| Penaeus vannamei is one of the most mature intensive and high-yield aquaculture species in the global industrial chain.While promoting the growth of its output,antibiotics also bring serious hidden dangers.Drug-resistant bacteria and their drug-resistant genes can not only remain in the aquaculture environment,but also spread in the aquatic product supply chain and pass to human beings through the food chain,causing serious harm to public health.Among them,class? integrons play an important role in drug resistance transmission because they can capture,integrate,excise and express drug resistance genes.Escherichia coli is also playing an important role as an important medium and potential source bank for drug resistance gene transmission.Therefore,P.vannamei from different sources such as farms,farmers markets and supermarkets respectively were collected in this study,screening E.coli from the samples and made its drug resistance spectrum,further studied the distribution characteristics of class? integrons of shrimp-derived E.coli and the genetic relationship between class? integron gene cassette positive E.coli isolates,and clarified the drug resistance transmission mechanism of E.coli isolated from P.vannamei mediated by class? integrons.The main research contents and results are as follows:A total of 307 strains of E.coli were isolated from 400 samples of P.vannamei from farms,farmers markets and supermarkets.The isolation rates were 87.6 %,69.0 % and 59.9 % respectively.The isolation rate of E.coli from P.vannamei samples from farms was significantly higher than that from farmers' markets and supermarkets(p < 0.05).The drug resistance spectrum of E.coli to 8 major types of 18 antibiotics(antibacterial drugs)was prepared,and the multi-drug resistance rate was 32.2 %.Among them,E.coli from different sources generally had higher drug resistance rates to tetracycline antibiotics,sulfonamides and ?-lactam cephalosporins,amoxicillin and ampicillin,while the drug resistance rate to imipenem and polymyxin was the lowest.40,43 and 32 strains of class? integron-positive E.coli were isolated from the samples of P.vannamei in farms,farmers markets and supermarkets respectively,of which 6,4 and 6 strains carried class? integron gene cassettes in turn.Sequencing results showed that 87.5 %(14/16)of the class? integron gene cassettes carried drug resistance genes and were related to the spread of drug resistance of aminoglycosides,trimethoprim and sulfanilamide,among which dfrA1-aadA1 was the main gene cassette array.In addition,the multi-drug resistance of E.coli is positively correlated with the class? integron(p < 0.01).The 16 class? integron gene cassette-positive E.coli isolates are divided into 10 PFGE pulse types and 10 ST types.The typing results show that their overall affinity is far away.Although there is a small amount of vertical transmission,horizontal transmission may be mainly carried out by horizontal gene transfer mediated by class? integron.Southern blotting was carried out on the class? integron gene cassette of 16 strains of E.coli isolated from P.vannamei.The results showed that the class? integron gene cassette of 12 strains of E.coli was located on the plasmid,while the other 4 strains were located on the genome.All plasmids carrying class? integron gene cassette of 12 strains of E.coli can obtain conjugon and transformant through conjugation transfer and transformation.The typing results of plasmid replicon showed that all 12 plasmids were IncF plasmids.The flanking sequence of class? integron gene cassette was analyzed by PCR walking.It was found that the class? integron could be mediated by mobile genetic elements such as insertion sequences(IS6 and IS26)and transposon elements(Tn3 and Tn3-like)to carry out horizontal transmission. |
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