| Object: In this study,E.coli was isolated and identified from anal swabs of dairy cows,and its serotype,phylogenetic group,MLST,distribution of virulence genes and drug resistance were studied,and it is a potential pathogenicity of E.coli derived from cow feces.Provide a basis for the rational use of antibiotics in the breeding farms.Methods:(1)Anal swabs of cows in Karamay and Aksu of Xinjiang were collected for isolation and culture of Escherichia coli,and biochemical identification and 16 S r RNA PCR identification of isolates were performed.(2)Serotype identification by glass agglutination test;The chuA,yjaA and Tsp E4.C2 genes were detected by triple PCR method,and the phylogeny of E.coli was determined according to the electrophoresis pattern.7 housekeeping genes(adk,fumC,gyrB,icdA,mdh,purA,and recA)were amplified and sequenced by PCR,and MLST typing was completed by online submission of sequences.According to the sequence number of five or more housekeeping genes with the same sequence number among the ST types,they were grouped into a group.ST type cluster analysis of isolates was performed with Goe Burst software.(3)PCR was used to detect 17 E.coli virulence genes,including sfaS,crl and fliC.(4)The drug resistance of E.coli isolates was determined by Kirby-Bauer method.And bla TEM,bla SHV,sul 1,sul 2,aac(3)-?,cml A,tet A,tet B,qnr B and other 24 drug resistance genes targeting ?-lactams,sulfonamides,aminoglycosides,chloramphenicol,tetracycline and quinolone antibiotics were detected by PCR.Results:(1)101 and 106 anal swabs were collected from Aksu and Karamay cattle farms,respectively.After biochemical identification and 16 S r RNA PCR detection,55 isolates of E.coli in Karamay were obtained,and the isolation rate was 51.89%.There were 51 isolates of E.coli in Aksu,and the isolation rate was 50.50%.(2)Serotype results showed that 31 strains of the 55 E.coli isolates from Karamay were detected as pathogenic E.coli serotype,and 24 strains were not detected as serotype.32 strains of 51 E.coli isolates were detected as pathogenic E.coli serotype,and 19 strains were not detected as serotype.The highest detection rate was EPEC serotype,accounting for 23.64%(13/55)and 33.33%(17/51),respectively.EHEC was not detected in all cases.The dominant serotypes of E.coli isolates from Karamay and Aksu were O142: K86(B)and O15: K? And O125:K70(B15),O86:K61(B7),O164:K?.The phylogenetic group results showed that the 106 strains of E.coli belonged to 4groups: 69.81%(74/106)belonged to group B1,17.92%(19/106)belonged to group A,10.38%(11/106)belonged to group D and 1.89%(2/106)belonged to group B2.The majority of E.coli isolates from Karamay and Aksu were B1 group,accounting for 68.63%(35/51)and70.91%(39/55),respectively.The isolates of group B2 only came from cattle farms in Aksu area.MLST typing showed that the 106 E.coli isolates belonged to 62 different ST types,of which 30 ST types existed in the existing database,accounting for 48.39%.32 new ST types(STn)were found,numbered STn-1 ? 32,accounting for 51.61%.ST-154 was the dominant ST type,accounting for 12.26%(13/106).The ST types of E.coli isolates from Karamay were relatively concentrated,while the ST types of E.coli isolates from Aksu were scattered in each type.Goe Burst analysis showed that the 106 E.coli isolates were divided into 8 clone groups and 20 individual types.(3)PCR detection of 17 virulence genes,such as hly A,afa,eae A and ipa H,showed that the detection rates of fli C,omp A and crl were 100%,99.06% and 91.51%,respectively,which were much higher than other virulence genes.Five virulence genes stx-1,iha,sfa,iuc D and afa were not detected in this experiment.According to the distribution of specific pathogenetic genes,there were four pathogenetic hybridization patterns: EAEC/UPEC,EPEC/ETEC,EIEC/UPEC and UPEC/NMEC.(4)The drug resistance rate of 106 E.coli isolates to penicillin is above 97%;The drug resistance rate of ceftithiophene was more than 60%.The multiple drug resistance rate was37.74%.The results showed that 11 genes including TEM,bla OXA,tet A,aac(3)-II,and sul 1were detected.13 genes such as bla CTX-M,qnr A,oqx A,and aac(3)-I were not detected.There is some consistency between phenotype and drug resistance gene.Conclusion:(1)In this study,106 strains of Escherichia coli were isolated from anal swabs derived from cow feces,with an isolation rate of 51.21%.The serotype,phylogenetic group and MLST typing showed significant diversity of isolates,with O86: K61(B7),B1 and ST154 being the most prevalent isolates.(2)The results of serotype and virulence gene indicated that there was pathogenic risk of E.coli from cow feces.(3)E.coli isolates showed certain drug resistance to penicillin and cefothiophene,and the proportion of multiple drug resistance was 37.74%.There were drug-resistant genes TEM,OXA,qnrS,aac(6')-Ib-cr,tetA,tetB,sul 1,sul 2,sul 3,cmlA.(4)The degree of variation of E.coli isolates from the two regions were different.The ST type,virulence gene and drug resistance gene analysis indicated that the evolution of Karamay isolates were relatively concentrated,while the evolution of Aksu isolates were relatively dispersed. |
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