Screening And Mechanism Of Quinolone Resistance In Foodborne Salmonella Isolates

The emergence and spread of antimicrobial-resistant Salmonella is one of the greatest concerns in public health.Continuous monitoring of antimicrobial-resistant Salmonella in food is important for food safety.Quinolones were used as first-line drugs in the treatment of Salmonella infection.It is of great significance to control the epidemic of quinolone-resistant Salmonella based on understanding the mechanisms on resistance formation.A total of 143 Salmonella isolates collected from retail foods in Shanghai between 2016-2017 were screened for their resistance to antimicrobials.The main contents and results of this study were as follows:1.Antimicrobial susceptibilities of 143 isolates to antimicrobials were determined using agar dilution method.Among 143 Salmonella isolates,a total of 69(48.2%)isolates showed resistance to at least three classes of antimicrobials and 31(21.7%)were resistant to at least five classes of antimicrobials.Isolates from frozen convenience food(92.9%)and ready-to-eat food(68.4%)showed higher multidrug-resistance rates compared with that from fresh meat(37.4%)(P<0.05).With regard to quinolones,the resistance rate of 143 isolates to nalidixic acid,ciprofloxacin and ofloxacin was 51.0%,25.9%and 14.7%,respectively.ST11(45.5%)and ST34(18.9%)were prevalent types,moreover,a total of 27 ST34 isolates were multidrug-resistant S.Typhimurium,carrying quinolone-resistant phenotype.All ST34isolates were further analyzed by PFGE,and 16 isolates exhibited a similarity of 100%in their PFGE patterns.2.The presence of antimicrobial resistance genes in Salmonella isolates resistant to each class of antimicrobials and mutations in quinolone resistance-determining region(QRDR)of quinolone-resistant isolates were identified.All?-lactamase-resistant isolates carried bla_TEM-1;aac(6')-Ib(100%)and aad A(95.7%)were identified in aminoglycoside-resistant isolates;cml A(100%)and flo R(100%)were carried by chloramphenicol-resistant isolates;tet A(97.4%),tet B(97.4%)and tet C(15.8%)were found in tetracycline-resistant isolates.In quinolone-resistant isolates,plasmid-mediated quinolone resistance(PMQR)gene:aac(6')-Ib-cr(100%),oqx AB(83.3%)and qnr S1(21.8%)were identified.Moreover,QRDR mutation was observed in 71(91.0%)quinolone-resistant isolates,mainly in Gyr A(85.9%),and the mutation types included Asp87Asn(51.3%)and Asp87Tyr(34.6%).Additionally,7 isolates did not harbor any QRDR mutation,among which,4 isolates showed resistance to ciprofloxacin while susceptibility to nalidixic acid,and it was speculated that RND efflux pumps might play a major role in these isolates.3.The contribution of RND efflux activities to ciprofloxacin resistance in Salmonella was evaluated by the inhibition of RND efflux pumps and the construction of the pump repressor gene ram R deletion mutant.Twenty-two isolates showing resistance to ciprofloxacin were selected.The MIC of ciprofloxacin to Salmonella was compared before and after the treatment with PA?N(an inhibitor of RND efflux pumps),and results indicated a decline in MIC in all isolates,ranging from 2 to 32 folds.One S.Enteritidis isolate SJTUF 13552 with no QRDR mutation was therefore chosen for the deletion of ram R gene.The quinolone-resistant level and relative expression level of RND efflux pump genes(acr B,acr D,acr F,mdt B and mds B)and regulate gene ram A were then determined.It was found that the ciprofloxacin and nalidixic acid MIC of the mutant were same as the wildtype strain.Relative expression level of all tested genes had no significant change(P>0.05).There might be ram R-independent pathways to regulate RND efflux pumps.In conclusion,Salmonella isolates from retail foods showed high resistance rate to quinolone.Quinolone-resistant isolates had high mutation rate in QRDR,and the generation of resistance in quinolone-resistant Salmonella may be caused by activities of RND efflux pumps.