A Specimen-level Analysis of Polymicrobial Bacterial and Viral Detections in Hospital-acquired Pneumonia Using Multiplex PCR

Abstract

Background: Hospital-acquired pneumonia (HAP), including ventilator-associated pneumonia (VAP), remains a significant cause of morbidity and mortality among critically ill patients. Conventional culture-based microbiological methods are often time-consuming and may underestimate microbial burden, particularly in patients who have received prior antibiotic therapy. Multiplex molecular diagnostic platforms such as the BioFire® FilmArray Pneumonia Panel (BFPP) enable rapid pathogen detection and may provide a more comprehensive understanding of microbial complexity in HAP. This study aimed to characterize specimen-level microbial patterns in ICU patients with suspected HAP, focusing on polymicrobial bacterial detections, viral co-detection, and antimicrobial resistance-gene profiles.

Methods: This secondary exploratory analysis utilized data from the intervention arm of a previously conducted single-center randomized controlled trial performed in an adult anesthesia ICU at a tertiary government teaching hospital in Kanpur, India, between January 2020 and October 2021. Thirty respiratory specimens from patients with suspected HAP were analyzed using BFPP. Bacterial targets detected at a semi-quantitative threshold of ≥10⁶ copies/mL were considered clinically significant. Outcomes assessed included bacterial detection, number of bacterial targets per specimen, polymicrobial detection, viral detection, bacteria–virus co-detection, virus-only detection, and resistance-gene detection.

Results: Of the 30 specimens analyzed, 21 (70.0%) demonstrated at least one clinically significant bacterial target, while 9 (30.0%) showed none. Polymicrobial bacterial detection was observed in 13/30 specimens (43.3%) and in 13/21 bacteria-positive specimens (61.9%). A total of 43 bacterial detections were recorded, with the most common pathogens being Acinetobacter calcoaceticus–baumannii complex, Pseudomonas aeruginosa, Klebsiella pneumoniae group, Escherichia coli, and Staphylococcus aureus. Viral targets were identified in 7/30 specimens (23.3%), including bacteria–virus co-detection in 4/30 (13.3%) and virus-only detection in 3/30 (10.0%) specimens. Six resistance-gene detections were recorded. BFPP demonstrated 100% positive percent agreement and 92.9% negative percent agreement compared with conventional culture methods.

Conclusion: BFPP frequently detected complex microbial patterns in ICU patients with suspected HAP, including substantial rates of polymicrobial bacterial infection and clinically relevant bacteria–virus co-detection. Rapid multiplex molecular testing may facilitate more precise interpretation of lower respiratory tract infections and support earlier, antimicrobial stewardship-guided clinical decision-making.