| Enterobacteriaceae is the most common conditional pathogens widely distributed in the environment of the duck slaughterhouse.Antibiotics have been widely used in livestock and poultry breeding environment as growth promoters and infectious drugs,but the abuse of antibiotics has caused serious bacterial resistance,which poses a great threat to livestock and poultry breeding and public health.The purpose of this study was to explore the bacterial epidemic and multidrug resistance on the production chain of a duck slaughterhouse in Zhejiang province,and to explore the mechanism of carbapenems producing plasmids mediating the transmission of drug resistance of Enterobacteriaceae,and to study the phenotype and genetic structure of the multidrug-resistant plasmids contained in them through bioinformatics.(1)The samples such as duck’s feces,soil,slaughterhouse utensils,workers’ feces and insects were collected from duck slaughterhouses,and were separated and purified,the detection rate of duck carcasses and slaughter utensils in the slaughtering process was 38.6% and 56.0% respectively.The total number of bacterial colonies in the samples were tested,Enterobacteriaceae were the main genus,E.coli,proteus mirabilis and Klebsiella pneumoniae were the main bacteria species identified by 16 S rRNA,and 108,29 and 38 strains were identified,respectively.Most isolates in duck feces,slaughterhouse and insect samples were E.coli,which was speculated that E.coli was the dominant bacteria in the duck slaughter chain.(2)K-B and multiple RPA assay were used to detect the resistance phenotype and genotype,and results showed that the apparent resistance of strains to tetracycline,amoxicillin,and compound sulfamethoxazole and cephalosporin antibiotics were as high as 90% above,the resistant phenotype was almost the same to genotype,9 categories of resistant genes were all detected,the multidrug resistant bacteria in premortem management were concentrated in duck feces and soil samples,with a resistance of 18 or more,mainly to tetracycline,fosfomycin and aminoglycoside.There were more than 15 multidrug resistant bacteria in carcass and slaughterhouse,mainly to tetracycline,fosfomycin and chloramphenicol.Tetracycline and fosfomycin resistance genes were detected and widely distributed throughout the slaughter chain,as well as in insects and workers in the environment.including Carbapenems drug-resistant genes in duck’s feces and slaughtering equipment isolated strains were found in samples,but the class of antibiotics in livestock farming have disabled,so these strains would be as subsequent analysis.Through Carba NP and plasmid conjugation transfer assay,the drug resistance genes of Carbapenems on the plasmid were verified,and the transmissibility of drug resistance genes was confirmed.(3)4 plasmids from 2 strains which producing carbapenems were sequenced and the obtained sequences were carefully annotated by second-generation high-throughput technology.Plasmids p11014-IMP and p10229-IMP belonged to the IncA/C2 and IncHIA types respectively,they both contained the blaIMP-3 and in downstream of type 1 integrase Intl1,and the MDR regions had a high degree of homology.p11014-CTXM belonged to IncFⅡ type,ISEcp1 promoted blaCTX-M-55 to move,and provided a strong promoter to drug resistance gene expression and improved the transfer efficiency.p10229-NDM belonged to IncX3 type,ISAba125 caused the rapid diffusion of blaNDM-1,and combined with IS26 and Tn125,blaNDM-1 was recombined and integrated into the plasmid.The MDR region of the 4 plasmids fully indicated that the backbone region would integrate and eventually make expression of the drug-resistant genes after the mobile elements carry them into plasmids.Such resistance mechanism enabled the drug-resistant genes to be continuously transmitted and transferred between strains and spreaded in a large range.The study showed that the existence of multi-drug resistant bacteria will accelerate the evolution of strains.Under the pressure of antibiotic,strains keep acquiring drug resistant genes,and keep spreading,so the situation of global drug resistance will be more serious. |
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