Contribution of Enhanced Efflux to Reduced Susceptibility of Salmonella enterica Serovar Choleraesuis to Fluoroquinolone and Other Antimicrobials

Ministry of Agriculture, Forestry and Fisheries, Tokyo, Japan.
Journal of Veterinary Medical Science (Impact Factor: 0.78). 02/2011; 73(2):279-82. DOI: 10.1292/jvms.10-0309
Source: PubMed


We examined antimicrobial susceptibility and efflux systems in laboratory-derived mutants of Salmonella enterica serovar Choleraesuis selected by culture on fluoroquinolone-containing plates. The mutants exhibited decreased susceptibilities to quinolones and several other antimicrobials. Mutations in the gyrA gene were not always found in the mutants. Accumulation assays revealed that intracellular enrofloxacin concentrations were significantly lower in the mutants compared with parent isolates. Increased expression of acrB mRNA can explain the decreased susceptibilities to several antimicrobials but not in the case of carbonyl cyanide m-chlorophenylhydrazone (CCCP). Decreased susceptibility to CCCP may result from the increased expression of emrA mRNA. These results suggest that the enhancement of multiple efflux pumps is responsible for decreased susceptibilities to several antimicrobials in the laboratory-derived mutants.

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    • "acrA-R GTACCGGACTGCGGGAATT Usui et al. (2011) acrB-rt1 GGCATTGGGTATGACTGGAC Zheng et al. (2009) acrB-rt2 GCATTACGGAGAACGGGATAG Zheng et al. (2009) ramA-rt1 TTTCCGCTCAGGTTATCGAC Zheng et al. (2009) ramA-rt2 CGGGCAATATCATCAATACG Zheng et al. (2009) soxS-rt1 AAATCGGGCTACTCCAAG Zheng et al. (2009) soxS-rt2 TACTCGCCTAATGTTTGATG Zheng et al. (2009) marA-rt1 ATTCTCTATCTGGCGGAAC Zheng et al. (2009) marA-rt2 CGGGTCAATGTTTGCTGTG Zheng et al. (2009) robA-rt1 TATTCCGCCAGTGCTTTATG Zheng et al. (2009) robA-rt2 CCTGCTCATCGTCTTTCTCC Zheng et al. (2009) 16S rRNA-F CCAGCAGCCGCGGTAAT Usui et al. (2010) 16S rRNA-R TTTACGCCCAGTAATTCCGATT Usui et al. (2010) "
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    ABSTRACT: Understanding the impact of antimicrobial use on the emergence of resistant bacteria is imperative to prevent its emergence. For instance, activation of the AcrAB efflux pumps is responsible for the emergence of antimicrobial-resistant Salmonella strains. Here, we examined the expression levels of acrB and its multiple regulator genes (RamA, SoxS, MarA, and Rob) in 17 field isolates of S. Choleraesuis by using quantitative PCR methods. The expression of acrB increased in 8 of the field isolates (P < 0.05). The expression of acrB was associated with that of ramA in 1 isolate, soxS in 1 isolate, and both these genes in 6 isolates. Thereafter, to examine the effect of selected antimicrobials (enrofloxacin, ampicillin, oxytetracycline, kanamycin, and spectinomycin) on the expression of acrB and its regulator genes, mutants derived from 5 isolates of S. Choleraesuis were selected by culture on antimicrobial-containing plates. The expression of acrB and ramA was higher in the mutants selected using enrofloxacin (3.3–6.3- and 24.5–37.7-fold, respectively), ampicillin (1.8–7.7- and 16.1–55.9-fold, respectively), oxytetracycline (1.7–3.3- and 3.2–31.1-fold, respectively), and kanamycin (1.6–2.2- and 5.6–26.4-fold, respectively), which are AcrAB substrates, than in each of the parental strains (P < 0.05). In contrast, in AcrAB substrate-selected mutants, the expression of soxS, marA, and rob remained similar to that in parental strains. Of the 4 antimicrobials, the level of ramA expression was significantly higher in the enrofloxacin- and ampicillin-selected mutants than in the oxytetracycline- and kanamycin-selected mutants (P < 0.05), whereas the expression levels of acrB and multiple regulator genes in spectinomycin-selected mutants were similar to those in each parental strain. These data suggest that exposure to antimicrobials that are AcrAB substrates enhance the activation of the AcrAB efflux pump via RamA, but not via SoxS, MarA, or Rob in S. Choleraesuis.
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