Detection of viable Staphylococcus aureus/methicillin-resistant Staphylococcus aureus and antimicrobial resistance directly by PMA-TaqMan qPCR from bronchoalveolar lavage fluid

Timely detection of viable Staphylococcus aureus (MSSA)/ methicillin-resistant Staphylococcus aureus (MRSA) and antimicrobial resistance are crucial for the treatment of patients with pulmonary infection. In the current study, we have developed and evaluated the potential clinical applicability of the PMA TaqMan-based real-time PCR (PMA-TaqMan qPCR) assay for detection of viable MSSA/MRSA and antimicrobial susceptibility directly from bronchoalveolar lavage fluid. The PMA-TaqMan qPCR assay demonstrated high sensitivity with a detection limit of 800 CFU/mL within 115 min. Furthermore, the assay required only 45 min of antibiotic exposure and completed antimicrobial susceptibility testing within 160 min. Upon detection of 10 spiked bronchoalveolar lavage fluid samples treated by clindamycin after 24 h, the PMA-TaqMan qPCR assay showed better agreement with culture-based methods in detecting viable bacteria. Subsequent antimicrobial susceptibility results of bronchoalveolar lavage fluid samples spiked by 16 MRSA isolates and 11 MSSA isolates based on the current assay, compared with the standard broth microdilution method, showed an overall agreement of 100% (95% confidence intervals [CI]: 80.64 to 100) for MRSA and 90.91% (95% CI: 62.27 to 98.38) for MSSA. Our findings demonstrate that the PMA-TaqMan qPCR assay is a highly sensitive and specific method for the direct detection of viable MSSA/MRSA and antimicrobial susceptibility testing from bronchoalveolar lavage fluid.