexpression can cause diverse aspects of ALS, including late-onset and progressive motor dysfunction, neuroinflammation, and neurodegeneration. Our findings suggest that persistent modest elevations in TDP-43 expression can lead to ALS and other neurological disorders involving TDP-43 proteinopathy. Because of the predictable and progressive clinical paralytic phenotype, this transgenic mouse model will be useful in preclinical trial of therapeutics targeting neurological disorders associated with elevated levels of TDP-43.

category: genetics


title: Physiological and Metabolomic Consequences of Reduced Expression of the Drosophila brummer Triglyceride Lipase

abstract: Disruption of lipolysis has widespread effects on intermediary metabolism and organismal phenotypes. Defects in lipolysis can be modeled in Drosophila melanogaster through genetic manipulations of brummer (bmm), which encodes a triglyceride lipase orthologous to mammalian Adipose Triglyceride Lipase. RNAi-mediated knock-down of bmm in all tissues or metabolic specific tissues results in reduced locomotor activity, altered sleep patterns and reduced lifespan. Metabolomic analysis on flies in which bmm is downregulated reveals a marked reduction in medium chain fatty acids, long chain saturated fatty acids and long chain monounsaturated and polyunsaturated fatty acids, and an increase in diacylglycerol levels. Elevated carbohydrate metabolites and tricarboxylic acid intermediates indicate that impairment of fatty acid mobilization as an energy source may result in upregulation of compensatory carbohydrate catabolism. bmm downregulation also results in elevated levels of serotonin and dopamine neurotransmitters, possibly accounting for the impairment of locomotor activity and sleep patterns. Physiological phenotypes and metabolomic changes upon reduction of bmm expression show extensive sexual dimorphism. Altered metabolic states in the Drosophila model are relevant for understanding human metabolic disorders, since pathways of intermediary metabolism are conserved across phyla.


title: Kdm6a Deficiency Restricted to Mouse Hematopoietic Cells Causes an Age- and Sex-dependent Myelodysplastic Syndrome-Like Phenotype

abstract: AbstractKdm6a/Utx, a gene on the X chromosome, encodes a histone K27me3 demethylase that has an orthologue on the Y chromosome (Uty)[1]. We previously identified inactivating mutations of Kdm6a in approximately 50% of mouse acute promyelocytic leukemia samples; however, somatic mutations of KDM6A are more rare in human AML samples, ranging in frequency from 2-15% in different series of patients, where their role in pathogenesis is not yet clear. In this study, we show that female Kdm6aflox/flox mice (with allele inactivation initiated by Vav1-Cre in hematopoietic stem and progenitor cells (HSPCs) have a sex-specific phenotype that emerges with aging, with features resembling a myelodysplastic syndrome (MDS). Female Kdm6a-knockout (KO) mice have an age-dependent expansion of their HSPCs with aberrant self-renewal, but they did not differentiate normally into downstream progeny. These mice became mildly anemic and thrombocytopenic, but did not develop overt leukemia, or die from these cytopenias. ChIP-seq and ATAC-seq studies showed only minor changes in H3K27me3, H3K27ac, H3K4me, H3K4me3 and chromatin accessibility between Kdm6a-WT and Kdm6a-KO mice. Utilizing scRNA-seq, Kdm6a loss was linked to the transcriptional repression of genes that mediate hematopoietic cell fate determination. These data demonstrate that Kdm6a plays an important role in normal hematopoiesis, and that its inactivation may contribute to AML pathogenesis by altering the epigenetic state of HSPCs.


title: Identification of key genes and immune infiltration modulated by CPAP in Obstructive sleep apnea by integrated bioinformatics analysis

abstract: Patients with obstructive sleep apnea (OSA) experience partial or complete upper airway collapses during sleep resulting in nocturnal hypoxia-normoxia cycling. And continuous positive airway pressure (CPAP) is the golden treatment for OSA. Nevertheless, the exact mechanisms of action, especially the transcriptome effect of CPAP on OSA patients, remains elusive. The goal of this study was to evaluate the longitudinal alterations in peripheral blood mononuclear cells transcriptome profiles of OSA patients in order to identify the hub gene and immune response. GSE133601 were downloaded from Gene Expression Omnibus (GEO). We identified black module via weighted gene co-expression network analysis (WGCNA), the genes in which were correlated significantly with the clinical trait of CPAP treatment. Finally, eleven hub genes (TRAV10, SNORA36A, RPL10, OBP2B, IGLV1-40, H2BC8, ESAM, DNASE1L3, CD22, ANK3, ACP3) were traced and used to construct a random forest model to predict therapeutic efficacy of CPAP in OSA with a good performance with AUC of 0.92. We further studied the immune cells infiltration in OSA patients with CIBERSORT, and monocytes were found to be related with the remission of OSA and partially correlated with the hub genes identified. In conclusion, these key genes and immune infiltration may be of great importance in the remission of OSA and related research of these genes may provide a new therapeutic target for OSA in the future.


title: Sex- and age- dependent effect of pre-gestational chronic stress and mirtazapine treatment on neurobehavioral development of offspring

abstract: Hormonal fluctuations, such as the perinatal period, may increase susceptibility of women to depression, which in turn exert a negative impact on childs neurodevelopment, becoming a risk factor in development of neuropsychiatric disorders. Moreover, the use of antidepressants during this critical period presents a serious health concern for both the mother and the child, due to the consequences of treatment in terms of the reliability and safety for the proper neurodevelopment of the organism being not well known. Atypical antidepressants, such as mirtazapine, that targets both serotonergic and noradrenergic systems in the central nervous system (CNS), represent a novel focus of research due to its unique pharmacological profile. The aim of this work was to study the effects of maternal depression and/or perinatal antidepressant mirtazapine treatment on the neurobehavioral development of the offspring. Pre-gestationally chronically stressed or non-stressed Wistar rat dams were treated with either mirtazapine (10 mg/kg/day) or vehicle during pregnancy and lactation followed by analysis of offsprings behavior at juvenile and adolescent age. We found mirtazapine induced alterations of nursing behavior. In offspring, pregestational stress (PS) had an anxiogenic effect on adolescent males and increased their active behavior in forced swim test. Interaction between pregestational stress and mirtazapine treatment variously induced anxiolytic changes of juvenile and adolescent females and impairment of spatial memory in adolescent females as well. Hippocampal density of synaptophysin, pre-synaptic protein marker, was decreased mainly by mirtazapine treatment. In conclusion, our results show mirtazapine induced alterations in maternal behavior and several sex- and age-dependent changes in neurobehavioral development of offspring caused by both prenatal mirtazapine treatment and/or chronic pregestational stress.


title: Statistical potentials from the Gaussian scaling behaviour of chain fragments buried within protein globules

abstract: Knowledge-based approaches use the statistics collected from protein data-bank structures to estimate effective interaction potentials between amino acid pairs. Empirical relations are typically employed that are based on the crucial choice of a reference state associated to the null interaction case. Despite their significant effectiveness, the physical interpretation of knowledge-based potentials has been repeatedly questioned, with no consensus on the choice of the reference state. Here we use the fact that the Flory theorem, originally derived for chains in a dense polymer melt, holds also for chain fragments within the core of globular proteins, if the average over buried fragments collected from different non-redundant native structures is considered. After verifying that the ensuing Gaussian statistics, a hallmark of effectively non-interacting polymer chains, holds for a wide range of fragment lengths, we use it to define a  bona fide reference state. Notably, despite the latter does depend on fragment length, deviations from it do not. This allows to estimate an effective interaction potential which is not biased by the presence of correlations due to the connectivity of the protein chain. We show how different sequence-independent effective statistical potentials can be derived using this approach by coarse-graining the protein representation at varying levels. The possibility of defining sequence-dependent potentials is explored.


title: 11β-HSD1 inhibition does not affect murine tumour angiogenesis but may exert a selective effect on tumour growth by modulating inflammation and fibrosis

abstract:
Glucocorticoids inhibit angiogenesis by activating the glucocorticoid receptor. Inhibition of the glucocorticoid-activating enzyme 11{beta}-hydroxysteroid dehydrogenase type 1 (11{beta}-HSD1) reduces tissue-specific glucocorticoid action and promotes angiogenesis in murine models of myocardial infarction. Angiogenesis is important in the growth of some solid tumours. This study used murine models of squamous cell carcinoma (SCC) and pancreatic ductal adenocarcinoma (PDAC) to test the hypothesis that 11{beta}-HSD1 inhibition promotes angiogenesis and subsequent tumour growth. SCC or PDAC cells were injected into female FVB/N or C57BL6/J mice fed either standard diet, or diet containing the 11{beta}-HSD1 inhibitor UE2316. SCC tumours grew more rapidly in UE2316-treated mice, reaching a larger (P<0.01) final volume (0.158 {+/-} 0.037 cm3) than in control mice (0.051 {+/-} 0.007 cm3). However, PDAC tumour growth was unaffected. Immunofluorescent analysis of SCC tumours did not show differences in vessel density (CD31/alpha-smooth muscle actin) or cell proliferation (Ki67) after 11{beta}-HSD1 inhibition, and immunohistochemistry of SCC tumours did not show changes in inflammatory cell (CD3- or F4/80-positive) infiltration. In culture, the growth/viability (assessed by live cell imaging) of SCC cells was not affected by UE2316 or corticosterone. Second Harmonic Generation microscopy showed that UE2316 reduced Type I collagen (P<0.001), whilst RNA-sequencing revealed that multiple factors involved in the innate immune/inflammatory response were reduced in UE2316-treated SCC tumours.

11{beta}-HSD1 inhibition increases SCC tumour growth, likely via suppression of inflammatory/immune cell signalling and extracellular matrix deposition, but does not promote tumour angiogenesis or growth of all solid tumours.


title: Temperate tree seedlings have similar drought vulnerability despite having different hydraulic drought responses in adults

abstract: Climate change is projected result in higher frequencies of drought events across the world and lead to reduced performance in many temperate tree species. However, many studies in this area focus specifically on adult tree drought responses and overlook how trees in other age classes might differ in their vulnerability. Evidence shows that seedling drought response can differ from that of adults and furthermore that demographic performance in the seedling age class will have disproportionately strong effects on the assembly dynamics of future forests, together suggesting that understanding seedling drought responses will be critical to our ability to predict how forests will respond to climate change. In this study, we measured four indices of hydraulic response to drought (leaf water potential, photosynthetic capacity, non-structural carbohydrate concentration, and hydraulic conductivity), as well as interaction effects with shade treatments, for seedlings of two temperate tree species that differ in their adult drought response: isohydric Acer saccharum and anisohydric Quercus rubra. We found a strong isohydric response in A. saccharum seedlings that included conservation of leaf water potentials (>-1.8 MPa) and reductions in non-structural carbohydrate concentrations consistent with reduction of stomatal conductance. Quercus rubra seedlings were able to survive to more negative water potentials, but only rarely, and they showed a similar reduction in photosynthetic capacity as was found for A. saccharum. Our results suggest that, although Q. rubra seedlings display some anisohydric responses to drought, they are more isohydric than adults. Both species seem to be relatively similar in their vulnerability to drought despite the differences predicted from adult drought response, and our results suggest that seedlings of both species will be similarly vulnerable to future drought events.


title: Appetite regulating genes in zebrafish gut; a gene expression study

abstract: The underlying molecular pathophysiology of feeding disorders, particularly in peripheral organs, is still largely unknown. A range of molecular factors encoded by appetite-regulating genes are already described to control feeding behaviour in the brain. However, the important role of the gastrointestinal tract in the regulation of appetite and feeding in connection to the brain has gained more attention in the recent years. An example of such inter-organ molecular interaction can be the signals mediated by leptin, a key regulator of body weight, food intake and metabolism, with conserved anorexigenic effects in vertebrates. Leptin signal functions through its receptor (lepr) in multiple organs, including the brain and the gastrointestinal tract. So far, the regulatory connections between leptin signal and other appetite-regulating genes remain unclear, particularly in the gastrointestinal system. In this study, we used a zebrafish mutant with impaired function of leptin receptor to explore gut expression patterns of appetite-regulating genes, under different feeding conditions (normal feeding, 7-day fasting, 2 and 6-hours refeeding). We compared these expression patterns to those from wild-type zebrafish, in order to identify leptin-dependent differentially expressed genes located in the zebrafish gut. We provide evidence that most appetite-regulating genes are expressed in the zebrafish gut. On one hand, we did not observed significant differences in the expression of orexigenic genes after changes in the feeding condition, and only one orexigenic gene, hcrt, displayed differential expression under impaired leptin signal. On the other hand, we found 8 anorexigenic genes in wild-types (cart2, cart3, dbi, oxt, nmu, nucb2a, pacap and pomc), as well as 4 genes in lepr mutants (cart3, kiss1, kiss1r and nucb2a), to be differentially expressed in the zebrafish gut after changes in feeding conditions. Most of these genes also showed significant differences in their expression between wild-type and lepr mutant in at least one of the feeding conditions. Finally, we observed that impaired leptin signalling influences potential regulatory connections between anorexigenic genes in zebrafish gut, particularly connections involving cart2, cart3, kiss1, kiss1r, mchr2, nmu, nucb2a and oxt. Altogether, these transcriptional changes propose a potential role of the gastrointestinal tract in the regulation of feeding through changes in expression of certain anorexigenic genes in zebrafish.


title: The complete mitogenome of   Lysmata vittata (Crustacea: Decapoda: Hippolytidae) and its phylogenetic position in Decapoda

abstract: In this study, the complete mitogenome of Lysmata vittata (Crustacea: Decapoda: Hippolytidae) has been determined. The genome sequence was 22003 base pairs (bp) and it included thirteen protein-coding genes (PCGs), twenty-two transfer RNA genes (tRNAs), two ribosomal RNA genes (rRNAs) and three putative control regions (CRs). The nucleotide composition of AT was 71.50%, with a slightly negative AT skewness (-0.04). Usually the standard start codon of the PCGs was ATN, while cox1, nad4L and cox3 began with TTG, TTG and GTG. The canonical termination codon was TAA, while nad5 and nad4 ended with incomplete stop codon T, and cox1 ended with TAG. We compared the order of genes of Decapoda ancestor and found that the positions of the two tRNAs genes (trnA and trnR) of the L. vittata were translocated. The phylogenetic tree showed that L. vittata was an independent clade, namely Hippolytidae.


title: Comparison of genetic variation between rare and common congeners of Dipodomys with estimates of contemporary and historical effective population size

abstract: Organisms with low effective population sizes are at greater risk of extinction because of reduced genetic diversity. Dipodomys elator is a kangaroo rat that is classified as threatened in Texas and field surveys from the past 50 years indicate that the distribution of this species has decreased. This suggests geographic range reductions that could have caused population fluctuations, potentially impacting effective population size. Conversely, the more common and widespread D. ordii is thought to exhibit relative geographic and demographic stability. Genetic variation between D. elator and D. ordii samples was assessed using 3RAD, a modified restriction site associated sequencing approach. It was hypothesized that D. elator would show lower levels of nucleotide diversity, observed heterozygosity, and effective population size when compared to D. ordii. Also of interest was identifying population structure within contemporary samples of D. elator and detecting genetic variation between temporal samples that could indicate demographic dynamics. Up to 61,000 single nucleotide polymorphisms were analyzed. It was determined that genetic variability and effective population size in contemporary D. elator populations were lower than that of D. ordii, that there is only slight, if any, structure within contemporary D. elator populations, and there is little genetic differentiation between spatial or temporal historical samples suggesting little change in nuclear genetic diversity over 30 years. Results suggest that genetic diversity of D. elator has remained stable despite claims of reduced population size and/or abundance, which may indicate a metapopulation-like system, whose fluctuations might counteract any immediate decrease in fitness.


title: Slightly different metabolomic profiles are associated with high or low weights duck foie gras

abstract: Understanding the evolution of fatty liver metabolism of ducks is a recurrent issue for researchers and industry. Indeed, the increase in weight during the overfeeding period leads to an important change in the liver metabolism. However, liver weight is highly variable at the end of overfeeding within a batch of animals reared, force-fed and slaughtered in the same way. For this study, we performed a proton nuclear magnetic resonance (1H-NMR) analysis on two classes of fatty liver samples, called low-weight liver (weight between 550 and 599 g) and high-weight liver (weight above 700 g). The aim of this study was to identify the differences in metabolism between two classes of liver weight (low and high). Firstly, the results showed that increased liver weight is associated with higher glucose uptake leading to greater lipid synthesis. Secondly, this increase is probably also due to a decline in the level of export of triglycerides and cholesterol from the liver by maintaining them at high hepatic concentration levels. Finally, the increase in liver weight could lead to a significant decrease in the efficiency of aerobic energy metabolism associated with a significant increase in the level of oxidative stress.


title: Evaluation of freshness and physicochemical characteristics of blankets of pirarucu (Arapaima gigas) bred in captivity at different ages in relation to the seasons

abstract: This study aimed to evaluate the freshness and physicochemical characterization of pirarucu blankets at different ages bred in captivity in excavated tanks in northern Mato Grosso State, Brazil, in different seasons (rain and dry seasons). Four harvests were performed in two different tanks and five specimens obtained in each evaluated period, with animals aged 18 and 24 months, respectively, at the beginning of the experiment. The animals were slaughtered in a local establishment that had an inspection service following humane standards, and the blankets were later sent to analysis. The samples were analyzed for freshness via total volatile nitrogenous bases and physicochemical characteristics (pH in 24 h, water activity, water retention capacity, dripping loss, cooking loss, and shear force). There was a statistical difference for total nitrogenous volatile bases and shear force with higher values in the rainy season (24 month-of-old animals). Additionally, pH and water activity differed statistically at 24 h in the same period, albeit for 18-month-of-old animals. It was possible to conclude that climate variation affected the freshness and physicochemical characteristics of the pirarucu blankets while age did not.


title: Change in the bladder function of rats with focal cerebral infarction induced by photochemically-induced thrombosis

abstract: The photochemically-induced thrombosis(photothrombosis) method can create focal cerebral infarcts anywhere in the relatively superficial layers of the cerebrum; it is easy to implement and minimally invasive. Taking advantage of this versatility, we aimed to establish a new rat model of urinary frequency with focal cerebral infarction, which was characterized by its simplicity, nonlethal nature, and high reproducibility. The prefrontal cortex and the anterior cingulate cortex, which are urinary centers, were targeted for focal cerebral infarction, and urinary parameters were measured by cystometrogram. Cystometric analysis indicated that micturition intervals significantly shortened in photothrombosis-treated rats compared with those in the sham operative group on Days 1 and 7 (P < 0.01), but prolonged after 14 days, with no difference between the two groups. Immunopathological evaluation showed an accumulation of activated microglia, followed by an increase in reactive astrocytes at the peri-infarct zone after photothrombotic stroke. Throughout this study, all postphotothrombosis rats showed cerebral infarction in the prefrontal cortex and anterior cingulate cortex; there were no cases of rats with fatal cerebral infarction. This model corresponded to the clinical presentation, in that the micturition status changed after stroke. In conclusion, this novel model combining nonlethality and high reproducibility may be a suitable model of urinary frequency after focal cerebral infarction.


title: Extraction and high-throughput sequencing of oak heartwood DNA: assessing the feasibility of genome-wide DNA methylation profiling

abstract: Tree ring features are affected by environmental factors and therefore are the basis for dendrochronological studies to reconstruct past environmental conditions. Oak wood often provides the data for these studies because of the durability of oak heartwood and hence the availability of samples spanning long time periods of the distant past. Wood formation is regulated in part by epigenetic mechanisms such as DNA methylation. Studies of the methylation state of DNA preserved in oak heartwood thus could identify epigenetic tree ring features informing on past environmental conditions. In this study, we aimed to establish protocols for the extraction of DNA, the high-throughput sequencing of whole-genome DNA libraries (WGS) and the profiling of DNA methylation by whole-genome bisulfite sequencing (WGBS) for oak (Quercus spp.) heartwood drill cores taken from the trunks of living standing trees spanning the AD 1776-2014 time period. Heartwood contains little DNA, and large amounts of phenolic compounds known to hinder the preparation of high-throughput sequencing libraries. Whole-genome and DNA methylome library preparation and sequencing consistently failed for oak heartwood samples more than 100 and 50 years of age, respectively. DNA fragmentation increased with sample age and was exacerbated by the additional bisulfite treatment step during methylome library preparation. Relative coverage of the non-repetitive portion of the oak genome was sparse. These results suggest that quantitative methylome studies of oak hardwood will likely be limited to relatively recent samples and will require a high sequencing depth to achieve sufficient genome coverage.


title: Distinctive neural correlates of phonological processing and reading impairment in fetal alcohol-exposed adolescents with and without facial dysmorphology

abstract:
AbstractO_ST_ABSBackgroundC_ST_ABSPrenatal alcohol exposure (PAE) has been linked to atypical brain development and a wide range of cognitive and behavioral impairments, including poor reading performance in childhood and adolescence. However, little is known about how structural and/or functional teratogenesis in the brain mediate reading impairment in fetal alcohol spectrum disorders (FASD) and whether neural correlates of reading and phonological processing differ between FASD subtypes with different clinical presentations in facial morphology.

MethodsThe current study utilized functional magnetic resonance imaging (fMRI) and diffusion tensor imaging (DTI) to characterize functional and structural mechanisms mediating reading deficits in 26 syndromal adolescents with PAE-related facial dysmorphology (i.e., fetal alcohol syndrome (FAS) or partial FAS (PFAS)) and 30 heavily exposed (HE) without this dysmorphology, in comparison with 19 typically developing controls. Importantly, the levels of PAE and reading ability were comparable between the FAS/PFAS and HE groups in the current study.

ResultsCompared to the nonsyndromal HE and control groups, the syndromal adolescents showed greater activation in the right precentral gyrus during an fMRI phonological processing task and rightward lateralization in an important reading-related tract (inferior longitudinal fasciculus, ILF), suggesting an atypical reliance on the right hemisphere during reading. By contrast, in the HE group, better reading skills were associated with increased neural activation in the left angular gyrus (LAG) and higher fractional anisotropy in the white matter organization of the left ILF. However, the brain function-behavior relation was weaker in the HE than among the controls, suggesting less efficient function of the typical reading neural network that may contribute to the observed reading impairments.

ConclusionsOur findings provide the first evidence for the distinctive functional and structural mechanisms underlying atypical reading and phonological processing in PAE adolescents with and without FAS facial dysmorphology.

HighlightsO_LIPrenatal alcohol exposure is associated with altered neural reading networks
C_LIO_LIFASD subtypes exhibit distinctive neural correlates of phonological processing
C_LIO_LIGreater right-hemispheric reliance was observed in FASD with facial dysmorphology
C_LIO_LINon-syndromal FASD showed deficits in the typical left-hemispheric reading network
C_LI


title: Efficient differentiation of human primordial germ cells through geometric control reveals a key role for NODAL signaling

abstract: Human primordial germ cells (hPGCs) form around the time of implantation and are the precursors of eggs and sperm. Many aspects of hPGC specification remain poorly understood. Here we show that micropatterned human pluripotent stem cells (hPSCs) treated with BMP4 give rise to hPGC-like cells (hPGCLC) and use these as a quantitatively reproducible and simple in vitro model to interrogate this important developmental event. We characterize micropatterned hPSCs up to 96h for the first time and show that hPGCLC populations are stable and continue to mature. By perturbing signaling during hPGCLC differentiation, we identify a previously unappreciated role for NODAL signaling and find that the relative timing and duration of BMP and NODAL signaling are critical parameters controlling the number of hPGCLCs. We formulate a mathematical model for a network of cross-repressive fates driven by NODAL and BMP signaling which predicts the measured fate patterns after signaling perturbations. Finally, we show that hPSC colony size dictates the efficiency of hPGCLC specification, which led us to dramatically improve the efficiency of hPGCLC differentiation over current protocols.

category: developmental biology