The Analysis On Antimicrobial Resisitance And PFGE Molecular Typing Of Salmonella Enteritidis

Objectives1.Analyze and study the characteristics of Salmonella enteritidis resistance from human,food and animal in JiangSu province from 2012 to 2015,to guide clinicians for using antibiotics rationally and provide scientific reference for preventing and controlling the infectious disease caused by Salmonella enteritidis.2.To analyze the status of the resistantance genes,the relationship between phenotype and genetype,and explore the cause of multi-drug resistance mechanism of Salmonella enteritidis from different sources,according to the results of resistance phenotype and resistance genes(?-lactamase,aminoglycoside,chloramphenicol,sulfamethoxazole/trimethoprim tablet,tetracycline and quinolone/fluoroquinolone resistance).3.Applying the PFGE,to compare of different sources of Salmonella enteritidis homology,make epidemiological analysis,find the main molecular types,and study the relationship between molecular type with phenotype and genotype.It is possible to provide scientific reference for outbreak surveillance and systematic analysis of epidemiology basing on the results above.Methods1.Ninety-three isolates were isolated and collected from the Salmonella baseline surveys of food animals,the diarrhea patients of foodborne diseases,and the human carriers of food employees,according to the criterion of national food sanitation examination method(GB4789—2010)during 2012 to 2015.All isolates were tested for resisitance to 16 antimicrobial agents including ?-lactamase(ampicillin,amoxicillin/clavulanic acid,cefotaxime,cefoxitin,cefepime,cefoperazone,ceftriaxone),aminoglycoside(gentamicin,kanamycin,amikacin,streptomycin),chloramphenicol,sulfamethoxazole,tetracycline and quinolone/fluoroquinol(nalidixic acid,ciprofloxacin)with the Kirby-Bauer(K-B)method and judged by CLSI 2013.2.All isolates were tested 29 resistantance genes,?-lactamase(blaTEM-1-like,blaCTX-M,blaOXA-1-like,blacMY-2,blaPSE-1,blaSHV),sulfamethoxazole(sull,sul2,sul3),chloramphenicol(catA1,cmlA1,floR),trimethoprim(dfvAl-like,dfrA12,dfrA17),aminoglycoside(aadA1-like,aaclA2,aadB,aacC4,aac(6')-1b),tetracycline(tet(A),tet(G),tet(B),tet(C)and quinolone/fluoroquinolone(qnrA,qnrB,qnrS,qnrC,qnrD)by PCR.3.All isolates were performed with the international PulseNet standard operating program of Salmonella,Salmonella PFGE sub typing.Xbal was used as the restriction endonuclease digestion.Gel image analysis system generated the image bands of PFGE.National pathogen identification network software was adopted to process and analysis image strips.Results1.Phenotypic characterization of antimicrobial resistance showed that Salmonella enteritidis isolates most often displayed resistance to nalidixic acid(93.55%),ampicillin(74.19%),streptomycin(65.59%),cefoperazone(51.61%).tetracycline(49.46%),and 22 antimicrobial resistance profiles were identified for the 16 antimicrobial agents.Certain recurring antimicrobial resistance profiles were evident,namely,AMP-CFP-STR-NA-TE(21/93),AMP-STR-NA-TE(15/93),AMP-CFP-STR-NA(10/93),resistant to three or more antimicrobials(MDR)isolates was 75.27%(70/93).2.Resistant genes by PCR,sul2 and sul3 were the high carried genes with 80%,respectively,blaTEM-1-like(75.27%),tet(A)(40.86%),tet(G)(35.4%)and aac(6')-1b(9.8%)were followed.Sul1 was related to sulfamethoxazole resistance phenotype with the coincidence of 93.55%,while bla-TEM-I-like gene was associated with resistance phenotype of ampicillin with the coincidence of 91.40%,and tetracycline resistance phenotype was related to tet(A)/tet(G)genes with the coincidence of 66.67%.3.PFGE molecular types,93 Salmonella enteritidis isolates were divided into two clusters,A and B by PFGE.B19 and B23 were the main sub-types,in which the strains were isolated from different areas and different host sources such as chicken,human and food.The strains of B19 were resistant to AMP-CFP-STR-NA-TE mainly,carrying the resistance genes,blaTEM-1-like,sul2,sul3 and tet(A),and the strains of B23 were resistant to AMP-CFP-NA-TE mainly,carrying the resistance genes,blaTEM-1-like,sul2,sul3,tet(A)and tet(G).Conclusions1.Salmonella enteritidis isoaltes were resistant to nalidixic acid,ampicillin,streptomycin mostly.The status of Samonella enteritidis resistance,especially multi-drug resistance was serious,most of which were resisitant to 4 kinds of antibiotics.Some fixed resistant patterns were formed such as AMP-CFP-STR-NA-TE,AMP-STR-NA-TE,AMP-CFP-STR-NA.2.The antibiotic resistance genes sul2,sul3,blaTEM-1-like,tet(A)and tet(G)were occured commonly in Salmonella enteritidis isolates.There was some correlation between the resistance phenotype and resistance genes of Salmonella enteritidis.The gene cluster blaTEM-1-like/blaCTx-M-tet(A)/tet(G)-sul1-aac(6')-1b could be the main cause of MDR Salmonella enteritidis.3.The results of PFGE showed that the differentiation degree of Salmonella enteritidis was not high.The main molecular types were B19 and B23,of which the strains were highly similar with the phenotypes and genotypes.Results showed that the strains from different origins and sources could have the same PFGE profiles,which suggested that the isolates of the same clone could spread among different hosts and different regions.