川西北高原牦牛和藏猪源大肠杆菌生物被膜表型、耐药基因、整合酶基因和毒力基因检测  

Biofilm-Forming Phenotype,Antibacterial Resistance Genes,Integrase Genes and Virulence Genes Detection of Escherichia coli Isolated from Yaks and Tibetan Pigs in Northwest Sichuan Plateau

作　　者：陈朝喜[1] 李宇涵 谭敏 汪露 黄志宏[1] CHEN ChaoXi;LI YuHan;TAN Min;WANG Lu;HUANG ZhiHong(College of Animal and Veterinary Sciences,Southwest Minzu University,Chengdu 610041)

机构地区：[1]西南民族大学畜牧兽医学院,成都610041

出　　处：《中国农业科学》2021年第23期5144-5162,共19页Scientia Agricultura Sinica

基　　金：国家现代农业产业技术体系四川创新团队项目(CARS-SVDIP);四川省科技厅项目(2016KZ0007);西南民族大学中央高校基本科研业务费专项基金项目(2020NYB27)。

摘　　要：【目的】对329株采自川西北高原牦牛和藏猪源大肠杆菌进行生物被膜形成能力、抗生素耐药基因、整合酶基因、毒力基因和遗传谱系分型分析,以期了解其耐药现状、毒力特性和优势遗传谱系分布等生物学特征。【方法】利用麦康凯培养基和15e肠杆菌科细菌生化编码鉴定管对牦牛和藏猪粪便和胃肠道内容物样本进行大肠杆菌分离和鉴定;采用改良结晶紫半定量染色法和微量肉汤稀释法分别对分离菌株进行生物被膜形成能力鉴定及其对24种抗菌药物的敏感性试验;采用普通PCR或多重PCR方法对28个耐药基因、2个整合酶基因、15个毒力基因进行检测和遗传谱系分型分析。【结果】(1)从471份牦牛、藏猪粪便和胃肠道内容物样本分离鉴定329株大肠杆菌,分离率为78.9%。(2)329株大肠杆菌大多表现出弱或无生物被膜形成能力,仅2株为强成膜能力表型(其中牦牛和藏猪源各1株)。(3)329株大肠杆菌对24种抗菌药物多表现出一定的耐药性并呈现多重耐药现象,其中对磺胺甲唑、磺胺二甲嘧啶、链霉素、氯霉素、氨苄西林、利福平和土霉素耐药率较高,对氨基糖苷类(卡那霉素、阿米卡星和壮观霉素)、β-内酰胺类(头孢噻呋、头孢曲松、头孢唑啉)、喹诺酮类(萘啶酸、沙拉沙星、恩诺沙星、环丙沙星、达氟沙星、左氧氟沙星)和多黏菌素B等敏感。(4)除cat1、cat2、bla_(CMY-2)、bla_(SHV)、tetC、tetG、tetX等7个耐药基因外,其他21个耐药基因检测结果均为阳性,其中以aac(6')-Ib最为流行,其次是sul1和floR,检出率均在30%以上。藏猪源大肠杆菌对喹诺酮类抗生素耐药与qnrA相关,牦牛源大肠杆菌对β-内酰胺类耐药性与bla_(TEM)和bla_(DHA)相关。(5)整合酶基因intⅠ1和intⅠ2的检出率分别为30.09%(99/329)和4.56%(15/329),其中10株大肠杆菌(牦牛源2株,藏猪源8株)同时检测到intⅠ1和intⅠ2。(6)毒力基因agn43、sitA、om【Objective】To improve the understanding of biological properties of drug resistance status,virulence characteristics and the predominant phylogroups of Escherichia coli(E.coli)isolated from yaks and Tibetan pigs in northwest Sichuan Plateau,biofilm-forming ability,antibacterial resistance genes,virulence genes,integrase genes and phylogenetic analyses were carried out in current study.【Method】Fecal samples and gastrointestinal contents from yaks and Tibetan pigs were collected for E.coli isolation and identification using MacConkey agar and 15e enterobacteriaceae bacterial biochemical coding identification tube.Modified semi-quantitative crystal violet staining method and microdilution broth method were used for biofilm-forming ability and antibacterial sensitivity testing to 24 antibacterial agents,respectively.Meanwhile,the detection of 28 antibacterial resistance genes,2 integrase genes and 15 virulence genes and phylogenetic analyses were performed by conventional PCR or multiple PCR method.【Result】The results showed that:(1)329 strains of E.coli were isolated and identified from 471 feces and gastrointestinal samples collected from yaks and Tibetan pigs,and the isolation rate was 78.9%.(2)Most of the 329 strains of E.coli performed weak or absent biofilm-forming ability,and only 2 strains showed strong biofilm formation phenotype(one isolated from yak and the other isolated from Tibetan pig).(3)Most of the 329 strains of E.coli revealed drug resistance to 24 antibacterial agents and were multi-drug resistant,among which,the drug resistance rates to Sulfamethoxazole,Sulfadimidine,Streptomycin,Chloramphenicol,Ampicillin,Rifampicin,and Oxytetracycline were relatively high,and were sensitive to Aminoglycosides(Kanamycin,Amikacin,Spectinomycin),β-lactams(Ceftiofur,Ceftriaxone,Cefazolin),Quinolones(Nalidixic acid,Sarafloxacin,Enrofloxacin,Ciprofloxacin,Danofloxacin,Levofloxacin)and Colistin B.(4)Twenty-one antibacterial resistance genes(ARGs)were detected positive and the other seven ARGs(cat1,cat2,bl

关 键 词：大肠杆菌 生物被膜 抗生素耐药基因(ARGs) 整合酶基因 毒力基因 遗传背景 

分 类 号：S852.61[农业科学—基础兽医学]