Mutations Associated with Decreased Susceptibility to Seven Antimicrobial Families in Field and Laboratory-Derived Mycoplasma bovis Strains

The molecular mechanisms of resistance to fluoroquinolones, tetracyclines, an aminocyclitol, macrolides, a lincosamide, a phenicol, and pleuromutilins were investigated inMycoplasma bovis . For the identification of mutations responsible for the high MICs of certain antibiotics, whole-genome sequencing of 35M. bovis field isolates and 36 laboratory-derived antibiotic-resistant mutants was performed.In vitro resistant mutants were selected by serial passages ofM. bovis in broth medium containing subinhibitory concentrations of the antibiotics. Mutations associated with high fluoroquinolones MICs were found at positions 244 to 260 and at positions 232 to 250 (according toEscherichia coli numbering) of the quinolone resistance-determining regions of thegyrA andparC genes, respectively. Alterations related to elevated tetracycline MICs were described at positions 962 to 967, 1058, 1195, 1196, and 1199 of genes encoding the 16S rRNA and forming the primary tetracycline binding site. Single transversion at position 1192 of therrs1 gene resulted in a spectinomycin MIC of 256 μg/ml. Mutations responsible for high macrolide, lincomycin, florfenicol, and pleuromutilin antibiotic MICs were identified in genes encoding 23S rRNA. Understanding antibiotic resistance mechanisms is an important tool for future developments of genetic-based diagnostic assays for the rapid detection of resistantM. bovis strains.