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    细菌耐药性及兽药相关Bacterial resistance & Veterinary drug

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    The vital role of CovS in the establishment of Streptococcus equi subsp. zooepidemicus virulence
    XU Bin, MA Zhe, ZHOU Hong, LIN Hui-xing, FAN Hong-jie
    2023, 22 (2): 568-584.   DOI: 10.1016/j.jia.2022.08.109
    Abstract331)      PDF in ScienceDirect      

    Streptococcus equi subsp. zooepidemicus (SEZ) is an important zoonotic agent.  Here, a virulence-attenuated strain M35246 derived from natural variation of wild-type SEZ ATCC35246 was found.  M35246 showed a deletion of 25 contiguous genes as well as a loss-of-function mutation in covS.  Subsequently, a 25-gene-deleted strain (ΔPI), a covS-mutant strain (McovS), and relevant complementary strains were constructed and investigated.  M35246 and McovS were significantly less encapsulated and exhibited poorer anti-phagocytic capacity compared to wild-type SEZ.  McovS was significantly more sensitive to β-lactams, aminoglycosides, macrolides, and lincosamides than wild-type SEZ.  M35246, McovS, and ΔPI exhibited an increase in median lethal dose (LD50) in mice by 105, 105, and 5 times when compared to wild-type SEZ, respectively.  Neither M35246 nor McovS were isolated from mice 48 h after being challenged with approximately 2 000 times the LD50 of wild-type SEZ.  Transcriptome analysis showed that 668 significantly differentially expressed genes existed between McovS and wild-type SEZ.  Numerous virulence factor-encoding genes and anabolic-related genes in McovS that were involved in anti-phagocytosis, capsule formation, pathogenicity, and antibiotic resistance were downregulated significantly relative to the wild-type strain.  This study revealed that the CovS plays a vital role in the establishment of SEZ virulence

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    Characterization of a blaCTX-M-3, blaKPC-2 and blaTEM-1B co-producing IncN plasmid in Escherichia coli of chicken origin

    WANG Wen-jing, WANG Yi-fu, JIN Ya-jie, SONG Wu-qiang, LIN Jia-meng, ZHANG Yan, TONG Xin-ru, TU Jian, LI Rui-chao, LI Tao
    2023, 22 (1): 320-324.   DOI: 10.1016/j.jia.2022.08.075
    Abstract368)      PDF in ScienceDirect      

    An extensively drug-resistant (XDR) Escherichia coli strain 258E was isolated from an anal swab sample of a chicken farm of Anhui province in China. Genomic analyses indicated that the strain 258E harbors an incompatibility group N (IncN) plasmid pEC258-3, which co-produces blaCTX-M-3, blaKPC-2, blaTEM-1B, qnrS1, aac(6')-Ib-cr, dfrA14, arr-3, and aac(6')-Ib3. Multiple genome arrangement analyses indicated that pEC258-3 is highly homologous with pCRKP-1-KPC discovered in Klebsiella pneumoniae from a patient. Furthermore, conjugation experiments proved that plasmid pEC258-3 can be transferred horizontally and may pose a significant potential threat in animals, community and hospital settings.

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    Molecular epidemiology, characterization of virulence factors and antibiotic-resistance profile of Streptococcus agalactiae isolated from dairy farms in China and Pakistan
    Ambreen LEGHARI, Shakeel Ahmed LAKHO, Faiz Muhammad KHAND, Khaliq ur Rehman BHUTTO, Sameen Qayoom LONE, Muhammad Tahir ALEEM, Iqra BANO, Muhammad Ali CHANDIO, Jan Muhammad SHAH, LIN Hui-xing, FAN Hong-jie
    2023, 22 (5): 1514-1528.   DOI: 10.1016/j.jia.2022.10.004
    Abstract260)      PDF in ScienceDirect      

    Streptococcus agalactiae is one of the most common pathogens that cause bovine mastitis worldwide. Identifying pathogen prevalence and virulence factors is critical for developing prevention and control approaches. Herein, 1161 milk samples from various dairy farms in China (n=558) and Pakistan (n=603) were collected between 2019-2021 and were subjected to S. agalactiae isolation. Prevalence, serotyping, virulence genes, and antibiotic-resistant genes of S. agalactiae were evaluated by PCR assay. All isolates were characterized for haemolysis, biofilm production, cytotoxicity, adhesion, and invasion on bovine mammary epithelial cells. The prevalence of S. agalactiae-induced mastitis in cattle was found to be considerably higher in Pakistan than in China. Jiangsu and Sindh provinces had the highest area-wise prevalence in China and Pakistan, respectively. Serotypes Ia and II were prevalent in both countries, whereas serotype III was found only in Pakistan. Moreover, all isolates tested positive for PI-2b gene but negative for PI-1 and PI-2a genes. All isolates harboured cfb, cylE, hylB, and fbsB virulent genes, whereas many of them lacked bibA, rib and bca. However, the absence of bac and scp genes in Chinese isolates and cspA in Pakistani isolates was noted, while spb1 and lmb were not detected in isolates of both countries. Pakistani isolates, particularly serotype Ia-positive, had a considerably higher ability to produce biofilm, haemolysis, cytotoxicity, adhesion, and invasion than Chinese isolates. Most of the isolates were phenotypically resistant to tetracycline, erythromycin, and clindamycin and genotypic resistance was confirmed by the presence of ermA, ermB, tetM and tetO genes. Our study highlights the antimicrobial resistance profile and virulence-related factors contributing to the epidemiological spread of mastitis-causing S. agalactiae in China and Pakistan. The findings may facilitate future studies designed to develop improved treatment and control strategies against this pathogen. 

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    Insights into the effects of pulsed antimicrobials on the chicken resistome and microbiota from fecal metagenomes
    ZHAO Ruo-nan, CHEN Si-yuan, TONG Cui-hong, HAO Jie, LI Pei-si, XIE Long-fei, XIAO Dan-yu, ZENG Zhen-ling, XIONG Wen-guang
    2023, 22 (6): 1857-1869.   DOI: 10.1016/j.jia.2022.11.006
    Abstract252)      PDF in ScienceDirect      

    Antimicrobial resistance has become a global problem that poses great threats to human health.  Antimicrobials are widely used in broiler chicken production and consequently affect their gut microbiota and resistome.  To better understand how continuous antimicrobial use in farm animals alters their microbial ecology, we used a metagenomic approach to investigate the effects of pulsed antimicrobial administration on the bacterial community, antibiotic resistance genes (ARGs) and ARG bacterial hosts in the feces of broiler chickens.  Chickens received three 5-day courses of individual or combined antimicrobials, including amoxicillin, chlortetracycline and florfenicol.  The florfenicol administration significantly increased the abundance of mcr-1 gene accompanied by floR gene, while amoxicillin significantly increased the abundance of genes encoding the AcrAB-tolC multidrug efflux pump (marA, soxS, sdiA, rob, evgS and phoP).  These three antimicrobials all led to an increase in Proteobacteria.  The increase in ARG host, Escherichia, was mainly attributed to the β-lactam, chloramphenicol and tetracycline resistance genes harbored by Escherichia under the pulsed antimicrobial treatments.  These results indicated that pulsed antimicrobial administration with amoxicillin, chlortetracycline, florfenicol or their combinations significantly increased the abundance of Proteobacteria and enhanced the abundance of particular ARGs.  The ARG types were occupied by the multidrug resistance genes and had significant correlations with the total ARGs in the antimicrobial-treated groups.  The results of this study provide comprehensive insight into pulsed antimicrobial-mediated alteration of chicken fecal microbiota and resistome.

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    A novel short transcript isoform of chicken IRF7 negatively regulates interferon-β production
    MA Yu-chen, CHEN Hua-yuan, GAO Shen-yan, ZHANG Xiao-zhan, LI Yong-tao, YANG Xia, ZHAO Jun, WANG Zeng
    2023, 22 (7): 2213-2220.   DOI: 10.1016/j.jia.2022.12.015
    Abstract145)      PDF in ScienceDirect      
    Type I interferon (IFN-I) provides an important first line to protect avian species against pathogens invasion. IFN regulatory factor 7 (IRF7) has been identified as the most important regulator for both DNA and RNA virus-induced IFN-I production in chickens. Although four splicing variants of IRF7 have been identified in mammals, it is still unclear whether alternative splicing patterns and the function of IRF7 isoform(s) exist in chickens. In this study, we reported a novel short transcript isoform of chicken IRF7 (chIRF7), termed chIRF7-iso, which contained an intact N-terminal DNAbinding domain (DBD) and 14 amino acids different from chIRF7 in the C-terminal. Overexpression of chIRF7 in chicken leghorn male hepatocellular (LMH) cells activated the IFN-β promoter and significantly inhibited Newcastle disease virus (NDV) and fowl adenovirus serotype 4 (FAdV-4) replication. Conversely, overexpression of chIRF7-iso blocked the IFN-β promoter activity and was favorable for NDV and FAdV-4 replication in vitro. Collectively, our results confirm that a novel chIRF7 isoform-mediated negative regulates IFN-β production, which will contribute to understanding the role of chIRF7 in innate antiviral response in chicken.
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    Dynamic transcriptome profiles and novel markers in bovine spermatogenesis revealed by single-cell sequencing
    Yuan Gao, Fuxia Bai, Qi Zhang, Xiaoya An, Zhaofei Wang, Chuzhao Lei, Ruihua Dang
    2024, 23 (7): 2362-2378.   DOI: 10.1016/j.jia.2023.04.036
    Abstract239)      PDF in ScienceDirect      
    Testicular development is an important biological process in male and requires interaction between the male germ cells and somatic cells. However, the mechanisms of testicular development in livestock, particularly in cattle, are poorly understood. Furthermore, cellular heterogeneity hinders the profiling of different cell types at different developmental stages. In this study, we first performed a single-cell transcriptomic study of the bovine testis development during puberty by using 10× genomics single-cell RNA sequencing (scRNA-seq). By collecting the scRNA-seq data from 11,083 cells from prepubertal and pubertal bovine testes, a high-resolution scRNA-seq atlas was described, identifying 9 somatic and 13 spermatogenic clusters. We also distinguished several stage-specific marker genes for bovine germ cells and somatic cells, such as GRAF2 and MORC1 for SSC (spermatogonial stem cells), HJURP and TCF19 for differentiating spermatogonia, ARSE for immature Sertoli, CLEC12B for mature Sertoli, LOC112441470 for Leydig. In conclusion, we have examined the transcription levels and constructed the single-cell developmental maps of germ cells and somatic cells during testicular development in Angus cattle. The datasets provided new insights into spermatogenesis and testicular somatic cell development in cattle.
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    Identification of blaIMI-mediated carbapenem-resistant Enterobacter from a duck farm in China
    HUANG Hong-hao, LU Yi-xing, WU Su-juan, MA Zhen-bao, ZENG Dong-ping, ZENG Zhen-ling
    2023, 22 (8): 2500-2508.   DOI: 10.1016/j.jia.2023.06.013
    Abstract175)      PDF in ScienceDirect      

    Carbapenem- and colistin-resistant Enterobacter has been a clinical and therapy problem in recent years. Here, we report the carbapenem- and colistin-resistant Enterobacter harboring blaIMI isolated from intestinal samples and the environment of a duck farm in China. Four blaIMI-positive Enterobacter isolates were resistant to carbapenem and colistin. Three blaIMI subtypes were detected in different molecular categories of Enterobacter. The detection of the various IMI producers highlights the diversity of carbapenemases in a duck farm. Whole-genome sequencing demonstrated the blaIMI genes were present in chromosomes or plasmids in these strains. The conjugation experiment demonstrated the ability of blaIMI-carrying plasmid to transmit horizontally. The molecular evolution characteristics were examined through comparative genetic analysis. The study demonstrated the presence of chromosomal and plasmid blaIMI and the blaIMI-carrying plasmid exhibits a horizontal transmission between Enterobacter and Escherichia coli C600. The similar genetic content was discovered between two blaIMI-16-positive Enterobacter asburiae. In addition, a blaIMI-16-carrying plasmid is an IncFII(Yp) plasmid, and a substantial amount of mobile genetic elements were identified around blaIMI-16. The IS-like elements and IncFII(Yp) plasmid are significant in the propagation of blaIMI. Our study provides evidence for the transmission of diverse blaIMI genes in China and supplies additional reference data for blaIMI-positive antimicrobial-resistant Enterobacter. Routine surveys of blaIMI-positive Enterobacter from animal-raising environments must be given more focus

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    Characteristics of Mycobacterium tuberculosis serine protease Rv1043c in enzymology and pathogenicity in mice
    TANG Yang-yang, CUI Ying-ying, JIANG Yan-yan, SHAO Ming-zhu, ZANG Xin-xin, DANG Guang-hui, LIU Si-guo
    2023, 22 (12): 3755-3768.   DOI: 10.1016/j.jia.2023.06.025
    Abstract161)      PDF in ScienceDirect      

    The serine proteases of Mycobacteria tuberculosis (Mtb) are important contributors to the process of bacterial invasion and its pathogenesis.  In the present study, we systematically characterized the role of the Rv1043c protein in Mycobacterium infection by purifying the Rv1043c protein in Escherichia coli and constructing a Mycobacterium smegmatis (Msg) strain overexpressing Rv1043c (Msg_Rv1043c).  We found that Rv1043c had serine protease activity and localized to the surface of Mtb.  We determined that the optimal pH and temperature for the Rv1043c serine protease were 9.0 and 45°C, respectively.  Moreover, the serine protease activity of Rv1043c was enhanced by divalent metal ions of Ca2+ and Mg2+.  Site-directed mutagenesis studies demonstrated that the serine 279 residue in Rv1043c plays a catalytic role.  Additionally, mouse model studies confirmed that Rv1043c significantly enhanced the survival of Msg in vivo, induced pulmonary injury and lung cell apoptosis, and promoted the release of pro-inflammatory cytokines interleukin-1β and interleukin-6 in mice.  This study presents novel insights into the relationship between mycobacterial serine protease and the pathogenesis of the disease.

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    The virulence regulator AbsR in avian pathogenic Escherichia coli has pleiotropic effects on bacterial physiology

    Dongfang Zhao, Haobo Zhang, Xinyang Zhang, Fengwei Jiang, Yijing Li, Wentong Cai, Ganwu Li
    2024, 23 (2): 649-668.   DOI: 10.1016/j.jia.2023.07.035
    Abstract179)      PDF in ScienceDirect      

    Avian pathogenic Escherichia coli (APEC) belonging to extraintestinal pathogenic Ecoli (ExPEC) can cause severe infections in extraintestinal tissues in birds and humans, such as the lungs and blood.  MprA (microcin production regulation, locus A, herein renamed AbsR, a blood survival regulator), a member of the MarR (multiple antibiotic resistance regulator) transcriptional regulator family, governs the expression of capsule biosynthetic genes in human ExPEC and represents a promising druggable target for antimicrobials.  However, a deep understanding of the AbsR regulatory mechanism as well as its regulon is lacking.  In this study, we present a systems-level analysis of the APEC AbsR regulon using ChIP-Seq (chromatin immunoprecipitation sequencing) and RNA-Seq (RNA sequencing) methods.  We found that AbsR directly regulates 99 genes and indirectly regulates 667 genes.  Furthermore, we showed that: 1) AbsR contributes to antiphagocytotic effects by macrophages and virulence in a mouse model for systemic infection by directly activating the capsular gene cluster; 2) AbsR positively impacts biofilm formation via direct regulation of the T2SS (type II secretion system) but plays a marginal role in virulence; and 3) AbsR directly upregulates the acid tolerance signaling system EvgAS to withstand acid stress but is dispensable in ExPEC virulence.  Finally, our data indicate that the role of AbsR in virulence gene regulation is relatively conserved in ExPEC strains.  Altogether, this study provides a comprehensive analysis of the AbsR regulon and regulatory mechanism, and our data suggest that AbsR likely influences virulence primarily through the control of capsule production.  Interestingly, we found that AbsR severely represses the expression of the type I-F CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR associated) systems, which could have implications in CRISPR biology and application.

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    Membrane vesicles derived from Streptococcus suis serotype 2 induce cell pyroptosis in endothelial cells via the NLRP3/Caspase-1/GSDMD pathway

    Keda Shi, Yan Li, Minsheng Xu, Kunli Zhang, Hongchao Gou, Chunling Li, Shaolun Zhai
    2024, 23 (4): 1338-1353.   DOI: 10.1016/j.jia.2023.09.022
    Abstract210)      PDF in ScienceDirect      
    Streptococcus suis serotype 2 (S. suis 2) is a zoonotic pathogen that clinically causes severe swine and human infections (such as meningitis, endocarditis, and septicemia).  In order to cause widespread diseases in different organs, S. suis 2 must colonize the host, break the blood barrier, and cause exaggerated inflammation.  In the last few years, most studies have focused on a single virulence factor and its influences on the host.  Membrane vesicles (MVs) can be actively secreted into the extracellular environment contributing to bacteria-host interactions.  Gram-negative bacteria-derived outer membrane vesicles (OMVs) were recently shown to activate host Caspase-11-mediated non-canonical inflammasome pathway via deliverance of OMV-bound lipopolysaccharide (LPS), causing host cell pyroptosis.  However, little is known about the effect of the MVs from S. suis 2 (Gram-positive bacteria without LPS) on cell pyroptosis.  Thus, we investigated the molecular mechanism by which S. suis 2 MVs participate in endothelial cell pyroptosis.  In this study, we used proteomics, electron scanning microscopy, fluorescence microscope, Western blotting, and bioassays, to investigate the MVs secreted by S. suis 2.  First, we demonstrated that S. suis 2 secreted MVs with an average diameter of 72.04 nm, and 200 proteins in MVs were identified.  Then, we showed that MVs were transported to cells via mainly dynamin-dependent endocytosis.  The S. suis 2 MVs activated NLRP3/Caspase-1/GSDMD canonical inflammasome signaling pathway, resulting in cell pyroptosis, but it did not activate the Caspase-4/-5 pathway.  More importantly, endothelial cells produce large amounts of reactive oxygen species (ROS) and lost their mitochondrial membrane potential under induction by S. suis 2 MVs.  The results in this study suggest for the first time that MVs from S. suis 2 were internalized by endothelial cells via mainly dynamin-dependent endocytosis and might promote NLRP3/Caspase-1/GSDMD pathway by mitochondrial damage, which produced mtDNA and ROS  under induction, leading to the pyroptosis of endothelial cells.
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    First identification of the oxazolidinone/phenicol resistance gene optrA in Streptococcus pluranimalium worldwide
    Kuan Zhao, Longyu Zhou, Shixia Zhang, Wanjiang Zhang, Yao Zhu
    2024, 23 (2): 731-734.   DOI: 10.1016/j.jia.2023.11.042
    Abstract168)      PDF in ScienceDirect      
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    Identification of a novel multi-drug resistant plasmid co-harbouring extended-spectrum β-Lactamase resistance genes blaPER-4 and blaOXA-10 in Moellerella wisconsensis of sheep
    Xueliang Zhao, Yongqiang Miao, Hongmei Chen, Honghu Shan, Juan Wang, Yang Wang, Jianzhong Shen, Zengqi Yang
    2024, 23 (9): 3238-3242.   DOI: 10.1016/j.jia.2024.03.040
    Abstract106)      PDF in ScienceDirect      
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    Emergence of a novel multi-resistance-mediating integrative and conjugative element ICEPmu3 in Pasteurella multocida
    Jiao He, Zhishuang Yang, Mingshu Wang, Renyong Jia, Shun Chen, Mafeng Liu, Xinxin Zhao, Qiao Yang, Ying Wu, Shaqiu Zhang, Juan Huang, Xumin Ou, Di Sun, Bin Tian, Yu He, Zhen Wu, Anchun Cheng, Dekang Zhu
    2024, 23 (11): 3938-3942.   DOI: 10.1016/j.jia.2024.07.008
    Abstract134)      PDF in ScienceDirect      
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