新疆克拉玛依地区奶牛源致病性大肠杆菌的血清型、系统进化群及MLST分析

doi:10.16431/j.cnki.1671-7236.2021.04.029

摘要/Abstract

摘要： 试验对新疆克拉玛地区106头健康成年荷斯坦奶牛的新鲜粪便进行了致病性大肠杆菌的分离鉴定、分型及遗传进化分析，初步掌握了调查地区奶牛致病性大肠杆菌的流行情况。参考USDA检测法，对106份粪便样品选择性增菌，分别利用麦康凯和伊红美蓝琼脂培养基对大肠杆菌进行分离纯化鉴别，通过PCR方法鉴定，获得55株大肠杆菌分离株，对这些分离株进行致病性大肠杆菌血清型鉴定、系统进化群及多位点序列分型（multilocus sequence typing，MLST）分析，利用goeBURST和Mega 7.0生物学软件进行聚类和进化研究，并进行流行病学分析。结果显示，106头健康荷斯坦奶牛的新鲜粪便中大肠杆菌的检出率为51.89%，55株大肠杆菌分离株共鉴定出9种与致病性相关的血清型，检出率为54.55%，其中肠致病性大肠杆菌（EPEC）、肠侵袭性大肠杆菌（EIEC）和肠产毒性大肠杆菌（ETEC）血清型检出率分别为21.82%、18.81%和14.55%，优势血清型为O142∶K86（B）和O124∶K72（均为10.91%），前者属于肠致病性大肠杆菌，后者属于肠侵袭性大肠杆菌，而肠出血性大肠杆菌（EHEC）血清型检出率为0；系统发育群结果表明，55株大肠杆菌中B1群所占比例最高，为70.91%，其次为A群，占21.82%，D群所占比例较少，占7.27%，B2群未检测出；MLST分析可知，分离株中存在24个ST型，其中优势ST型为ST154（18.18%）；进化树分析显示有7个进化分支。结果表明，新疆克拉玛依地区健康成年奶牛粪便大肠杆菌多样性丰富，不仅存在多种致病性大肠杆菌，而且大肠杆菌菌株之间还存在一定的进化关系。因此，健康成年奶牛粪便也具有潜在的安全风险，应加强对奶牛粪便致病性大肠杆菌的检测和监控。

关键词: 奶牛源大肠杆菌; 血清型; 系统发育群; 多位点序列分型(MLST)

Abstract: The pathogenic Escherichia coli (E.coli) was isolated,identified,typed and analyzed for genetic evolution of 106 healthy adult Holstein cows in Karama,Xinjiang,and the epidemic situation of pathogenic Escherichia coli in cows in the investigation area was preliminarily known.With reference to the USDA detection method,106 stool samples were selectively enriched,and the E.coli was separated,purified and identified using MacConkey and Eosin Methylene Blue agar medium.The PCR method was used to identify 55 strains of E.coli.The isolates were subjected to pathogenic E.coli serotype identification,phylogenetic group and MLST type analysis,clustering and evolution studies were performed using goeBURST and Mega 7.0 biological software,and epidemiological analysis was performed.The results showed that the detection rate of Escherichia coli in fresh feces of 106 healthy Holstein cows was 51.89%.Nine serotypes related to pathogenicity were identified in 55 E.coli isolates,the detection rate was 54.55%.The detection rates of EPEC,EIEC and ETEC were 21.82%,18.81% and 14.55%,respectively.The dominant serotypes were O142∶K86 (B) and O124∶K72 (both 10.91%).The former belonged to EPEC,the latter belonged to EIEC,and the detection rate of EHEC serotype was 0.The phylogenetic group results show that the B1 group of 55 strains of E.coli accounted for the highest proportion,which was 70.91%,followed by group A,accounting for 21.82%,group D,which proportion was small,accounting for 7.27%,and the B2 group was not detected.MLST analysis showed that there were 24 ST types in the isolates,among which the dominant ST type was ST154 (18.18%).The evolutionary tree analysis revealed seven branches of evolution.In Conclusion,the feces of healthy dairy cows in Karamay,Xinjiang had a rich diversity of E.coli.There were not only pathogenic E.coli but also a certain evolutionary relationship between E.coli strains.Therefore,the excrement of healthy adult cows also had potential safety risk,so the detection and monitoring of pathogenic E.coli in cows excrement should be strengthened.

Key words: Escherichia coli from cows; serotype; phylogenetic group; multilocus sequence typing (MLST)

中图分类号:

S885.1

江婉琳, 马勋, 王静, 平丽莹, 蔡思奇. 新疆克拉玛依地区奶牛源致病性大肠杆菌的血清型、系统进化群及MLST分析[J]. 中国畜牧兽医, 2021, 48(4): 1414-1422.

JIANG Wanlin, MA Xun, WANG Jing, PING Liying, CAI Siqi. Serotype,Phylogenetic Group and MLST Analysis of Pathogenic Escherichia coli Isolated from Dairy Cattle in Karamay,Xinjiang[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(4): 1414-1422.

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