Genetic characterization, antimicrobial susceptibility, and pathogenicity of avian Mycoplasma synoviae strains

Abstract Mycoplasma synoviae ( M. synoviae ) infection remains a major cause of economic losses in intensive poultry production worldwide. The increased incidence of infections by M. synoviae has facilitated the emergence of diverse field strains. Characterizing the biological features of currently circulating strains will provide data support for region-specific disease control. In this study, 11 M. synoviae field strains were isolated from Northern China between 2018 and 2023 and comprehensively analyzed their genetic genotypes, antimicrobial susceptibility profiles, and potential resistance-associated mutations. Additionally, four representative strains (3S, GJZ, JB913, and YFS) were evaluated for pathogenicity through four inoculation routes: intravenous, footpad, intranasal, and intratracheal administration. Multilocus sequence typing (MLST) assigned the 11 isolates to six distinct sequence types (STs), including four STs (ST34, ST93, ST151 and ST153) exclusively found in China and two previously unreported novel STs (ST273 and ST274). Antimicrobial susceptibility assays demonstrated that most strains had elevated minimum inhibitory concentrations (MICs) against enrofloxacin and florfenicol, whereas most strains showed low MICs to tilmicosin, tylvalosin, tylosin, oxytetracycline, doxycycline, lincomycin, and tiamulin. Analysis of quinolone resistance-determining regions (QRDRs) presented a strong association between high enrofloxacin MICs (MIC ≥ 2 µg/mL) and specific amino acid mutations, including ParC Thr85Ile and GyrB Ser417Asn substitutions, as well as a putative ParE Pro446Ser point mutation based on M. synoviae reference numbering. Pathogenicity assays showed that intravenous and footpad inoculation of the four tested strains consistently induced severe systemic pathological lesions, including footpad/joint swelling and keel bursa edema, while the YFS strain additionally caused airsacculitis. In contrast, intranasal and intratracheal inoculation only caused negligible or no observable tissue damage. Notably, the GJZ strain induced significant upregulation of pro-inflammatory cytokines (IL-6, IL-17 A) and immunomodulatory cytokines (IL-4, IFN-γ) following intranasal inoculation, without evidence of clinical disease. This immunogenic yet attenuated phenotype is comparable to that of the MS-H vaccine strain administered via eye drops, indicating that GJZ holds promising potential as a novel vaccine candidate. Overall, this study provides a basic characterization of M. synoviae field strains from Northern China and identifies a single strain with favorable characteristics for further vaccine development research.