Point-of-Care Air Surveillance of Respiratory Pathogens Using the GeneXpert(R) System

Advances in air-based surveillance of pathogen genetic material are hindered by reliance on centralized, time-consuming molecular techniques. Point-of-care (POC) diagnostic platforms, like the Cepheid(R) GeneXpert(R), offer rapid, simplified testing in clinical settings but have not been evaluated for use with air samples. Here, we paired the ThermoFisherTM AerosolSenseTM air sampler with the Xpert(R) Xpress SARS-CoV-2/Flu/RSV Plus test to evaluate near-real-time air surveillance. To assess analytical sensitivity, we spiked collection substrates with inactivated viruses and performed overnight sampling using the air sampler. As few as 10 copies of influenza A/B (IAV/IBV) and RSV applied to the substrate were detectable by GeneXpert, while SARS-CoV-2 required at least 100 copies for detection. Longitudinal air surveillance was conducted across congregate settings in Columbia, Missouri, and Madison, Wisconsin, in 2024-2025, collecting 281 air samples. SARS-CoV-2 was detected most frequently, followed by IAV. To assess concordance, 191 samples with paired GeneXpert and RT-qPCR results were analyzed across multiple Ct value cutoffs. Agreement between GeneXpert and RT-qPCR for SARS-CoV-2 was fair to moderate (K = 0.306-0.443). Optimal GeneXpert Ct cutoffs for the best balance between sensitivity and specificity, determined using analyses such as Youden's index, were site-specific: 45 for Wisconsin (67% sensitivity, 83% specificity) and 41 for Missouri (76% sensitivity, 62% specificity), reflecting differences in laboratory protocols. For IAV, agreement was moderate (K = 0.56) with GeneXpert Ct cutoff of 40, achieving 85% sensitivity and 81% specificity. Further studies across diverse settings and viral targets are needed to establish GeneXpert's role in routine air surveillance.