Bioaerosol sampling for the detection of aerosolized influenza virus

Abstract

Background: Influenza virus was used to characterize the efficacy of a cyclone-based, two-stage personal bioaerosol sampler for the collection and size fractionation of aerosolized viral particles.

Methods: A Collison single-jet nebulizer was used to aerosolize the attenuated FluMist vaccine into a calm-air settling chamber. Viral particles were captured with bioaerosol samplers that utilize 2 microcentrifuge tubes to collect airborne particulates. The first tube (T1) collects particles greater than 1.8 microm in diameter, while the second tube (T2) collects particles between 1.0 and 1.8 microm, and the back-up filter (F) collects submicron particles. Following aerosolization, quantitative PCR was used to detect and quantify H1N1 and H3N2 influenza strains.

Results: Based on qPCR results, we demonstrate that aerosolized viral particles were efficiently collected and separated according to aerodynamic size using the two-stage bioaerosol sampler. Most viral particles were collected in T2 (1-1.8 microm) and on the back-up filter (< 1 microm) of the bioaerosol sampler. Furthermore, we found that the detection of viral particles with the two-stage sampler was directly proportional to the collection time. Consequently, viral particle counts were significantly greater at 40 minutes in comparison to 5, 10 and 20 minute aerosol collection points.

Conclusions: Due to a lack of empirical data, aerosol transmission of influenza is often questioned. Using FluMist, we demonstrated that a newly developed bioaerosol sampler is able to recover and size fractionate aerosolized viral particles. This sampler should be an important tool for studying viral transmission in clinical settings and may significantly contribute towards understanding the modes of influenza virus transmission.

Figure 1

Relative number of H3N2 viral particles collected on the back‐up filters. FluMist® was aerosolized in the calm air chamber, samples were collected on filters of different composition after 40 minutes and analyzed using qPCR.

Figure 2

Regression analysis of influenza particle accumulation over time. Values from three replicate experiments were combined and the average relative number of particles for each stage is presented. (A) H1N1; (B) H3N2 viral particles.

Figure 3

Separation of aerosolized particles by stage. Spores of Aspergillus versicolor (10⁷) and FluMist® were co‐aerosolized together into the calm air chamber. Samples were collected at 40 minutes and analyzed for influenza viruses using qPCR and A. versicolor using qPCR and hemacytometer counts. Data for each stage is presented as the percentage of total number of particles collected in the three stages. Spore counts were the average of eight replicate hemacytometer counts. Values for the FluMist® with no fungal spores were taken from the previous experiment presented in Table 2 and represent the combined average values for H1N1 and H3N2 strains.