Assessment of bioaerosol sampling techniques for viable Legionella pneumophila by ethidium monoazide quantitative PCR

Abstract

Legionella pneumophila causes severe pneumonia and Pontiac fever in humans. Rapid and sensitive bioaerosol monitoring techniques for viable L. pneumophila are unavailable. Coupled with a newly developed viable assay called ethidium monoazide with quantitative PCR (EMA-qPCR), this study applies EMA-qPCR to aerobiology for the first time to evaluate the effects of the method of sampling (all-glass impinger (AGI-30), BioSampler, and MAS-100 sampler) and sampling time (3, 30, 60 min) on the collection of viable L. pneumophila. The effects of the collection fluid (deionized water (DW) and Tween mixture) and the replenishment of DW every 15 min during 60-min sampling were also assessed. Escherichia coli, as a model microorganism in bioaerosol research, was also tested. Using the Tween mixture (DW containing 1% peptone, 0.01% Tween 80, and 0.005% antifoam), the AGI-30 and BioSampler performed significantly better than the MAS-100 sampler for collecting viable L. pneumophila and viable E. coli (P < 0.05). An increase in sampling time adversely affected the quantification of both bacterial species (P < 0.05). The collection with DW yielded greater recovery of viable L. pneumophila than the Tween mixture in both AGI-30 and BioSampler, regardless of sampling time, by a factor of 1.4-6.9 (P < 0.05). The replenishment of DW every 15 min further improved the collection of viable L. pneumophila. This study demonstrates that viable L. pneumophila can be efficiently sampled by the AGI-30 and BioSampler and successfully quantified by EMA-qPCR. Copyright © American Association for Aerosol Research.