Assembly of a pangenome for global cattle reveals missing sequences and novel structural variations, providing new insights into their diversity and evolutionary history.

A cattle pangenome representation was created based on the genome sequences of 898 cattle representing 57 breeds. The pangenome identified 83 Mb of sequence not found in the cattle reference genome, representing 3.1% novel sequence compared with the 2.71-Gb reference. A catalog of structural variants developed from this cattle population identified 3.3 million deletions, 0.12 million inversions, and 0.18 million duplications. Estimates of breed ancestry and hybridization between cattle breeds using insertion/deletions as markers were similar to those produced by single nucleotide polymorphism-based analysis. Hundreds of deletions were observed to have stratification based on subspecies and breed. For example, an insertion of a Bov-tA1 repeat element was identified in the first intron of the APPL2 gene and correlated with cattle breed geographic distribution. This insertion falls within a segment overlapping predicted enhancer and promoter regions of the gene, and could affect important traits such as immune response, olfactory functions, cell proliferation, and glucose metabolism in muscle. The results indicate that pangenomes are a valuable resource for studying diversity and evolutionary history, and help to delineate how domestication, trait-based breeding, and adaptive introgression have shaped the cattle genome.

Expression and prognostic value of APOBEC2 in gastric adenocarcinoma and its association with tumor-infiltrating immune cells.

BACKGROUND: Apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 2 (APOBEC2) is associated with nucleotide alterations in the transcripts of tumor-related genes which are contributed to carcinogenesis. Expression and prognosis value of APOBEC2 in stomach adenocarcinoma (STAD) remains unclear. METHODS: The APOBEC2 gene alteration frequency of STAD and APOBEC2 gene expression in STAD and normal tissues were investigated in cBioportal and GEPIA, respectively. We detected expression of APOBEC2, infiltration of CD66b+ tumor-associated neutrophils and CD163+ tumor-associated macrophages in tissue microarrays by immunohistochemistry. APOBEC2 gene expression was explored by western blot and qRT-PCR. Relationships between APOBEC2 and CD66b, CD163, and other clinicopathological characteristics were investigated. Associations among APOBEC2 expression status and patient survival outcome were further analyzed. RESULTS: APOBEC2 gene alteration frequency was 5%, and APOBEC2 gene was downexpressed in STAD compared to normal tissues (P < 0.05). APOBEC2 expression status were associated with the infiltration of CD66b+ TANs, differentiation grade, TNM stage, histological type and gender (all P < 0.05) in STAD. Little or no APOBEC2 expression was detected in STAD and adjacent normal tissues by western blot. We failed to show that APOBEC2 was an independent risk factor for OS (Hazard Ratio 0.816, 95%CI 0.574-1.161, P = 0.259) or DFS (Hazard Ratio 0.821, 95%CI 0.578-1.166, P = 0.270) in STAD by multivariate Cox regression analysis, but APOBEC2 negative subgroup has a worse OS and DFS among patients with adjuvant chemotherapy. CONCLUSIONS: APOBEC2 correlates with CD66b, differentiation grade, TNM stages, histological classification, and gender in STAD. APOBEC2 is not an independent prognostic factor for STAD, our results suggest that patients with positive APOBEC2 can benefit from postoperative chemotherapy, and combination of APOBEC2 and CD66b is helpful to further stratify patients into different groups with distinct prognoses.

Gut microbiome variations in Rhinopithecus roxellanae caused by changes in the environment.

BACKGROUND: The snub-nosed monkey (Rhinopithecus roxellanae) is an endangered animal species mainly distributed in China and needs to be protected. Gut microbiome is an important determinant of animal health and population survival as it affects the adaptation of the animals to different foods and environments under kinetic changes of intrinsic and extrinsic factors. Therefore, this study aimed to elucidate gut fecal microbiome profiles of snub-nosed monkeys affected by several extrinsic and intrinsic factors, including raising patterns (captive vs. wild), age, sex, and diarrheal status to provide a reference for making protection strategies. RESULTS: The 16S rRNA gene sequencing was firstly used to pre-check clustering of 38 fecal samples from the monkeys including 30 wild and 8 captive (5 healthy and 3 diarrheal) from three Regions of Shennongjia Nature Reserve, Hubei Province, China. Then the 24 samples with high-quality DNA from 18 wild and 6 captive (4 healthy and 2 diarrheal) monkeys were subjected to shotgun metagenomic sequencing to characterize bacterial gut microbial communities. We discovered that the raising pattern (captive and wild) rather than age and sex was the predominant factor attributed to gut microbiome structure and proportionality. Wild monkeys had significantly higher bacterial diversity and lower Bacteroidetes/Firmicutes ratios than captive animals. Moreover, the gut microbiomes in wild healthy monkeys were enriched for the genes involved in fatty acid production, while in captive animals, genes were enriched for vitamin biosynthesis and metabolism and amino acid biosynthesis from carbohydrate intermediates. Additionally, a total of 37 antibiotic resistant genes (ARG) types were detected. Unlike the microbiome diversity, the captive monkeys have a higher diversity of ARG than the wild animals. CONCLUSION: Taken together, we highlight the importance of self-reprogramed metabolism in the snub-nosed monkey gut microbiome to help captive and wild monkeys adapt to different intrinsic and extrinsic environmental change.

Unique Fluorescent Protein-Encoded Microbeads with Supramaximal Encoding Capacity and High Sensitivity for Multiplex Bioassays.

Fluorescence-encoded microbeads (FEBs) have been widely used as a critical component in multiplexed biomolecular assays. Here, we propose a simple, sustainable, low-cost, and safe strategy for preparing FEBs by assembling fluorescent proteins (FPs) onto magnetic microbeads (MBs) via chemical coupling. Combining the type of FP, the concentration of FP, and the size of the magnetic microbeads as encoding elements, an ultralarge encoding capacity with 506 barcodes was obtained. We demonstrate that the FP-based FEBs have good stability during long-term storage and tolerate the use of an organic solution. Multiplex detection of femtomolar ssDNA molecules was achieved via flow cytometry, and the detection procedure is simple and fast because it does not require amplification and washing strategies. The advantages of this advanced method for multiplex detections including high sensitivity, specificity, accuracy, repeatability, rapidity, and cost-effectiveness show a broad application prospect in basic and applied research fields such as disease diagnosis, food safety, environmental protection, proteomics, genomics, and drug screening.

Soluble expression and purification of recombinant bovine ferritin H-chain.

Ferritin is a potential medicine delivery vehicle and vaccine platform, and its efficient expression is a prerequisite for widespread application. This study introduces a soluble expression strategy for recombinant bovine ferritin heavy chain (rFTH) in a prokaryotic system and an improved protein purification method. The amplified rFTH gene was ligated into the prokaryotic expression vector pET30a. The recombinant vectors with the N-terminal His-tag(N-His) or C-terminal His-tag(C-His) were translated and expressed separately. The results showed that the solubility of rFTH with C-His was significantly higher than that with N-His. The expression of rFTH with C-His was attempted at 37 °C and 16 °C, respectively. The results showed that the proportion of soluble protein expressed at 37 °C was more than 90%, higher than that expressed at 16 °C. Then rFTH with C-His was purified successfully using anion exchange chromatography, modified PEG precipitation, and dialysis. The rFTH protein was characterized using SDS-PAGE, Native-PAGE, Western blot, transmission electron microscopy, and dynamic light scattering. The results demonstrated that the purified rFTH protein self-assembled into ferritin nanoparticles with a regular shape and uniform size. This study sheds new light on the soluble expression of ferritin and provides a foundation for the construction of bovine ferritin nanoparticle production platforms.

Impact of health literacy and social support on medication adherence in patients with hypertension: a cross-sectional community-based study.

BACKGROUND: Previous studies have examined the associations of health literacy and social support with medication adherence among patients with hypertension. However, limited evidence exists regarding the mechanisms underlying the relationship between these factors and medication adherence. PURPOSE: To explore the prevalence of medication adherence and its determinants in patients with hypertension in Shanghai. METHODS: A community-based cross-sectional study was conducted among 1697 participants with hypertension. We collected sociodemographic and clinical characteristics as well as data regarding health literacy, social support, and medication adherence using questionnaires. We examined interactions among the factors using a structural equation model. RESULTS: The participants included 654 (38.54%) patients with a low degree of medication adherence and 1043 (61.46%) patients with a medium/high degree of adherence. Social support directly influenced adherence (ß = 0.165, P < 0.001) and indirectly influenced adherence through health literacy (ß = 0.087, P < 0.001). Health literacy directly influenced adherence (ß = 0.291, P < 0.001). Education indirectly affected adherence through both social support (ß = 0.048, P < 0.001) and health literacy (ß = 0.080, P < 0.001). Moreover, there was a sequential mediating effect of social support and health literacy on the association between education and adherence (ß = 0.025, P < 0.001). After controlling for age and marital status, similar results were also obtained, indicating a good model fit. CONCLUSIONS: The degree of medication adherence among hypertensive patients needs to improve. Health literacy and social support had both direct and indirect effects on adherence, and thus, these factors should be considered as tools to improve adherence.

Identification of Differential Circular RNA Expression Profiles and Functional Networks in Human Macrophages Induced by Virulent and Avirulent Mycobacterium tuberculosis Strains.

Circular RNAs (circRNAs) are noncoding RNAs with diverse functions. However, most Mycobacterium tuberculosis (M.tb)-related circRNAs remain undiscovered. In this study, we infected THP-1 cells with virulent and avirulent M.tb strains and then sequenced the cellular circRNAs. Bioinformatic analysis predicted 58,009 circRNAs in all the cells. In total, 2035 differentially expressed circRNAs were identified between the M.tb-infected and uninfected THP-1 cells and 1258 circRNAs were identified in the virulent and avirulent M.tb strains. Further, the top 10 circRNAs were confirmed by Sanger sequencing, among which four circRNAs, namely circSOD2, circCHSY1, circTNFRSF21, and circDHTKD1, which were highly differentially expressed in infected cells compared with those in uninfected cells, were further confirmed by ring formation, specific primers, and RNase R digestion. Next, circRNA-miRNA-mRNA subnetworks were constructed, such as circDHTKD1/miR-660-3p/IL-12B axis. Some of the individual downstream genes, such as miR-660-3p and IL-12B, were previously reported to be associated with cellular defense against pathological processes induced by M.tb infection. Because macrophages are important immune cells and the major host cells of M.tb, these findings provide novel ideas for exploring the M.tb pathogenesis and host defense by focusing on the regulation of circRNAs during M.tb infection.

An emerging role for cyclic dinucleotide phosphodiesterase and nanoRNase activities in Mycoplasma bovis: Securing survival in cell culture.

Mycoplasmas are host-restricted prokaryotes with a nearly minimal genome. To overcome their metabolic limitations, these wall-less bacteria establish intimate interactions with epithelial cells at mucosal surfaces. The alarming rate of antimicrobial resistance among pathogenic species is of particular concern in the medical and veterinary fields. Taking advantage of the reduced mycoplasma genome, random transposon mutagenesis was combined with high-throughput screening in order to identify key determinants of mycoplasma survival in the host-cell environment and potential targets for drug development. With the use of the ruminant pathogen Mycoplasma bovis as a model, three phosphodiesterases of the DHH superfamily were identified as essential for the proliferation of this species under cell culture conditions, while dispensable for axenic growth. Despite a similar domain architecture, recombinant Mbov_0327 and Mbov_0328 products displayed different substrate specificities. While rMbovP328 protein exhibited activity towards cyclic dinucleotides and nanoRNAs, rMbovP327 protein was only able to degrade nanoRNAs. The Mbov_0276 product was identified as a member of the membrane-associated GdpP family of phosphodiesterases that was found to participate in cyclic dinucleotide and nanoRNA degradation, an activity which might therefore be redundant in the genome-reduced M. bovis. Remarkably, all these enzymes were able to convert their substrates into mononucleotides, and medium supplementation with nucleoside monophosphates or nucleosides fully restored the capacity of a Mbov_0328/0327 knock-out mutant to grow under cell culture conditions. Since mycoplasmas are unable to synthesize DNA/RNA precursors de novo, cyclic dinucleotide and nanoRNA degradation are likely contributing to the survival of M. bovis by securing the recycling of purines and pyrimidines. These results point toward proteins of the DHH superfamily as promising targets for the development of new antimicrobials against multidrug-resistant pathogenic mycoplasma species.

Prevalence of bovine astroviruses and their genotypes in sampled Chinese calves with and without diarrhoea.

Bovine astrovirus (BoAstV) belongs to genus Mamastravirus (MAstV). It can be detected in the faeces of both diarrhoeal and healthy calves. However, its prevalence, genetic diversity, and association with cattle diarrhoea are poorly understood. In this study, faecal samples of 87 diarrhoeal and 77 asymptomatic calves from 20 farms in 12 provinces were collected, and BoAstV was detected with reverse transcription-polymerase chain reaction (RT-PCR). The overall prevalence rate of this virus in diarrhoeal and asymptomatic calves was 55.17 % (95 % CI: 44.13, 65.85 %) and 36.36 % (95 % CI: 25.70, 48.12 %), respectively, indicating a correlation between BoAstV infection and calf diarrhoea (OR=2.15, P=0.024). BoAstV existed mainly in the form of co-infection (85.53 %) with one to five of nine viruses, and there was a strong positive correlation between BoAstV co-infection and calf diarrhoea (OR=2.83, P=0.004). Binary logistic regression analysis confirmed this correlation between BoAstV co-infection and calf diarrhoea (OR=2.41, P=0.038). The co-infection of BoAstV and bovine rotavirus (BRV) with or without other viruses accounted for 70.77 % of all the co-infection cases. The diarrhoea risk for the calves co-infected with BoAstV and BRV was 8.14-fold higher than that for the calves co-infected with BoAstV and other viruses (OR=8.14, P=0.001). Further, the co-infection of BoAstV/BRV/bovine kobuvirus (BKoV) might increase the risk of calf diarrhoea by 14.82-fold, compared with that of BoAstV and other viruses (OR=14.82, P <0.001). Then, nearly complete genomic sequences of nine BoAstV strains were assembled by using next-generation sequencing (NGS) method. Sequence alignment against known astrovirus (AstV) strains at the levels of both amino acids and nucleotides showed a high genetic diversity. Four genotypes were identified, including two known genotypes MAstV-28 (n=3) and MAstV-33 (n=2) and two novel genotypes designated tentatively as MAstV-34 (n=1) and MAstV-35 (n=3). In addition, seven out of nine BoAstV strains showed possible inter-genotype recombination and cross-species recombination. Therefore, our results increase the knowledge about the prevalence and the genetic evolution of BoAstV and provide evidence for the association between BoAstV infection and calf diarrhoea.

Design of bovine lactoferricin-derived peptide and its expression and activity in Pichia pastoris.

Bovine lactoferrin peptide has been shown to be a broad-spectrum antimicrobial peptide. Based on the relationship between the structure and function of antimicrobial peptides, the antimicrobial peptide databases and protein analysis software were used to optimize the design of bovine lactoferricin peptide (LfcinB). The designed bovine lactoferricin-derived peptide (LfcinBD) gene fragment was inserted into a pPIC9K-His plasmid to construct a recombinant expression vector. After linearization of the Recombinant plasmid, Pichia pastoris GS115 cells were transfected with linearized recombinant plasmid by using electroporation and LfcinBD gene expression was induced with methanol. After the fermentation, supernatant was separated by low-temperature high-speed centrifugation. Ultrafiltration and freeze drying of the fermentation supernatant were performed, purified. Experimental results showed that the LfcinBD had stronger bacteriostatic activity against Staphylococcus aureus than the natural bovine lactoferrin peptide (LfcinB) produced under the same fermentation conditions. The effective expression of the optimized bovine lactoferricin-derived peptide was detected using SDS-PAGE electrophoresis. This study lays the foundation for further exploration to improve the biological activities of antimicrobial peptides.

Genome-Wide Analysis of LncRNA in Bovine Mammary Epithelial Cell Injuries Induced by Escherichia Coli and Staphylococcus Aureus.

Escherichia coli and Staphylococcus aureus are two common pathogenic microorganisms that cause mastitis in dairy cows. They can cause clinical mastitis and subclinical mastitis. In recent studies, lncRNAs have been found to play an important role in the immune responses triggered by microbial inducers. However, the actions of lncRNAs in bovine mastitis remain unclear. The purpose of this study was to investigate the effects of bovine mammary epithelial cell injuries induced by treatment with E. coli and S. aureus, and to explore the lncRNA profile on cell injuries. The lncRNA transcriptome analysis showed a total of 2597 lncRNAs. There were 2234 lncRNAs differentially expressed in the E. coli group and 2334 in the S. aureus group. Moreover, we found that the E. coli and S. aureus groups of maternal genes targeted signaling pathways with similar functions according to KEGG and GO analyses. Two lncRNA-miRNA-mRNA interaction networks were constructed in order to predict the potential molecular mechanisms of regulation in the cell injuries. We believe that this is the first report demonstrating the dysregulation of lncRNAs in cells upon E. coli and S. aureus infections, suggesting that they have the potential to become important diagnostic markers and to provide novel insights into controlling and preventing mastitis.

Identification of Unique Key miRNAs, TFs, and mRNAs in Virulent MTB Infection Macrophages by Network Analysis.

Although Mycobacterium tuberculosis (MTB) has existed for thousands of years, its immune escape mechanism remains obscure. Increasing evidence signifies that microRNAs (miRNAs) play pivotal roles in the progression of tuberculosis (TB). RNA sequencing was used to sequence miRNAs in human acute monocytic leukemia cells (THP-1) infected by the virulent MTB-1458 strain and the avirulent vaccine strain Mycobacterium bovis Bacillus Calmette-Guérin (BCG). Sets of differentially expressed miRNAs (DE-miRNAs) between MTB-1458/BCG-infected groups and uninfected groups were identified, among which 18 were differentially expressed only in the MTB-1458-infected THP-1 group. Then, 13 transcription factors (TFs) and 81 target genes of these 18 DE-miRNAs were matched. Gene Ontology classification as well as Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that the candidate targets were predominantly involved in apoptotic-associated and interferon-γ-mediated signaling pathways. A TF-miRNA-mRNA interaction network was constructed to analyze the relationships among these 18 DE-miRNAs and their targets and TFs, as well as display the hub miRNAs, TFs, and target genes. Considering the degrees from network analysis and the reported functions, this study focused on the BHLHE40-miR-378d-BHLHE40 regulation axis and confirmed that BHLHE40 was a target of miR-378d. This cross-talk among DE-miRNAs, mRNAs, and TFs might be an important feature in TB, and the findings merited further study and provided new insights into immune defense and evasion underlying host-pathogen interactions.

Transcriptome Profiling of m6A mRNA Modification in Bovine Mammary Epithelial Cells Treated with Escherichia coli.

Mastitis is a common disease in dairy cows that is mostly caused by E. coli, and it brings massive losses to the dairy industry. N6-Methyladenosine (m6A), a methylation at the N6 position of RNA adenine, is a type of modification strongly associated with many diseases. However, the role of m6A in mastitis has not been investigated. In this study, we used MeRIP-seq to sequence the RNA of bovine mammary epithelial cells treated with inactivated E. coli for 24 h. In this in vitro infection model, there were 16,691 m6A peaks within 7066 mRNA transcripts in the Con group and 10,029 peaks within 4891 transcripts in the E. coli group. Compared with the Con group, 474 mRNAs were hypermethylated and 2101 mRNAs were hypomethylated in the E. coli group. Biological function analyses revealed differential m6A-modified genes mainly enriched in the MAPK, NF-κB, and TGF-ß signaling pathways. In order to explore the relationship between m6A and mRNA expression, combined MeRIP-seq and mRNA-seq analyses revealed 212 genes with concomitant changes in the mRNA expression and m6A modification. This study is the first to present a map of RNA m6A modification in mastitis treated with E. coli, providing a basis for future research.

Differential nitric oxide induced by Mycobacterium bovis and BCG leading to dendritic cells apoptosis in a caspase dependent manner.

Dendritic cells (DCs) are critical for both innate and adaptive immunity. Meanwhile, nitric oxide (NO) is a member of reactive nitrogen species (RNS) generally considered to play a key role in the bactericidal process in innate immunity against Mycobacterium tuberculosis complex infection. The present study therefore investigated the mechanism of NO production in murine DCs induced by Mycobacterium bovis (M.bovis) and its attenuated strain Bacillus Calmette-Guérin (BCG) infection. The expression of genes Slc7A1, Slc7A2, iNOS, and ArgI essential to NO synthesis was up-regulated in M.bovis/BCG infected DCs. IFN-γ addition further increased, while the iNOS inhibitor L-NMMA significantly inhibited their expression. Accordingly, the end products of arginine metabolism, NO and urea, were found to be significantly increased. In addition, BCG induced significantly higher levels of apoptosis in DCs compared to M.bovis shown by higher levels of DNA fragmentation using flow cytometry and release of mitochondrial Cytochrome C, and up-regulation of the genes caspase-3, caspase-8, caspase-9 and dffa critical to apoptosis by qRT-PCR detection and western blot analysis. Furthermore, IFN-γ increased, but L-NMMA decreased apoptosis of M.bovis/BCG infected DCs. In addition, mycobacterial intracellular survival was significantly reduced by IFN-γ treatment in BCG infected DCs, while slightly increased by L-NMMA treatment. Taken altogether, our data show that NO synthesis was differentially increased and associated with apoptosis in M.bovis/BCG infected DCs. These findings may significantly contribute to elucidate the pathogenesis of M.bovis.

TRIM25 upregulation by Mycobacterium tuberculosis infection promotes intracellular survival of M.tb in RAW264.7 cells.

Tripartite motif 25 (TRIM25) is a TRIM family member which is involved in innate immunity. However, its role in the modulation of host defense against Mycobacterium tuberculosis (M.tb) infection has not been investigated. Therefore, this study aimed to demonstrate the significance of TRIM25 in the regulation of macrophage responses to M.tb infection. TRIM25 was found to be significantly overexpressed (3.476-fold) in peripheral blood mononuclear cells (PBMCs) of 67 patients with pulmonary tuberculosis compared with 48 healthy controls. TRIM25 expression was enhanced following M.tb infection of RAW264.7 cells, a macrophage cell line. Overexpression of TRIM25 in M.tb-infected RAW264.7 cells led to a significant increase in phosphorylated p38 levels; however, the production of IL-6, IL-1ß, and TNF-α were significantly reduced. Finally, M.tb intracellular survival increased by 90% at 12 h post-infection (PI) (p < 0.01). To validate the previous results, TRIM25 levels in M.tb-infected RAW264.7 macrophages were down-regulated using small interfering RNA (siRNA). Therefore, it was concluded that TRIM25 promotes intracellular survival of M.tb in RAW264.7 cells, likely by enhancing p38 pathways and thereby inhibiting the production of proinflammatory cytokines. These results contribute to the further understanding of the host defense against M.tb infection.

Progresses on bacterial secretomes enlighten research on Mycoplasma secretome.

Bacterial secretome is a comprehensive catalog of bacterial proteins that are released or secreted outside the cells. They offer a number of factors that possess several significant roles in virulence as well as cell to cell communication and hence play a core role in bacterial pathogenesis. Sometimes these proteins are bounded with membranes giving them the shape of vesicles called extracellular vesicles (EVs) or outer membrane vesicles (OMVs). Bacteria secrete these proteins via Sec and Tat pathways into the periplasm. Secreted proteins have found to be important as diagnostic markers as well as antigenic factors for the development of an effective candidate vaccine. Recently, the research in the field of secretomics is growing up and getting more interesting due to their direct involvement in the pathogenesis of the microorganisms leading to the infection. Many pathogenic bacteria have been studied for their secretome and the results illustrated novel antigens. This review highlights the secretome studies of different pathogenic bacteria in humans and animals, general secretion mechanisms, different approaches and challenges in the secretome of Mycoplasma sp.

Identification of 60 secreted proteins for Mycoplasma bovis with secretome assay.

Mycoplasma bovis is a risky pathogen mainly responsible for pneumonia and mastitis in cattle. Up to date, its pathogenesis is not clear. Since secreted proteins have a tricky role in M. bovis pathogenesis, this study was designed to systematically reveal M. bovis secretome and potential role in virulence of the pathogen. By using bioinformatics tools, a total of 246 secreted proteins were predicted based on M. bovis genome. Among them, 14 were classical, 154 non-classical and 78 both pathways. Then by using 2-dimensional gel electrophoresis (2-DE) and Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF- MS), 169 proteins were revealed. Of them, 60 were predicted to be secreted including 3 classical, 43 non-classical, and 14 both classical and non-classical. Further 8 proteins (MbovP0038, MbovP0338, MbovP0341, MbovP0520, MbovP0581, MbovP0674, MbovP0693, MbovP0845) were predicted to be virulence-related factors with VFDB. In addition, MbovP0581 (ABC transporter protein) was validated experimentally as secreted in nature and highly immunogenic reacting with sera of cattle experimentally infected with M. bovis. In conclusion, this study might be a crucial step towards a better understanding of pathogenesis and leading to the development of novel diagnostic marker and potent vaccine against M. bovis.

Clinical significance and biological function of WD repeat domain 54 as an oncogene in colorectal cancer.

In recent years, protein-protein interactions have become an attractive candidate for identifying biomarkers and drug targets for various diseases. However, WD40 repeat (WDR) domain proteins, some of the most abundant mediators of protein interactions, are largely unexplored. In our study, 57 of 361 known WDR proteins were identified as hub nodes, and a hub (WDR54) with elevated mRNA in colorectal cancer (CRC) was selected for further study. Immunohistochemistry of specimens from 945 patients confirmed the elevated expression of WDR54 in CRC, and we found that patients with WDR54-high tumors typically had a shorter disease-specific survival (DSS) than those with WDR54-low tumors, especially for the subgroup without well-differentiated tumors. Multivariate analysis showed that WDR54-high tumors were an independent risk factor for DSS, with a hazard ratio of 2.981 (95% confidence interval, 1.425-6.234; p = 0.004). Knockdown of WDR54 significantly inhibited the growth and aggressiveness of CRC cells and reduced tumor growth in a xenograft model. Each WDR54 isoform (a, b, and c) was found to reverse the inhibitory effect of WDR54 knockdown; however, only isoform c, which exhibited the highest expression, was increased in CRC cells. Sensitization of WDR54 knockdown to an SHP2 inhibitor was consistently found in CRC cells, and the underlying mechanism involved their common function in regulating AKT and ERK signaling. In conclusion, the present study is the first to investigate the significance of WDR54 in cancer and to conclude that WDR54 serves as an oncogene in CRC and may be a potential prognostic marker and therapeutic target.

Diversity and distribution of type A influenza viruses: an updated panorama analysis based on protein sequences.

BACKGROUND: Type A influenza viruses (IAVs) cause significant infections in humans and multiple species of animals including pigs, horses, birds, dogs and some marine animals. They are of complicated phylogenetic diversity and distribution, and analysis of their phylogenetic diversity and distribution from a panorama view has not been updated for multiple years. METHODS: 139,872 protein sequences of IAVs from GenBank were selected, and they were aligned and phylogenetically analyzed using the software tool MEGA 7.0. Lineages and subordinate lineages were classified according to the topology of the phylogenetic trees and the host, temporal and spatial distribution of the viruses, and designated using a novel universal nomenclature system. RESULTS: Large phylogenetic trees of the two external viral genes (HA and NA) and six internal genes (PB2, PB1, PA, NP, MP and NS) were constructed, and the diversity and the host, temporal and spatial distribution of these genes were calculated and statistically analyzed. Various features regarding the diversity and distribution of IAVs were confirmed, revised or added through this study, as compared with previous reports. Lineages and subordinate lineages were classified and designated for each of the genes based on the updated panorama views. CONCLUSIONS: The panorama views of phylogenetic diversity and distribution of IAVs and their nomenclature system were updated and assumed to be of significance for studies and communication of IAVs.

P27 (MBOV_RS03440) is a novel fibronectin binding adhesin of Mycoplasma bovis.

Mycoplasma bovis, one of the major pathogens of bovine respiratory disease, binds to respiratory epithelial cells resulting in severe pneumonia and tissue damage. This study was designed to identify the adhesive function of a putative 27-kDa M. bovis lipoprotein, encoded by the gene MBOV_RS03440 and designated as P27. The gene was cloned and overexpressed to produce antibodies against the recombinant P27 (rP27). The western blot and flow cytometry assay confirmed P27 to be a surface-localized protein, while ELISA confirmed it to be an immunogenic protein. Confocal immunofluorescence microscopy demonstrated that rP27 bound to embryonic bovine lung (EBL) cell monolayers in a dose-dependent manner. Furthermore, anti-rP27 antiserum inhibited the attachment of M. bovis to EBL cells demonstrating the binding specificity of P27 to EBL cells. The attachment of rP27 to EBL cells was mediated by fibronectin (Fn), an extracellular matrix component. The interaction between rP27 and Fn was qualitatively and quantitatively monitored by ligand immunoblot assay, ELISA, and biolayer interferometry. Collectively, these results indicate that P27 is a novel Fn-binding, immunogenic adhesive protein of M. bovis, thereby contributing to the further understanding of the molecular pathogenesis of M. bovis.