腹泻仔猪肠道大肠杆菌分离鉴定与毒力基因及耐药性分析

Identification,virulence gene and antibiotic resistance analysis of Escheri chia coli isolated from diarrheic piglets

为了解粤西部分地区腹泻仔猪中大肠杆菌流行情况及耐药谱,采用鉴别培养基、革兰氏染色镜检、快速生化特性鉴定、PCR鉴定以及药敏纸片琼脂扩散法等方法,对粤西地区不同养殖场中114份腹泻仔猪直肠拭子样本进行大肠杆菌分离鉴定、毒力基因以及耐药性分析。结果表明,所有样本均分离得到优势大肠杆菌,其生化特性与大肠杆菌标准质控菌株ATCC 25922一致。在毒力基因检测试验中,对所有大肠杆菌分离菌株(共114株)进行9种毒力基因检测,结果显示,毒力基因总检出率为90.35%(103/114)。根据毒力基因PCR鉴定结果确定各毒力基因检出率:STa(71.93%,82/114)、STb(61.40%,70/114)、LT(11.40%,13/114)、K99(7.02%,8/114)、F41(6.14%,7/114)、F18ac(5.26%,6/114)、K88(0.88%,1/114)、Stx2e(3.51%,4/114),而黏附素987P未在分离菌株中检出。药敏试验结果显示,大肠杆菌对磺胺甲基异恶唑、氨苄西林、四环素均高度耐药,耐药率为100%(114/114)、97.37%(111/114)、96.49%(110/114),对其余抗生素耐药率为新霉素48.25%(55/114)、左氧氟沙星43.86%(50/114)、多黏菌素B 42.98%(49/114)、庆大霉素42.11%(48/114)、环丙沙星39.47%(45/114)、头孢噻呋24.56%(28/114)。研究初步获得广东省西部部分地区腹泻仔猪肠道中大肠杆菌毒力基因分布及耐药谱数据。

To gain insight into the prevalence of infection and drug resistance profiles of Escherichia coli isolated from diarrheic piglets in parts of western region of Guangdong Province,the present study was aimed at isolation and identification of E.coli from 114 rectal swabs in diarrheic piglets through differential media,Gram staining and rapid biochemical characterization,and then to identify virulence genes and antibiotic resistance of pathogenic E.coli through PCR and agar disk diffusion methods.The identification results showed that the predominant E.coli could be isolated from each sample,and the biochemical characteristics of isolated E.coli were consistent with that of E.coli ATCC 25922.A total of nive virulence genes were detected in all the E.coli isolates(114 strains).The results showed that the overall detection rates of virulence genes were 90.35%(103/114),among which the detection rates were 71.93%(82/114)for STa,61.40%(70/114)for STb,11.40%(13/114)for LT,7.02%(8/114)for K99,6.14%(7/114)for F41,5.26%(6/114)for F18ac,0.88%for K88(1/114)and 0%(0/114)for 987P and 3.51%(4/114)for Stx2e.The results of drug sensitivity testing revealed that the E.coli isolates displayed high rates of resistance to sulfamethoxazole(100%,114/114),ampicillin(97.37%,111/114)and tetracycline(96.49%,110/114).The rates of resistance to other antibiotics were observed for:neomycin(48.25%,55/114),levofloxacin(43.86%,50/114),polymyxin B(42.98%,49/114),gentamicin(42.11%,48/114),cipro-floxacin(39.47%,45/114),and ceftiofur(24.56%,28/114).The present research obtained basic data of virulence gene distribution and drug resistance profiles of E.coli isolated from diarrheic piglets in parts of western region of Guangdong Province.