Epidemiological Characteristics And Drug Resistance Characteristics Of Escherichia Coli In Some Large-Scale Dairy Farms In Tai'an Area

Escherichia coli(E.coli)is the most important species of Escherichia,which is a component of normal flora in warm-blooded animals and human intestinal tract.Escherichia coli is abundant in animal feces and is often transmitted through direct or indirect contact and feces.In production,the harm caused by colibacillosis is very serious,its clinical symptoms are complex and the case characteristics are diverse.In recent years,with the general control of major diseases in intensive dairy farms,E.coli disease has shown an upward trend.Dairy products and meat products are necessties of our daily life.E.coli infections can affect people and further threaten public health through the food chain.Antibiotics are still effective in the treatment of E.coli disease,but the unreasonable use of drugs will not only lead to antibiotic residues in dairy products,but also cause high resistance rate of E.coli from dairy cows,broad spectrum of drug resistance and high proportion of muLti-drug resistant strains.It poses a great threat to aquacuLture and human health.In order to investigate the epidemiology of Colibacillosis in several large-scale dairy farms in Tai'an area,194 samples of fresh feces were collected from four large-scale dairy farms in Tai'an area.E.coli in feces were isolated and identified by enriching bacteria,selective cuLture of eosin methylene blue medium and identification of 16 SrRNA.Plate agglutination test was performed with serum diagnostic kit to detect EIEC,EPEC,ETEC and EHEC in the isolated strains.K-B method was used to detect the resistance of strains isolated to different antibiotics by drug susceptibility test of 9 classes and 14 kinds of drugs.Genomic DNA was extracted from the strains and 24 drug-resistant genes and class I integrons were detected to understand the carrying status of different drug-resistant genes and drug-resistant gene cassettes in the strains.To clarify the epidemic situation of E.coli in this region,muLti-locus sequencing technique was used to trace the origin of E.coli isolates.Isolated strains were cuLtured in sheep blood plate to detect the proportion of strains carryingIV hemolytic rings.The diameter of hemolytic rings in strains was measured.Three strains with the largest diameter of hemolytic rings were selected for pre-experiment to determine the safe dosage for 8-day-old SPF chickens and 30-day-weaned rabbits.The pathogenicity of the strains was verified by design experiments.The results of isolation and identification showed that 171 strains of Escherichia coli were detected from 194 feces collected from four large-scale healthy dairy farms in Tai'an,the isolation rate was 88.14%.Agglutination test showed that 79 strains of Escherichia coli were pathogenic serotypes,including 13 different serotypes,of which O128 and O143 had the highest detection rate,accounting for 7.0%(12/171),and EPEC(19.9%)was the most common serotype.The drug sensitivity test showed that the highest rate of ampicillin resistance was 33.9%,followed by tetracycline,the drug resistance rate was 24.0%.The isolates were more sensitive to cephalosporins,quinolones and chloramphenicol antibiotics,and the sensitivity rate was more than 90%.In addition,the proportion of muLti-drug resistant strains was 22.2%.The detection rates of beta-lactam resistance genes blaTEM,blaCTX-M-15,blaOXA-1 and blaCMY-2 were 59.1%,4.7%,1.8% and 0.6%,respectively.The detection rates of AAC(6')-Ib-cr,qnrS,oqxa and qnrD were 4.1%,1.8%,1.2% and 0.6% respectively.The detection rates of aminoglycoside resistance genes ant(2')and AAC(3)-II were 40.9% and 2.3%respectively.The detection rates of sul2,sul 3 and sul1 were 16.4%,10.5% and 3.5%respectively.The detection rate of chloramphenicol resistance gene cmlA was 0.6%.No tetracycline resistance gene was detected.No blaPSE,blaSHV,qnrA,qnrB,qnrC,AAC(3)-I,AAC(3)-III,AAC(3)-IV,tetA and tetB resistance genes were detected.The results of integron detection showed that 7 strains carried four drug-resistant gene cassettes,the detection rate was 4.1%(7/171).The most common drug-resistant gene cassette was dfrA12-aadA2-sul1(4/7).The molecuLar characteristics of integron-gene cassette were mainly dfrA and aadA gene types mediating sulfonamide-trimethoprim and aminoglycoside resistance.Only a few drug-resistant strains carried type I integron.Through MLST analysis,eight ST types were detected as ST58,ST5,ST23,ST1146,ST172,ST155,ST661 and ST3096,of which ST58 accounted for the highest proportion of26.3%,and different strains of dairy sites had the same ST type.Among them,ST8 and ST155 are a clonal complex,indicating that the strains of different dairy sites are closely related,and E.coli crosses in different dairy farms.Animal experiments showed that in 8-day-old SPF chickens,one died within 12 hours in group A1,the mortality rate was 16.67%,and in 30-day-old rabbits,five died within 12 hours in group A2,the mortality rate was 100%.The results showed that the pathogenicity of selected E.coli strains from cattle to rabbits was higher than that of SPF chickens,indicating that the strains producing hemolytic rings were pathogenic and had different pathogenicity to different species.The results of animal experiment showed that the death rate caused by selected E.coli strain was higher among rabbits than that among SPF chickens.Which indicated that the Screened strains of E.coli can infect rabbits and SPF chickens with different pathogenic abilities.This study analyzed the epidemic situation of E.coli from dairy cows in Tai'an area in2017 from various aspects,and got a preliminary understanding of the epidemic situation.This research has a reference significance for the prevention and control of E.coli from dairy farms,and revealed the potential hazards to public health and safety.