Isolation,Identification,Molecular Typing And Drug Resistance Of Extraintestinal Pathogenic Escherichia Coli From Cattle And Sheep In Part Of Xinjiang

Objective:Escherichia coli?E.coli?is one of the three major bacterial diseases that endanger the world's aquaculture industry and is an important zoonotic pathogen.The infection and epidemic caused by E.coli is a worldwide problem.Xinjiang is a large province for cattle and sheep breeding.E.coli disease often occurs in cattle and sheep,which not only causes significant economic losses to livestock production,but also poses a potential threat to public health.In order to understand the prevalence and biological characteristics of Extraintestinal Pathogenic Escherichia coli?ExPEC?of cattle and sheep in recent years in Xinjiang,pathogens,phylogenetic clustering,serotype and multilocus sequence typing?MLST?,virulence factors and biofilm?BF?formation ability analysis,the drug resistance phenotype and resistance gene carrying status and drug resistance plasmid transmission were studied.Methods:?1?Sterile collection of liver,spleen,lung,heart,brain,lymph and other tissues of the suspected cattle and sheep infected by E.coli in parts of Xinjiang from 2015 to 2019,the conventional bacterial isolation and identification technology combined with bacterial biochemical identification system and 16S rRNA amplification were used to identify pathogens,which pathogenicity was determined by mouse test,detection of ExPEC special virulence factors by PCR,the phylogenetic group of isolates was determined by multiplex PCR method,and the serotype was determined by slide agglutination,and specific gene sequencing method was used to identify MLST.?2?PCR gene amplification method was used to analyze the situation of carrying virulence factor genes of the isolates,and 96-well microplate method was used to analyze their ability to produce BF and the optimal time for formation.?3?The KB was used to detect the resistance phenotypes of 26 antibiotics,and the common drug resistance gene was analyzed with PCR.The drug-resistant plasmids carrying?-lactam genes were extracted from the isolates and transferred into blank E.coli DH5?for the detection and analysis of plasmid drug-resistant transmission characteristics.Results:?1?132 strains of E.coli were isolated and identified from liver,spleen,lung,heart,brain and lymph nodes of the cattle and sheep which were infected by E.coli in parts of Xinjiang from 2015 to 2019.According to the characteristics of carrying virulence factors,these isolates were determined as ExPEC and belonged to 7 phylogenetic groups,of which were part of the dominant groups A and B1.A total of 22different serogroups were identified,mainly O101?26 strains?,O154?14 strains?and O65?8 strains?.132strains of ExPEC were divided into 15 different ST types,ST10 was the most common sequence type,ST23 accounted for 13.2%?5/38?and ST457 for 10.5%?4/38?.?2?Among the isolates,the detection rate of the curli pili crl gene was the highest,reaching 97.4%,followed by the hemolysin hlyE with a detection rate of 94.69%;group B2 has the most virulence factor genes;105?79.5%?strains had the ability to produce BF,of which 15?11.4%?were strong film-forming ability,28?21.2%?showed intermediate film-forming ability and 62?47%?were weak film-forming ability.BF began to form at 6 h,the shape of BF tended to be complete and reached its peak at about 36 h,after that,the network structure of BF gradually dissociatedand disappeared at about 72 h.?3?The results of drug sensitivity tests showed that the resistance rates to?-lactams,aminoglycosides,tetracyclines and quinolones were generally high,including cefraditin?81.2%?,streptomycin?76.9%?,gentamicin?71.6%?and enrofloxacin?64.6%?.The genes with the high detection rate were gyrA from aminoglycoside?98.28%?,tetA rom tetracyclines?75%?,sul2 from sulfonamide?75%?,bla_TEMEM from?-lactam?72.73%?,aph?3'?-Ia from Aminoglycoside?62.12%?respectively.The isolates carried a maximum of 15 and a minimum of 2 drug-resistant genes.Conclusion:These ExPEC derived from the infeted cattles and sheeps in some areas of Xinjiang belonged to groups A and B1,while they have abundant serotypes and multisite sequence typing and carried abundant virulence factor genes and drug resistance genes.The isolates showed a high drug resistance rate and a wide range of multiple drug resistance rates,most of which could produce significant BF.