Abstract: Objective: In this study, the relationship among methyl-directed mismatch repair (MMR) deficiency, mutation frequency, and the minimal inhibitory concentration (MIC) of ciprofloxacin was investigated to reveal the antimicrobial resistance mechanism of Salmonella hypermutators (SHs). Methods: Ninety SHs were investigated for their susceptibility to ciprofloxacin via agar dilution method. Mutation frequencies were determined by using agar plates containing nalidixic acid and rifampicin. Mutations related to antibiotic resistance were tested by polymerase chain reaction (PCR). The wild-type mutant genes mutS, mutH, mutL, and uvrD were separately transferred into the representative hypermutator 103D2 by electrotransformation. The expression of antibiotic resistance genes in 103D2 and the resulting transformants were tested by real-time PCR so as to determine the effect of MMR gene deficiency on ciprofloxacin resistance. Results: Forty-nine of the 90 SHs (89 of which were ciprofloxacin resistant) were deficient in the MutS (MutS-), and GyrA (67.3%) and ParC (87.8%) mutations were more likely (P = 0.007) to occur in SHs with MutS- compared with the non-MutS wild-type strains. The mutation frequency of the transformants on Luria-Bertani-agar plates with nalidixic acid (20 μg/mL) was significantly (103D2:P-mutS) or extremely significantly (103D2:P-mutL and 103D2:P-uvrD) decreased compared with the original strains. In all transformants, the expression of marA was significantly down-regulated whereas the expression of marR and tolC were significantly upregulated except for 103D2:P-uvrD, leading to deceased ciprofloxacin resistance in 103D2. Conclusion: MMR gene deficiency may be a factor contributing to antibiotic resistance in SHs by affecting the efflux pumps and the expression of marA, tolC, and marR, associated with antibiotic susceptibility.

Key words: Salmonella; hypermutator; antimicrobial resistance; real-time polymerase chain reaction; methyl-directed mismatch repair genes