摘要
为了解河北地区犊牛源致病性大肠杆菌的优势血清型、ST型、毒力基因及耐药特性,本研究对河北4个地区(保定、衡水、承德和秦皇岛)采集的128份腹泻犊牛粪便样品进行细菌分离,并采用特异性引物经PCR鉴定,结果显示,在各培养基均生长出具有大肠杆菌特征的菌落,进一步经16S rRNA基因的PCR鉴定后确定分离到76株致病性大肠杆菌。采用PCR检测分离菌的毒力基因,采用玻片凝集法鉴定致病性大肠杆菌血清型,通过K-B纸片扩散法检测分离菌的耐药性,采用PCR检测分离菌共8类41种耐药基因。根据药敏试验与耐药基因检测结果分析二者之间的相关性,采用多位点序列分型(MLST)技术对分离菌分型。结果显示,76株致病性大肠杆菌分离株优势血清型为O29;共检测出15种毒力基因,其中irp2、hlyF、iucD基因的检出率相对较高,分别为36.84%(28/76)、21.05%(16/76)、21.05%(16/76);大部分(80.26%,61/76)大肠杆菌携带1种~2种毒力基因;药敏试验结果显示76株致病性大肠杆菌均至少对4种药物耐药,对链霉素、大观霉素、青霉素等11种药物耐药率均在50%以上,对丁胺卡那、庆大霉素、氟苯尼考等12种药物的敏感率在50%以上;共检出31种耐药基因,其中喹诺酮类耐药基因总检出率最高(gyrA:97.37%,74/76),其次为磺胺类耐药基因(dhpS:96.05%,73/76)和氨基糖苷类耐药基因(aac2:84.21%,64/76),76株致病性大肠杆菌均至少携带5种耐药基因;且耐药表型与耐药基因间无明显相关性;MLST分型结果显示河北保定等地区致病性大肠杆菌优势ST型为ST101。本研究结果表明河北保定、衡水、承德和秦皇岛4个地区致病性大肠杆菌的毒力基因、血清型和ST型分布均表现出较高的多样性,尽管携带毒力基因数量较少,但耐药性严重。本研究为河北地区养牛业合理使用抗生素防控致病性大肠杆菌病提供了参考依据。
In order to understand the dominant serotypes,ST types,virulence genes and drug resistance characteristics of calfderived pathogenic Escherichia coli in Hebei,128 fecal samples of calf diarrhea collected from four regions of Hebei(Baoding,Hengshui,Chengde and Qinhuangdao)were subjected to bacterial isolation in this study,and were further identified through isolation and culture by PCR with specific primers,which showed that colonies with the characteristics of Escherichia coli were grown in each culture medium.Further PCR identification confirmed the isolation of 76 pathogenic Escherichia coli strains.The virulence genes of the isolates were detected by PCR,the serotypes of pathogenic Escherichia coli were identified by slide agglutination,and the drug sensitivity were detected by K-B paper diffusion method and typed by multilocus sequence typing(MLST)technique.The results showed that the dominant serotype of 76 pathogenic Escherichia coli isolates was O29;a total of 15 virulence genes were detected,among which the detection rates of irp2,hlyF,and iucD genes were relatively high,36.84%(28/76),21.05%(16/76),and 21.05%(16/76),respectively;the majority of(80.26%,61/76)Escherichia coli carried the number of virulence genes of 1-2;the results of drug sensitivity test showed that all 76 strains of pathogenic Escherichia coli were resistant to at least 4 drugs,the resistance rate of 11 drugs such as streptomycin,macrolide and penicillin was above 50%,and the sensitivity rate of 12 drugs such as butylmethamphetamine,gentamicin,and fosfenacol was above 50%;a total of 31 drug-resistant genes were detected,of which the total rate of quinolone-resistant genes detection was(gyrA:97.37%,74/76),followed by sulfonamide resistance genes(dhpS:96.05%,73/76)and aminoglycoside resistance genes(aac2:84.21%,64/76),and all 76 strains of pathogenic Escherichia coli carried at least five resistance genes;the MLST typing results showed that the predominance of pathogenic Escherichia coli in Baoding,Hebei and other regions was ST101.The results of this study showed that the distribution of virulence genes,serotypes and ST-types of pathogenic Escherichia coli in the four regions of Hebei,Baoding,Hengshui,Chengde and Qinhuangdao,showed high diversity,carrying a small number of virulence genes but severe drug resistance.This study provides a reference basis for the rational use of antibiotics for the prevention and control of pathogenic Escherichia coli disease in cattle breeding in Hebei.
作者
韩平
王小立
赵允清
苏蒙蒙
李涵
候蔷
李彬
郑百芹
张洪军
吴同垒
张香斋
张艳英
HAN Ping;WANG Xiao-li;ZHAO Yun-qing;SU Meng-meng;LI Han;HOU Qiang;LI Bin;ZHENG Bai-qin;ZHANG Hong-jun;WU Tong-lei;ZHANG Xiang-zhai;ZHANG Yan-ying(Key Laboratory of Preventive Veterinary Medicine of Hebei Province,Hebei Normal University of Science and Technology,Qinhuangdao 066604,China;Tangshan Food and Drug Comprehensive Inspection and Testing Center,Tangshan 063000,China;Animal Epidemic Prevention Supervision station of left behind camp in Funing District,Qinhuangdao City,Qinhuangdao 066600,China;Animal Disease Prevention and Control Center ofWeichang Manchu-Mongolian Autonomous County,Chengde 068450,China)
出处
《中国预防兽医学报》
北大核心
2025年第6期568-576,共9页
Chinese Journal of Preventive Veterinary Medicine
基金
河北省科技厅重点研发计划项目(19226628D)
河北省科技厅重点研发乡村振兴技术创新专项(22326612D)
河北省现代农业产业技术体系建设专项资金(HBCT2024280205)
围场满族蒙古族自治县科技计划项目(致犊牛腹泻病原诊断技术研究与益生菌的筛选和应用,2024)。
关键词
大肠杆菌
毒力基因
耐药基因
耐药性
优势血清型
Escherichia coli
virulence gene
drug resistance gene
drug resistance
dominant serotype
