Preliminary Study On Pollution Testing And Pathogenicity Of Non-O157 Shiga Toxin-producing Escherichia Coli Isolated From Xinjiang Cattle And Sheep

Shiga toxin-producing Escherichia coli(STEC)is a class of food-borne pathogens that carry one or two Shiga toxin genes encoded by pre-phage.It has high pathogenicity,mainly manifested as Hemolytic uremic syndrome(HUS).The cattle and sheep breeding industry is the leading industry in the development of modern animal husbandry in Xinjiang.Because STEC strain exists in the storage and host cattle and sheep,it potentially affects the quality and safety of animal food and environmental health.More and more diseases caused by human infection have attracted worldwide attention.In this study,PCR method was used to detect the contamination and genotyping of non-O157 STEC from cattle and sheep in some areas of xinjiang.Further cloning and bioinformatics analysis of stx1 and stx2 were carried out.The difference of pathogenicity of non-O157 STEC in vivo and in vitro was studied by mouse challenge test and human intestinal epithelial cells(HIEC)infection,which laid a foundation for further study on the pathogenesis of STEC.The main research results are as follows:1.431 samples were collected from farms,slaughterhouses and markets of cattle and sheep,and 392 samples were identified(the separation rate was 91.0%).Non-O157 STEC 64 strains(the isolation rate was 16.3%).Non-O157 STEC strain 32 encoding stx1+stx2(isolation rate 49.2%),non-O157 STEC strain 29 encoding stx1(separation rate 44.6%),non-O157 STEC strain 4 encoding stx2(separation rate 6.2%).The detection rates of virulence genes eae,HlyA,subA,toxB,and iha were 12.5%,79.7%,14.1%,4.7%,and 51.6%,respectively.The drug resistance rates of non-O157 STEC to madecamycin,ampicillin,piperacillin,tobramycin,cefalothiophene,cefazolin,cefazolin and cefoxitin were 61%,3.1%,1.6%,4.7%,1.6%,4.7%respectively,and there were 8 strains with multiple drug resistance(14%).The results of eric-pcr showed polymorphism distribution,which was divided into two clusters:A(36 strains)and B(28 strains),and the same eric-pcr fingerprint strains existed in each cluster.2.6 strains of representative strains(H34,M22-5,M15-1,213,B21,B34)over stx1 sequence with the German outbreak strain O157:H7 type stxlc sequence homology of 100%,5 strains of representative strains(H34,M15-1,213,B21,B34)stx2 total length of the sequence and the outbreak strain O91:H21 and O91:H8,Australia,the outbreak strain O8:H19,the outbreak strain O78:H8 type stx2d sequence homology of 100%.3.The mortality rate of mice in H34,M22-5,M15-1 and 213 groups was 60%,60%,20%and 30%respectively,while no death occurred in mice in B21 and B34 groups.The survival rate of H34,M15-1,B34,M22-5 and B21 was 39.3%,40.2%and 40.8%,respectively.Conclusion:Non-O157 STEC is polluted in farms,slaughterhouses and markets.The pollution of farms is the most serious,and the source of cattle is more polluted than sheep.Cattle and sheep are closely related to each other.The stx1 of the six representative strains tested were all stx1c type.The stx2 of the five representative strains were all stx2d type.Six non-O157 STEC strains have different pathogenicity in mice,and four strains have different degrees of lethality in mice.Six non-O157 STECs have different pathogenicity to HIEC.H34 is the most pathogenic to HIEC.The optimal concentration of HIEC for HIEC is 106 CFU/mL,and the optimal time is 1 h.