Comparing rapid molecular and culture methods for detecting fungal contamination in healthcare environments

Abstract Background: Fungal contamination of healthcare environments is increasingly recognized as a potential contributor to healthcare-associated infections, yet standardized environmental surveillance strategies remain poorly defined. Culture-based methods are widely used but have limitations including low sensitivity and prolonged turnaround time. Molecular approaches such as quantitative PCR (qPCR) may improve detection but have not been extensively evaluated in real-world hospital environments. Methods: We conducted a longitudinal observational study across three inpatient units at a tertiary academic medical center from September 2023 through June 2024. Environmental samples were collected monthly from patient rooms and shared unit areas, including air, bathroom floors, HVAC components, patient bedrails, and linen storage areas. Samples were analyzed using direct-from-sample 18S qPCR and culture-based detection followed by 18S or ITS sequencing. Results: Among 742 samples collected, 474 (64%) were positive for fungi by qPCR compared with 213 (29%) by culture ( P < .01). qPCR showed higher detection rates across most sample types, including air (35% vs 3%), bathroom floors (86% vs 42%), HVAC exhaust vents (72% vs 41%), and patient bedrails (78% vs 10%). Culture methods identified a broader diversity of fungi, including Talaromyces , Candida , and Penicillium , while qPCR detections were dominated by Malassezia . Conclusions: Molecular and culture-based methods provide complementary insights into hospital fungal contamination. qPCR demonstrated greater sensitivity, while culture identified a broader range of viable fungi. Future surveillance strategies may benefit from leveraging qPCR sensitivity using targeted primers for clinically important fungi, reducing the need for broad sequencing and bioinformatic analysis.