Performance Characteristics of a Real-Time PCR Assay for Direct Detection of Streptococcus pneumoniae in Clinical Specimens

Community-acquired pneumonia and complications, such as bacteremia and meningitis due to Streptococcus pneumoniae infection, still occur in at-risk populations, despite the availability of effective vaccines. Laboratory confirmation of S. pneumoniae remains challenging despite advances in blood culture techniques and the availability of nucleic acid-amplification tests. The goal of this study was to determine the performance characteristics of a molecular assay designed as a diagnostic test using primary clinical specimens for invasive pneumococcal disease. The molecular assay adapted for the Luminex Aries instrument targets an S. pneumoniae-specific gene (autolysin, lytA) in clinical specimens. Using real-time PCR MultiCode technology, four different clinical specimen types were evaluated. Specimen types included bronchoalveolar lavage, whole blood, cerebrospinal fluid, and urine to cover the various presentations and appropriate specimen types for invasive pneumococcal infections. The lower limit of detection in urine was 10 colony forming units (CFU)/mL, while in bronchoalveolar lavage, cerebrospinal fluid, and whole blood, it was 100 CFU/mL. Accuracy and specificity were both 100%, and all specimen types were stable for 8 days at 4 degrees C. Finally, 38 clinical specimens were tested to further evaluate the assay. The performance characteristics met Clinical Laboratory Improvement Amendments standards for a clinical diagnostic assay, and the assay offers a sensitive and specific realtime PCR test for direct detection of S. pneumoniae in relevant clinical specimens. (J Mol Diagn 2024, 26: 552-562; https://doi.org/10.1016/j.jmoldx.2024.03.009)