Bacitracin Susceptibility Testing 

Bacitracin is a cyclic polypeptide antibiotic primarily active against Gram-positive bacteria, including Staphylococcus spp., Streptococcus spp., and Clostridium spp. It functions by inhibiting bacterial cell wall synthesis through interference with undecaprenyl pyrophosphate, a key lipid carrier in peptidoglycan biosynthesis. Due to its nephrotoxicity, bacitracin is mainly used topically or in selective decontamination regimens.

Testing Methods and Standards

The susceptibility of bacitracin is evaluated using multiple standardized methodologies:

• Disk Diffusion Method:
  • Utilizes a 10-unit bacitracin disk applied to Mueller-Hinton agar (MHA) or blood agar.
  • Incubation at 35°C for 18-24 hours.
• Broth Microdilution (BMD) Method:
  • The gold standard for quantitative determination of minimum inhibitory concentration (MIC) values.
  • Conducted in cation-adjusted Mueller-Hinton broth (CAMHB) with serial dilutions of bacitracin.
• Etest (Gradient Strip) Method:
  • Employs a strip with a continuous gradient of bacitracin.
  • Used for MIC determination, particularly in vancomycin-resistant enterococci (VRE) and coagulase-negative staphylococci.

Application and Clinical Relevance

• Identification of Group A Streptococcus (S. pyogenes):
  • The bacitracin disk test remains a crucial method for distinguishing S. pyogenes (susceptible) from other β-hemolytic streptococci (resistant).
• Surveillance of Bacitracin Resistance in Pathogens:
  • Increasing resistance in Staphylococcus aureus and Enterococcus spp. necessitates susceptibility testing for accurate resistance profiling.
• Guiding Antimicrobial Stewardship Programs:
  • Helps optimize topical antibiotic selection and minimize overuse in decolonization protocols.
  • Avoids empirical use in systemic infections where bacitracin has limited clinical application.
• Potential for Combination Therapy in MDR/XDR Infections:
  • Used in combination regimens for selective gut decontamination in critically ill patients.
  • Investigated as an alternative for topical MRSA decolonization.

Bacitracin susceptibility testing remains an essential component of diagnostic microbiology, particularly for streptococcal identification, resistance monitoring, and targeted antimicrobial therapy.