Optimized In Vitro Antibiotic Susceptibility Testing Method for Small-Colony Variant Staphylococcus aureus

Area of Science:

• Microbiology
• Infectious Diseases
• Antimicrobial Resistance

Background:

• Staphylococcus aureus small-colony variants (SCVs) are linked to persistent infections and poor outcomes.
• Standard antibiotic susceptibility testing (AST) methods are unreliable for SCVs due to poor growth and reversion.
• Predicting SCV antibiotic resistance based on clinical history or related normal-colony (NC) isolates is difficult.

Purpose of the Study:

• To establish reproducible methods for AST of S. aureus SCVs.
• To determine if SCV susceptibilities can be predicted by auxotrophy, treatment history, or isogenic NC isolate data.

Main Methods:

• Evaluated SCV growth and reversion on 11 agar media, selecting two optimal media.
• Performed disk diffusion and Etest AST on 86 SCVs and 28 isogenic NCs.
• Analyzed SCV auxotrophy, NC antibiograms, and patient treatment history for predictive value.

Main Results:

• Brain heart infusion and Mueller-Hinton agars supplemented with hemin, menadione, and thymidine supported optimal SCV growth and stability.
• SCVs showed predictable nonsusceptibility to trimethoprim-sulfamethoxazole or aminoglycosides based on auxotrophy.
• Varied nonsusceptibility was observed for fluoroquinolones, macrolides, and others; mecA-positive SCVs were cefoxitin-resistant.
• All isolates remained susceptible to quinupristin-dalfopristin, vancomycin, minocycline, linezolid, chloramphenicol, and tigecycline.

Conclusions:

• Developed a reliable AST method for S. aureus SCVs using supplemented BHI and MH agars.
• SCV auxotrophy predicts resistance to certain antibiotics, but other resistance patterns are variable.
• Predictive value of treatment history and isogenic NC isolates for SCV susceptibility is limited.
• This validated AST method and findings can guide treatment decisions for challenging SCV infections.