SUMMARY To better understand the underlying mechanisms of aerovirology, accurate sampling of airborne viruses is fundamental. The sampling instruments commonly used in aerobiology have also been used to recover viruses suspended in the air. We reviewed over 100 papers to evaluate the methods currently used for viral aerosol sampling. Differentiating infections caused by direct contact from those caused by airborne dissemination can be a very demanding task given the wide variety of sources of viral aerosols. While epidemiological data can help to determine the source of the contamination, direct data obtained from air samples can provide very useful information for risk assessment purposes. Many types of samplers have been used over the years, including liquid impingers, solid impactors, filters, electrostatic precipitators, and many others. The efficiencies of these samplers depend on a variety of environmental and methodological factors that can affect the integrity of the virus structure. The aerodynamic size distribution of the aerosol also has a direct effect on sampler efficiency. Viral aerosols can be studied under controlled laboratory conditions, using biological or nonbiological tracers and surrogate viruses, which are also discussed in this review. Lastly, general recommendations are made regarding future studies on the sampling of airborne viruses.

This study exploits the virulent bacteriophages phi 6 (dsRNA) and MS2 (ssRNA) as surrogates for airborne RNA viruses. Two different filter types, polytetrafluoroethylene (PTFE) and polycarbonate (PC), were tested for their efficiency in collecting aerosolized RNA phages. Two commercial kits were tested for total RNA isolation. Also, heat shock treatments were performed in three different media to obtain the most favorable conditions for reverse transcription assays of dsRNA. Our findings suggest that PC filters are more suitable to recover infectious airborne RNA viruses as determined by plaque assays. Both types of filters were equally efficient in recovering RNA from aerosolized phage phi 6 as established by qRT-PCR. Viral samples should be treated with QIAamp Viral RNA Mini Kit and a 5 min heat shock treatment at 110• C in TE buffer before RT-PCR to maximize detection of phage phi 6. Overall, the infectivity of the recovered phages was severely affected by the aerosolization/air sampling process and the presence of RNA viruses in air samples should be determined by qRT-PCR.

The dairy industry adds starter bacterial cultures to heat-treated milk to control the fermentation process during the manufacture of many cheeses. These highly concentrated bacterial populations are susceptible to virulent phages that are ubiquitous in cheese factories. In this study, the dissemination of these phages by the airborne route and their presence on working surfaces were investigated in a cheese factory. Several surfaces were swabbed, and five air samplers (polytetrafluoroethylene filter, polycarbonate filter, BioSampler, Coriolis cyclone sampler, and NIOSH two-stage cyclone bioaerosol personal sampler) were tested. Samples were then analyzed for the presence of two Lactococcus lactis phage groups (936 and c2), and quantification was done by quantitative PCR (qPCR). Both lactococcal phage groups were found on most swabbed surfaces, while airborne phages were detected at concentrations of at least 10 3 genomes/m 3 of air. The NIOSH sampler had the highest rate of air samples with detectable levels of lactococcal phages. This study demonstrates that virulent phages can circulate through the air and that they are ubiquitous in cheese manufacturing facilities.