Quinolone resistance in Riemerella anatipestifer from Thai ducks: Mutation analysis of gyrA, parC, and plasmid-mediated quinolone resistance genes.

Abstract

Background and aim: Riemerella anatipestifer is a Gram-negative bacterium causing systemic infections in ducks, often treated with quinolones. However, increasing resistance to quinolones poses a threat to effective treatment, and the molecular mechanisms underlying this resistance remain inadequately understood in Thailand. This study aimed to determine the minimum inhibitory concentrations (MICs) of nalidixic acid, ciprofloxacin, and enrofloxacin; identify mutations in the quinolone resistance-determining regions of gyrA and parC; and detect plasmid-mediated quinolone resistance (PMQR) genes in R. anatipestifer isolates from Thai ducks.

Materials and methods: A total of 37 clinical isolates of R. anatipestifer were collected from diseased ducks between 2021 and 2023. MICs were determined using the agar dilution method, following the guidelines of the Clinical and Laboratory Standards Institute. Polymerase chain reaction and Sanger sequencing were employed to detect mutations in gyrA and parC and to screen for PMQR genes (qnrA, qnrB, and qnrS). Phylogenetic analysis of the gyrA gene was performed to assess the relatedness among isolates.

Results: Nalidixic acid MICs ranged from 16 μg/mL to ≥128 μg/mL; ciprofloxacin from 1 μg/mL to 8 μg/mL; and enrofloxacin from 0.25 μg/mL to 4 μg/mL. All isolates had a single point mutation at codon 83 of gyrA, either C248T (Ser83Ile, n = 35) or C248G (Ser83Arg, n = 2). No mutations were observed in parC, and none of the PMQR genes were detected. Phylogenetic analysis grouped most Thai isolates into one major cluster, with a few aligning with Chinese strains and the American Type Culture Collection reference strain.

Conclusion: This study provides the first molecular evidence of quinolone resistance mechanisms in R. anatipestifer from ducks in Thailand. Resistance appears primarily associated with a single mutation at codon 83 of gyrA, while no parC mutations or PMQR genes were detected. These findings highlight the importance of ongoing resistance surveillance and prudent antimicrobial use. Despite limitations in sample size and gene scope, this study provides essential baseline data to inform treatment guidelines and supports the inclusion of R. anatipestifer monitoring in Thailand's national antimicrobial resistance action plan. Future research should explore additional resistance genes using advanced genomic tools.