A first look into the genomic characterization and fluoroquinolone resistance genotypes of Mycoplasma spp. in Colombian poultry

This study provides the first molecular characterization using multilocus sequence typing (MLST) of Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) from Colombian poultry, and examines genotypes associated with fluoroquinolone resistance. A total of 90 pooled DNA samples collected between 2019 and 2023 from breeders, commercial layers, and broilers were analyzed: 30 MG-positive, 30 MS-positive, and 30 with simultaneous MG-MS detection. MLST revealed 32 complete allelic profiles, of which 19 (59.4 %) were novel. For MG, three new sequence types (STs) (ST147, ST152, ST153) were identified, while ST5, linked to the F vaccine strain, was predominant. In MS, over 90 % of STs were new, and two samples corresponded to ST43, previously reported in Eastern Europe. Phylogenetic analyses showed distinct clustering of vaccine and field strains, with MS displaying greater genetic diversity than MG. Antimicrobial resistance (AMR) genotyping targeted the quinolone resistance-determining regions (QRDRs) of gyrA and parC. In MG, substitutions Ser83→Ile (gyrA) and Ser80→Leu (parC) both linked to increased minimum inhibitory concentrations were detected, particularly in novel STs, suggesting circulation of resistant field strains. A His59→Tyr change (gyrA) was also observed in vaccine-related ST5 but is not consistently associated with resistance. In MS, no gyrA mutations associated with AMR were found, but parC Thr80→Ile substitutions were identified in several new STs, indicating potential reduced susceptibility to fluoroquinolones. No mutations were detected at other key QRDR positions in any of both types. These results indicate the spread of field and vaccine strains of MG, along with a high prevalence of various strains of MS, many of which have mutations associated with AMR. The results highlight the importance of molecular surveillance that integrates genotypic and phenotypic information to develop specific control strategies and monitor AMR in poultry. These efforts are essential to mitigate the impact of the disease, limit the spread of resistance, and improve the control measures in Colombia and around the world about micoplasmosis.