Medication and adverse drug event (ADE) information extracted from electronic health record (EHR) notes can be a rich resource for drug safety surveillance. Existing observational studies have mainly relied on structured EHR data to obtain ADE information; however, ADEs are often buried in the EHR narratives and not recorded in structured data.

Different products of combined oral contraceptives (COCs) can improve clinical and biochemical findings in patients with polycystic ovary syndrome (PCOS) through suppression of the hypothalamic-pituitary-gonadal (HPG) axis.

Campomelic dysplasia is a rare skeletal dysplasia characterized by Pierre Robin sequence, craniofacial dysmorphism, shortening and angulation of long bones, tracheobronchomalacia, and occasionally sex reversal. The disease is due to mutations in SOX9 or chromosomal rearrangements involving the long arm of Chromosome 17 harboring the SOX9 locus. SOX9, a transcription factor, is indispensible in establishing and maintaining neural stem cells in the central nervous system. We present a patient with angulation of long bones and external female genitalia on prenatal ultrasound who was subsequently found to harbor the chromosomal abnormality 46, XY, t(6;17) (p21.1;q24.3) on prenatal genetic testing. Comparative genomic hybridization revealed deletions at 6p21.1 and 17q24.3, the latter being 2.3 Mb upstream of SOX9 Whole-exome sequencing did not identify pathogenic variants in SOX9, suggesting that the 17q24.3 deletion represents a translocation breakpoint farther upstream of SOX9 than previously identified. At 2 mo of age the patient developed progressive communicating ventriculomegaly and thinning of the cortical mantle without clinical signs of increased intracranial pressure. This case suggests ventriculomegaly in some cases represents not a primary impairment of cerebrospinal fluid dynamics, but an epiphenomenon driven by a genetic dysregulation of neural progenitor cell fate.

Multimerization is a key characteristic of most voltage-sensing proteins. The main exception was thought to be the Ciona intestinalis voltage-sensing phosphatase (Ci-VSP). In this study, we show that multimerization is also critical for Ci-VSP function. Using coimmunoprecipitation and single-molecule pull-down, we find that Ci-VSP stoichiometry is flexible. It exists as both monomers and dimers, with dimers favored at higher concentrations. We show strong dimerization via the voltage-sensing domain (VSD) and weak dimerization via the phosphatase domain. Using voltage-clamp fluorometry, we also find that VSDs cooperate to lower the voltage dependence of activation, thus favoring the activation of Ci-VSP. Finally, using activity assays, we find that dimerization alters Ci-VSP substrate specificity such that only dimeric Ci-VSP is able to dephosphorylate the 3-phosphate from PI(3,4,5)P3 or PI(3,4)P2 Our results indicate that dimerization plays a significant role in Ci-VSP function.

Bunyaviruses pose a significant threat to human health, prosperity, and food security. In response to viral infections, interferons (IFNs) upregulate the expression of hundreds of interferon-stimulated genes (ISGs), whose cumulative action can potently inhibit the replication of bunyaviruses. We used a flow cytometry-based method to screen the ability of ∼500 unique ISGs from humans and rhesus macaques to inhibit the replication of Bunyamwera orthobunyavirus (BUNV), the prototype of both the Peribunyaviridae family and the Bunyavirales order. Candidates possessing antibunyaviral activity were further examined using a panel of divergent bunyaviruses. Interestingly, one candidate, ISG20, exhibited potent antibunyaviral activity against most viruses examined from the Peribunyaviridae, Hantaviridae, and Nairoviridae families, whereas phleboviruses (Phenuiviridae) largely escaped inhibition. Similar to the case against other viruses known to be targeted by ISG20, the antibunyaviral activity of ISG20 is dependent upon its functional RNase activity. Through use of an infectious virus-like particle (VLP) assay (based on the BUNV minigenome system), we confirmed that gene expression from all 3 viral segments is strongly inhibited by ISG20. Using in vitro evolution, we generated a substantially ISG20-resistant BUNV and mapped the determinants of ISG20 sensitivity/resistance. Taking all the data together, we report that ISG20 is a broad and potent antibunyaviral factor but that some bunyaviruses are remarkably ISG20 resistant. Thus, ISG20 sensitivity/resistance may influence the pathogenesis of bunyaviruses, many of which are emerging viruses of clinical or veterinary significance.IMPORTANCE There are hundreds of bunyaviruses, many of which cause life-threatening acute diseases in humans and livestock. The interferon (IFN) system is a key component of innate immunity, and type I IFNs limit bunyaviral propagation both in vitro and in vivo Type I IFN signaling results in the upregulation of hundreds of IFN-stimulated genes (ISGs), whose concerted action generates an "antiviral state." Although IFNs are critical in limiting bunyaviral replication and pathogenesis, much is still unknown about which ISGs inhibit bunyaviruses. Using ISG-expression screening, we examined the ability of ∼500 unique ISGs to inhibit Bunyamwera orthobunyavirus (BUNV), the prototypical bunyavirus. Using this approach, we identified ISG20, an interferon-stimulated exonuclease, as a potent inhibitor of BUNV. Interestingly, ISG20 possesses highly selective antibunyaviral activity, with multiple bunyaviruses being potently inhibited while some largely escape inhibition. We speculate that the ability of some bunyaviruses to escape ISG20 may influence their pathogenesis.

Successful drug treatment for tuberculosis (TB) depends on the unique contributions of its component drugs. Drug resistance poses a threat to the efficacy of individual drugs and the regimens to which they contribute. Biologically and chemically validated targets capable of replacing individual components of current TB chemotherapy are a major unmet need in TB drug development. We demonstrate that chemical inhibition of the bacterial biotin protein ligase (BPL) with the inhibitor Bio-AMS (5'-[N-(d-biotinoyl)sulfamoyl]amino-5'-deoxyadenosine) killed Mycobacterium tuberculosis (Mtb), the bacterial pathogen causing TB. We also show that genetic silencing of BPL eliminated the pathogen efficiently from mice during acute and chronic infection with Mtb Partial chemical inactivation of BPL increased the potency of two first-line drugs, rifampicin and ethambutol, and genetic interference with protein biotinylation accelerated clearance of Mtb from mouse lungs and spleens by rifampicin. These studies validate BPL as a potential drug target that could serve as an alternate frontline target in the development of new drugs against Mtb.

Faithful genome transmission in dividing cells requires that the two copies of each chromosome's DNA package into separate but physically linked sister chromatids. The linkage between sister chromatids is mediated by cohesin, yet where sister chromatids are linked and how they resolve during cell cycle progression has remained unclear. In this study, we investigated sister chromatid organization in live human cells using dCas9-mEGFP labeling of endogenous genomic loci. We detected substantial sister locus separation during G2 phase irrespective of the proximity to cohesin enrichment sites. Almost all sister loci separated within a few hours after their respective replication and then rapidly equilibrated their average distances within dynamic chromatin polymers. Our findings explain why the topology of sister chromatid resolution in G2 largely reflects the DNA replication program. Furthermore, these data suggest that cohesin enrichment sites are not persistent cohesive sites in human cells. Rather, cohesion might occur at variable genomic positions within the cell population.

The laboratory diagnosis of infectious diseases, especially those caused by mixed infections, is challenging. Routinely, it requires submission of multiple samples to separate laboratories. Advances in next-generation sequencing (NGS) have provided the opportunity for development of a comprehensive method to identify infectious agents. This study describes the use of target-specific primers for PCR-mediated amplification with the NGS technology in which pathogen genomic regions of interest are enriched and selectively sequenced from clinical samples. In the study, 198 primers were designed to target 43 common bovine and small-ruminant bacterial, fungal, viral, and parasitic pathogens, and a bioinformatics tool was specifically constructed for the detection of targeted pathogens. The primers were confirmed to detect the intended pathogens by testing reference strains and isolates. The method was then validated using 60 clinical samples (including tissues, feces, and milk) that were also tested with other routine diagnostic techniques. The detection limits of the targeted NGS method were evaluated using 10 representative pathogens that were also tested by quantitative PCR (qPCR), and the NGS method was able to detect the organisms from samples with qPCR threshold cycle (CT ) values in the 30s. The method was successful for the detection of multiple pathogens in the clinical samples, including some additional pathogens missed by the routine techniques because the specific tests needed for the particular organisms were not performed. The results demonstrate the feasibility of the approach and indicate that it is possible to incorporate NGS as a diagnostic tool in a cost-effective manner into a veterinary diagnostic laboratory.

In the present study, an innovative top-down liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the identification of clinically relevant fungi is tested using a model set of dermatophyte strains. The methodology characterizes intact proteins derived from Trichophyton species, which are used as parameters of differentiation. To test its resolving power compared to that of traditional Sanger sequencing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF), 24 strains of closely related dermatophytes, Trichophyton rubrum, T. violaceum, T. tonsurans, T. equinum, and T. interdigitale, were subjected to this new approach. Using MS/MS and different deconvolution algorithms, we identified hundreds of individual proteins, with a subpopulation of these used as strain- or species-specific markers. Three species, i.e., T. rubrum, T. violaceum, and T. interdigitale, were identified correctly down to the species level. Moreover, all isolates associated with these three species were identified correctly down to the strain level. In the T. tonsurans-equinum complex, eight out of 12 strains showed nearly identical proteomes, indicating an unresolved taxonomic conflict already apparent from previous phylogenetic data. In this case, it was determined with high probability that only a single species can be present. Our study successfully demonstrates applicability of the mass spectrometric approach to identify clinically relevant filamentous fungi. Here, we present the first proof-of-principle study employing the mentioned technology to differentiate microbial pathogens. The ability to differentiate fungi at the strain level sets the stage to improve patient outcomes, such as early detection of strains that carry resistance to antifungals.

Neural stem cells (NSCs) in the adult vertebrate brain are found in a quiescent state and can preserve long-lasting progenitor potential (stemness). Whether and how these two properties are linked, and to what extent they can be independently controlled by NSC maintenance pathways, is unresolved. We have previously identified Notch3 signalling as a major quiescence-promoting pathway in adult NSCs of the zebrafish pallium. We now show that Notch3 also controls NSC stemness. Using parallel transcriptomic characterizations of notch3 mutant NSCs and adult NSC physiological states, we demonstrate that a set of potentially direct Notch3 target genes distinguishes quiescence and stemness control. As a proof of principle, we focus on one 'stemness' target, encoding the bHLH transcription factor Hey1, that has not yet been analysed in adult NSCs. We show that abrogation of Hey1 function in adult pallial NSCs in vivo, including quiescent NSCs, leads to their differentiation without affecting their proliferation state. These results demonstrate that quiescence and stemness are molecularly distinct outputs of Notch3 signalling, and identify Hey1 as a major Notch3 effector controlling NSC stemness in the vertebrate adult brain.

Hirschsprung disease (HSCR) is a genetic disorder characterized by the absence of neural crest cells in parts of the intestine. This study aims to investigate the association of vesicle-associated membrane protein 5 (VAMP5) and mutated in colorectal cancer (MCC) genetic polymorphisms and their correlated risks with HSCR. We examined the association in four polymorphisms (rs10206961, rs1254900 and rs14242 in VAMP5, rs11241200 in MCC) and HSCR susceptibility in a Southern Chinese population composed of 1473 cases and 1469 controls. Two variants in VAMP5 were replicated as associated with HSCR. Interestingly, we clarified SNPs rs10206961 and rs1254900 in VAMP5 are more essential for patients with long-segment aganglionosis (LHSCR). Relatively high expression correlation was observed between VAMP5 and MCC using data from public database showing there may exist potential genetic interactions. SNP interaction was cross-examined by logistic regression and multifactor dimensionality reduction analysis revealing that VAMP5 rs1254900 and MCC rs11241200 were interacting significantly, thereby contributing to the risk of HSCR. The results suggest that significant associations of the rs10206961 and rs14242 in VAMP5 with an increased risk of HSCR in Southern Chinese, especially in LHSCR patients. This study provided new evidence of epistatic association of VAMP5 and MCC with increased risk of HSCR.

Atherosclerosis is a chronic inflammatory process of the arterial wall and carotid intima-media thickness (cIMT) is regarded as its early marker. Several members of the IL-17 family are involved in pro-inflammatory functions. The specific aim of the study was to explore the relationships of common genetic variants on IL-17 genes with cIMT thickening.

The objective of this study was to evaluate the efficacy of a non-selective COX inhibitor (tolfenamic acid) and a selective COX-2 inhibitor (robenacoxib) for post-operative pain control in cats. Thirty cats undergoing ovariohysterectomy were randomly divided into three groups: the control (placebo) group, the tolfenamic acid (4 mg/kg/day) group, and the robenacoxib (1 mg/kg/day) group. Non-steroidal anti-inflammatory drugs (NSAIDs) were administered orally 2 hr before anesthesia induction and 24 and 48 hr post-operation. Buccal mucosal bleeding times (BMBTs) were assessed prior to anesthesia induction. Colorado pain scores and composite pain scores were evaluated in a blinded fashion before induction and 2, 8, 24, 30 and 48 hr post-operation. The Colorado pain scores of cats receiving robenacoxib were significantly lower than those of cats in the control group at 30 (P=0.0126) and 48 (P=0.0439) hr post-operation. The composite pain scores of cats from the robenacoxib group were lower than those of cats in the control group at 30 (P=0.0299) and 48 (P=0.0103) hr post-operation. The Colorado pain scores of cats receiving tolfenamic acid were significantly lower than those of cats in the control group at 30 hr (P=0.0186) post-operation. The composite pain scores in cats in the tolfenamic acid group were lower than the scores of cats in the control group at 24 (P=0.0403) and 48 (P=0.0413) hr post-operation. BMBTs remained within normal limits in all groups. Both tolfenamic acid and robenacoxib are useful for post-operative pain control in cats.

Epithelial-mesenchymal transition (EMT) is an orchestral and functional change in epithelial cells. Many signaling pathways are involved in EMT, and transforming growth factor-beta (TGF-β) is considered to be one of the most important factors in induction of EMT. In this study, we treated the rat intestinal epithelial cell line (IEC-6) with TGF-β1 as a signaling stimulant. Gross analysis of IEC-6 cells showed typical characteristics of epithelial cells such as cuboidal morphology and cell-cell contact, whereas treatment with TGF-β1 (10 ng/ml-1) for 7 days produced robust, spindle-shaped morphology. Immunocytochemistry analysis showed distinct E-cadherin staining in IEC-6 cells, but weak and faint in EMT cells. EMT cells showed positive expression of α-SMA and tenascin-C but IEC-6 cells did not. Quantitative real-time PCR analysis showed that myosin light chain kinase and C-kinase potentiated protein phosphatase-1 inhibitor (CPI-17) mRNAs were significantly upregulated in EMT cells. Immunocytochemistry analysis also showed that EMT cells strongly expressed CPI-17 but IEC-6 cells did not. A collagen gel contraction assay revealed that EMT cells had greatly increased contraction compared with control cells. These results suggest that the increased contractile activity induced by TGF-β in EMT cells may be attributable to the upregulation of molecules responsible for myosin phosphorylation/de-phosphorylation.

Fowl cholera caused by Pasteurella multocida has always been a disease of global importance for poultry production. The aim of this study was to obtain more information about the epidemiology of avian P. multocida infection in southwest China and the genetic characteristics of clinical isolates. P. multocida isolates were characterized by biochemical and molecular-biological methods. The distributions of the capsular serogroups, the phenotypic antimicrobial resistance profiles, lipopolysaccharide (LPS) genotyping and the presence of 19 virulence genes were investigated in 45 isolates of P. multocida that were associated with clinical disease in poultry. The genetic diversity of P. multocida strains was performed by 16S rRNA and rpoB gene sequence analysis as well as multilocus sequence typing (MLST). The results showed that most (80.0%) of the P. multocida isolates in this study represented special P. multocida subspecies, and 71.1% of the isolates showed multiple-drug resistance. 45 isolates belonged to capsular types: A (100%) and two LPS genotypes: L1 (95.6%) and L3 (4.4%). MLST revealed two new alleles (pmi77 and gdh57) and one new sequence type (ST342). ST129 types dominated in 45 P. multocida isolates. Isolates belonging to ST129 were with the genes ompH+plpB+ptfA+tonB, whereas ST342 included isolates with fur+hgbA+tonB genes. Population genetic analysis and the MLST results revealed that at least one new ST genotype was present in the avian P. multocida in China. These findings provide novel insights into the epidemiological characteristics of avian P. multocida isolates in southwest China.

Angiopoietin-like proteins (ANGPTLs) 3 and 4 are circulating factors that participate in the regulation of lipid and glucose metabolism.

Many bioinformatics resources with unique perspectives on the protein landscape are currently available. However, generating new knowledge from these resources requires interoperable workflows that support cross-resource queries. In this study, we employ federated queries linking information from the Protein Kinase Ontology, iPTMnet, Protein Ontology, neXtProt, and the Mouse Genome Informatics to identify key knowledge gaps in the functional coverage of the human kinome and prioritize understudied kinases, cancer variants and post-translational modifications (PTMs) for functional studies. We identify 32 functional domains enriched in cancer variants and PTMs and generate mechanistic hypotheses on overlapping variant and PTM sites by aggregating information at the residue, protein, pathway and species level from these resources. We experimentally test the hypothesis that S768 phosphorylation in the C-helix of EGFR is inhibitory by showing that oncogenic variants altering S768 phosphorylation increase basal EGFR activity. In contrast, oncogenic variants altering conserved phosphorylation sites in the 'hydrophobic motif' of PKCβII (S660F and S660C) are loss-of-function in that they reduce kinase activity and enhance membrane translocation. Our studies provide a framework for integrative, consistent, and reproducible annotation of the cancer kinomes.

Nonsyndromic craniosynostosis (NSC) is the most common craniosynostosis with the primary defect being one or more fused sutures. In contrast to syndromic craniosynostosis, the etiopathogenesis of NSC is largely unknown. Here we show that autophagy, a major catabolic process required for the maintenance of bone homeostasis and bone growth, is a pathological change associated with NSC. Using calvarial suture mesenchymal cells (SMCs) isolated from the fused and unfused sutures of NSC patients, we demonstrate that during SMC differentiation, the level of the autophagosomal marker LC3-II increases as osteogenic differentiation progresses, particularly at differentiation day 7, a stage concurrent with mineralization. In fused SMCs, autophagic induction was more robust than that in unfused SMCs, which consequently led to enhanced mineralized nodule formation. Perturbation of autophagy with rapamycin or wortmannin promoted or inhibited the ossification of SMCs, respectively. Our findings suggest that autophagy is essential for the osteogenic differentiation of SMCs and that overactive autophagy is a molecular abnormality underlying premature calvarial ossification in NSC.

Mulching is critical for increasing water availability and hence winter wheat production in dryland farming systems. A two-year study was conducted to assess the effects of mulches on soil water storage (SWS), temperature, water use efficiency (WUE) and yields of winter wheat on the Loess Plateau. Four treatments were examined: conventional flat planting (CK), straw mulch (FPS), transparent plastic film mulch (FPP) and ridge-furrow with plastic film-mulched ridge and straw-mulched furrow (RFPS). Compared with CK, RFPS greatly increased SWS from 0-60 cm, FPP increased SWS from 0-40 cm, and FPS slightly increased SWS from 0-60 cm; however, FPP significantly (P < 0.05) decreased SWS from 61-100 cm. RFPS and FPP increased soil temperatures in cold seasons relative to CK, especially in RFPS (2.0-2.3 °C). Meanwhile, the rate of soil temperature increase was greater in RFPS and FPP than in CK but was lower in FPS. Mean yields were significantly increased in RFPS (56.78%), FPP (44.72%) and FPS (9.57%), and WUE was significantly increased in RFPS (44.04%) and in FPP (37.50%) compared with CK (P < 0.05). We conclude that ridge-furrow planting with plastic film-mulched ridge and straw-mulched furrow has a good potential for raising winter wheat production on the Loess Plateau.

Treatment with PD-1 inhibitors can be hampered by severe auto-immune-related toxicities. Our objective was to identify single-nucleotide polymorphisms (SNPs) in genes previously associated with auto-immunity, which are associated with toxicities in nivolumab-treated NSCLC patients. This was in order to identify patients prone to develop severe toxicities and to gain more insight into the underlying pathobiology.