Clinical Detection Of Waterfowl Pathogenic Bacteria In Jiangsu And Molecular Epidemiology Of Escherichia Coli And Its Characteristics

In recent years,although the scale of waterfowl farming has increased,the diseases of waterfowl have not been improved,even bacterial diseases have become more complicated because of multiple pathogens secondary or concurrent.In addition,due to the data of drug sensitivity monitoring,comprehensive prevention measures and epidemiological investigation is not perfect yet,the abuse of antibiotic drugs had caused serious challenges to the prevention and control of waterborne bacterial diseases.In order to quickly detect four important pathogens of waterfowl in Jiangsu province,we collected a total of 768 samples from 64 farms in all waterfowl scale farming areas in Jiangsu from 2017 to 2019,including 510 samples of clinical healthy waterfowl source anal swabs,258 tissue samples.Synthesis of primers based on Escherichia coli(E.coli)F1 fimbriae subunit A(fimA)and P fimbriae subunit C(papC),the gene of outer membrane protein(ompA)in Riemerella anatipestifer,the HlyA gene of Listeria monocytogene(hlyA),and the enterotoxin gene of Salmonella(stn).Four important pathogens of waterfowl including pathogenic E.coli,Riemerella anatipestifer(RA),Listeria monocytogene(LM)and Salmonella were identified by PCR.The results showed that the average detection rate of F1,P,RA,LM and Salmonella in waterfowl samples was 74.21%(570/768),0.78%(6/768),2.73%(21/768),7.42%(57/768)and 5.60%(43/768),respectively.From the results we could see that the incidence of E.coli in watery and poultry bacterial diseases in Jiangsu is the highest,and the infection of LM shouldn't be ignored.All these data is expected to provide some reference for prevention and control of waterborne bacterial diseases.E.coliis very important in waterfowl bacterial diseases.In order to reveal the distribution of virulence genes in Escherichia coli,the main virulence genes(tsh,hlyE,ECs3703,ECs3737,irp2,and eae)of waterfowl Escherichia coli were detected by PCR in 355 positive Escherichia coli anal swabs samples and 213 positive tissue samples.Then the positive samples were isolated,purified and identified.A total of 342 E.colist rains were isolated from anal swab samples,the Isolateswere divided into 12 different v irulence factors,followed by F1+(217 strains),F1+ETT2+(56 strains),F1+HlyE+(3 strains),ETT2+HPI+(15 strains),F1+ETT2+HPI+(13 strains),F1+HPI+Tsh+(1 strains),F1+ETT2+HPI+Tsh+(7 strains),F1+ETT2+LEE+Tsh+(4 strains),F1+P+ETT2+Tsh+(2 strains),F1+ETT2+LEE+Tsh+HlyE+(3 strains),ETT2+(6 strains),F1+HPI+(11 strains).A total of 206 pathogenicE.colistrains were isolated from tissue samples,and the isolates werediv ided into 13 different virulence factors,followed by.F1+(13 strains),F1+ETT2+(39 strains),F1+HlyE+(2 strains),ETT2+HPI+(4 strains),F1+ETT2+HPI+(100 strains),F1+HPI+Tsh+(7 strains),F1+ETT2+HPI+Tsh+(7strains),F1+ETT2+LEE+Tsh+(5 strains),Fl+P+ETT2+Tsh+(4 strains),F1+ETT2+LEE+Tsh+HlyE+(3 strains),Tsh+(7 strains),F1+Tsh+(13 strains),ETT2+Tsh+(2 strains).A total of 15 gene combinations of E.coli isolates from healthy and diseased waterfowl were found by comparison.The top 10 gene combinations were the same,but the ratios were different.The single gene F1+strains inE.colianal swabisolates was up to 63.45%,secondly,F1+ETT2+accounted for 16.37%.The gene combination with the highest proportion in the tissue isolates was F1+ETT2+HPI+,the proportion was as high as 48.54%,F1+ETT2+accounted for 18.93%secondly.In order to study the drug susceptibility of pathogenic E.coli,and provide scientific guidance for clinical drug use,13 antibiotics were used to treat 256 strains of pathogenic E.coli isolated from anal swab samples and tissue samples in Jiangsu Province using drug sensitive paper agar diffusion method.The results showed that the overall clinical health and diseased waterfowl isolates had little difference in sensitivity to 13 drugs,more than 70%of E.coli isolates are resistant to doxycycline and tetracycline,and more than 70%isolates are sensitive to fosfomycin and polymyxin B.Further analysis found that 99.7%of the E.coli isolates were multidrug-resistant,the most common drug resistance combination is doxycycline+tetracycline+neomycin+compound neonomine.The sensitivity of drugs in different regions and different farms is different,but the resistance to doxycycline and tetracycline is universal.In order to study the pathogenicity of E.coliwith different virulence factors,E.colisolate of known virulence factors was subjected to an animal regression test to determine the difference in virulence of the 1-week-old ducklings.The LD50 was determined by one strain of pathogenic E.colisolates with virulence genes F1+,F1+ETT2+and F1+ETT2+HPI+.The results showed that pathogenic E.colisolate with the virulence gene F1+ETT2+HPI+was the most toxic(The LD50 was 2.88×108 CFU/0.5 mL),Followed by F1+ETT2+(The LD50 was 6.92×108 CFU/0.5 mL),the single F1+ pathogenicE.coli isolate had the weakest virulence(The LD50 was 4.28×109 CFU/0.5 mL).The results showed that the virulence of pathogenic E.coli was positively correlated with the number of virulence genes.