Characterization of plasmid-mediated quinolone resistance genes in Salmonella isolates from patients and animals in Thailand

Abstract

Nontyphoidal Salmonella is considered to be acquired from animals to human via food chain. Fluoroquinolones, drug of choice for Salmonella infection has become reduced susceptible and clinical failure has been reported. The chromosomal-mediated fluoroquinolone resistance mechanisms are commonly due to the drug target alterations in gyrA, gyrB, parC and parE genes which can cause high level of resistance. Other resistance mechanism is the presence of plasmid-mediated genes, including qnr genes and aac(6’)-Ib-cr genes which become more clinical concern because of the ability of horizontal gene transfer. This study characterized the plasmid-mediated quinolone resistance genes and investigated the prevalence of qnr genes and aac(6’)-Ib-cr genes in 356 nontyphoidal Salmonella isolated from 108 patients and 248 animals. Prevalence of nalidixic acid resistance in patient isolates was 86.1% and reduced susceptibility to ciprofloxacin was found in 72.2%. High rate of reduced susceptibility was found in S. Choleraesuis (30.56%). Prevalence of nalidixic acid resistance was 42.6%, ciprofloxacin resistance was 0.4% and reduced susceptibility to ciprofloxacin was found in 41.36%. Reduced susceptibility was mostly found in S. Enteritidis (8.06%, 20/248). Screening for the presence of qnr and aac(6’)-Ib-cr in 356 nontyphoidal Salmonella isolates showed that qnrS gene was found in 8.70% (31 isolates) and there was no isolate carrying aac(6’)-Ib-cr. The prevalence of qnrS genes in isolates from patients was 3.7% (4/108) which were found in S. Choleraesuis (1 isolate) and S. group D (3 isolates). The prevalence of qnrS genes in isolates from animals was 10.88% (27/248) and the most common serovar carrying qnr genes was S. Anatum (9 isolates). This study was the first report of the prevalence of qnrS in nontyphoidal Salmonella isolated from Thailand. DNA sequencing analysis of the qnrS gene of all 31 qnrS-positive revealed 100% nucleotide and amino acid identity to qnrS1 and QnrS1 submitted in GenBank. The in vitro selection of ciprofloxacin resistance demonstrated that 4 out of 5 qnrS1-positive parent strains raised ciprofloxacin MIC to 32-64 µg/ml in the third-generation selection whereas 2 out of 5 qnrS1-negative parent strains raised ciprofloxacin MIC to 32 µg/ml. Single amino acid substitutions in GyrA were found at S83F, Y and D87G when the ciprofloxacin MIC of mutants increased to > 1 µg/ml. The double mutations at S83 and D87 in GyrA led to higher ciprofloxacin MIC. However, there was an increase in ciprofloxacin MIC without any mutation in GyrA, suggesting that there were other mechanisms involved in the development of resistance such as mutations in QRDR of ParC, overexpression of efflux system and decreased outer membrane porins.