The rise in antimicrobial resistance (AMR) in Mycoplasma bovis underscores the urgent need for alternative treatments. This study evaluated the minimal inhibitory concentrations (MICs) of four metal ions (cobalt, copper, silver, and zinc) and colloidal silver against 15 clinical M. bovis isolates, alongside conventional antimicrobials (florfenicol, tetracycline, tulathromycin, and tylosin). Colloidal silver demonstrated the most effective antimicrobial activity, inhibiting 81.25% of isolates at 1.5 mg/L, while silver inhibited 93.7% of isolates at concentrations above 1.5 mg/L. Copper exhibited notable efficacy, inhibiting 37.5% of isolates at 1.5 mg/L, with a small proportion responding at 0.1 mg/L. Cobalt and zinc displayed variable activity, with MIC values ranging from 0.7 to 12.5 mg/L. In contrast, conventional antimicrobials showed limited effectiveness: tetracycline inhibited 31.25% of isolates at ≥16 mg/L, tylosin inhibited 25% at 16 mg/L, and tulathromycin MICs ranged from 0.5 to 8 mg/L. Time–kill assays revealed a reduction in M. bovis viability after eight hours of exposure to silver and colloidal silver, though higher concentrations (4×–8× MIC) were required for complete eradication. These findings highlight the significant potential of colloidal silver and copper as alternatives for treating M. bovis infections and combating AMR. Further research is essential to explore their standalone and synergistic applications for therapeutic use.

Mycoplasma hyopneumoniae, an important cause of enzootic pneumonia in pigs in many countries, has recently been shown to exhibit reduced susceptibility to several antimicrobial classes. In the present study, a total of 185 pig lung tissue samples were collected from abattoirs in Australia, from which 21 isolates of M. hyopneumoniae were obtained. The antimicrobial resistance profile of the isolates was determined for 12 antimicrobials using minimum inhibitory concentration (MIC) testing, and a subset (n = 14) underwent whole-genome sequence analysis. MIC testing revealed uniformly low values for enrofloxacin (≤1 μg/mL), florfenicol (≤8 μg/mL), lincomycin (≤4 μg/mL), spectinomycin (≤4 μg/mL), tetracycline (≤0.5 μg/mL), tiamulin (≤2 μg/mL), tildipirosin (≤4 μg/mL), tilmicosin (≤16 μg/mL) tulathromycin (≤2 μg/mL), and tylosin (≤2 μg/mL). Higher MICs were observed for erythromycin (MIC range: 16–32 μg/mL), gamithromycin, and tilmicosin (MIC range of both: 32–64 μg/mL). Whole-genome sequencing of the isolates and additional screening using mismatch amplification mutation assay PCR did not identify any known genetic resistance markers within 23S rRNA (macrolides), DNA gyrase A, and topoisomerase IV genes (fluoroquinolones). The WGS data also indicated that the Australian M. hyopneumoniae isolates exhibited limited genetic diversity and formed a distinct monophylectic clade when compared to isolates from other countries. These findings indicate that Australian M. hyopneumoniae likely remains susceptible to the major antimicrobials used to treat enzootic pneumonia in pigs and have evolved in isolation from strains identified in other pig-producing countries.