Characterization and expression of the gyrA gene from quinolone resistant Yersinia ruckeri strains isolated from Atlantic salmon (Salmo salar L.) in Norway

Abstract

Yersinia ruckeri, the causative agent of yersiniosis has been reported in a number of fish species but the most vulnerable are the salmonids including Atlantic salmon (Salmo salar L.). In the present study Y. ruckeri isolates were collected from diseased Atlantic salmon juveniles and characterized for resistance against quinolones. Isolates were screened using disk diffusion assays and MIC determination. The QRDR regions of the gyrA, gyrB, parC and parE genes were sequenced. Quinolone resistant isolates revealed a single bp mutation which replaced serine by arginine at position 83 in the GyrA protein while no mutations were found in gyrB, parC and parE genes. Isolates were also screened for plasmid encoded qnrA, qnrB and qnrS genes but they were found absent. Cloning of gyrA from susceptible and resistant isolates into heterologous Y. ruckeri was not successful. The different gyrA alleles from susceptible and resistant isolates of Y. ruckeri were cloned into Escherichia coli TOPO 10 and E. coli DH5 alpha. While cloning of the resistant allele into the sensitive host had no effect, cloning of the quinolone susceptible gyrA allele into quinolone resistant E. coli DH5 alpha increased the inhibition zone diameter from 25 mm to 38 mm and decreased the MIC from 4 mu g/ml to 2 mu g/ml, suggesting dominance of wild type gyrA over the mutant allele. It is assumed that the wild type GyrA protein has more affinity to form the gyrase-DNA complex than mutant GyrA even in the presence of high levels of the mutated enzyme. (C) 2012 Elsevier B.V. All rights reserved.