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.

Louis Pasteur described the first measurements of airborne microorganisms in 1861. A century later, the inhalation of spores from thermophilic microorganisms was shown to induce attacks of farmers' lung in patients with this disease, while endotoxins originating from Gram-negative bacteria were identified as causal agents for byssinosis in cotton workers. Further epidemiological and toxicological studies have demonstrated inflammatory, respiratory, and pathogenic effects following exposure to bioaerosols. Exposure assessment is often confounded by the diversity of bioaerosol agents in the environment. Microorganisms represent a highly diverse group that may vary in toxicity. Fungi and bacteria are mainly quantified as broad groups using a variety of viable and nonviable assessment methods. Endotoxins and β(1 → 3)-glucans are mainly measured by their activity in the Limulus amebocyte lysate assay, enzymes by immuno-chemical methods and mycotoxins by liquid chromatography-mass spectrometry. Few health-based occupational exposure limits (OELs) are available for risk assessment. For endotoxins, a health-based OEL of 90 endotoxin units m⁻³ has been proposed in the Netherlands. A criteria document for fungal spores recently proposed a lowest observed effect level of 100 000 spores m⁻³ for non-pathogenic and non-mycotoxin producing species based on inflammatory respiratory effects. Recent developments in bioaerosol assessment were presented at the Organic Dust Tromsø Symposium including molecular biological methods for infectious agents and organisms that are difficult to cultivate; studies of submicronic and hyphal fragments from fungi; the effect of biodiversity of microorganisms in asthma studies; and new/improved measurement methods for fungal antigens, enzymes and allergens. Although exposure assessment of bioaerosol agents is complex and limited by the availability of methods and criteria, the field is rapidly evolving.

Bacteriophages are perceived to be good models for the study of airborne viruses because they are safe to use, some of them display structural features similar to those of human and animal viruses, and they are relatively easy to produce in large quantities. Yet, only a few studies have investigated them as models. It has previously been demonstrated that aerosolization, environmental conditions, and sampling conditions affect viral infectivity, but viral infectivity is virus dependent. Thus, several virus models are likely needed to study their general behavior in aerosols. The aim of this study was to compare the effects of aerosolization and sampling on the infectivity of five tail-less bacteriophages and two pathogenic viruses: MS2 (a single-stranded RNA [ssRNA] phage of the Leviviridae family), Φ6 (a segmented double-stranded RNA [dsRNA] phage of the Cystoviridae family), ΦX174 (a single-stranded DNA [ssDNA] phage of the Microviridae family), PM2 (a double-stranded DNA [dsDNA] phage of the Corticoviridae family), PR772 (a dsDNA phage of the Tectiviridae family), human influenza A virus H1N1 (an ssRNA virus of the Orthomyxoviridae family), and the poultry virus Newcastle disease virus (NDV; an ssRNA virus of the Paramyxoviridae family). Three nebulizers and two nebulization salt buffers (with or without organic fluid) were tested, as were two aerosol sampling devices, a liquid cyclone (SKC BioSampler) and a dry cyclone (National Institute for Occupational Safety and Health two-stage cyclone bioaerosol sampler). The presence of viruses in collected air samples was detected by culture and quantitative PCR (qPCR). Our results showed that these selected five phages behave differently when aerosolized and sampled. RNA phage MS2 and ssDNA phage ΦX174 were the most resistant to aerosolization and sampling. The presence of organic fluid in the nebulization buffer protected phages PR772 and Φ6 throughout the aerosolization and sampling with dry cyclones. In this experimental setup, the behavior of the influenza virus resembled that of phages PR772 and Φ6, while the behavior of NDV was closer to that of phages MS2 and ΦX174. These results provide critical information for the selection of appropriate phage models to mimic the behavior of specific human and animal viruses in aerosols.

Summary: The bacterial bioaerosol community of eight swine confinement buildings (SCB) was monitored during two visits in the winter, and one during the summer. To our knowledge, culture‐independent approaches and molecular biology tools such as biomass quantification and biodiversity analyses have never been applied to swine building bioaerosol analyses. Total DNA of each sample was extracted and analysed by quantitative real‐time polymerase chain reaction, denaturing gradient gel electrophoresis (DGGE) and phylogenetic analysis using primers targeting the bacterial 16S rRNA gene. Even though the total bacterial concentration was higher in winter than in summer, the total bacterial concentration for both seasons was 100 to1000 times higher than the total cultural bacteria. The concentration of bioaerosol was influenced by the temperature indoors, which was regulated with an electronic fan system driving warm air and particles outside of the SCB. Comparison of the DGGE profiles showed the same biodiversity in each SCB during both seasons. The phylogenetic analysis revealed a large number of sequences (93.8%) related to Gram‐positive anaerobic bacteria, such as Clostridia , and dominated by the Clostridia cluster I ( C. disporicum ) and the Clostridia cluster XI ( C. glycolycum ). The bioaerosol diversity also contained also a low proportion of Bacteroidetes and Lactobacillales – Streptococcales sequences. Analyses of the global community and phylotype diversity showed that the main source of bioaerosols could come from the pig manure slurry.

The gut microbiota is associated with the modulation of mucosal immunity and the etiology of inflammatory bowel diseases (IBD). Previous studies focused on the impact of bacterial species on IBD but seldom suspected archaea, which can be a major constituent of intestinal microbiota, to be implicated in the diseases. Recent evidence supports that two main archaeal species found in the digestive system of humans, Methanobrevibacter smithii (MBS) and Methanosphaera stadtmanae (MSS) can have differential immunogenic properties in lungs of mice; with MSS but not MBS being a strong inducer of the inflammatory response. We thus aimed at documenting the immunogenic potential of MBS and MSS in humans and to explore their association with IBD. To validate the immunogenicity of MBS and MSS in humans, peripheral blood mononuclear cells from healthy subjects were stimulated with these two microorganisms and the production of inflammatory cytokine TNF was measured by ELISA. To verify MBS and MSS prevalence in IBD, stool samples from 29 healthy control subjects and 29 patients suffering from IBD were collected for DNA extraction. Plasma was also collected from these subjects to measure antigen-specific IgGs by ELISA. Quantitative PCR was used for bacteria, methanogens, MBS and MSS quantification. Mononuclear cells stimulated with MSS produced higher concentrations of TNF (39.5 ng/ml) compared to MBS stimulation (9.1 ng/ml). Bacterial concentrations and frequency of MBS-containing stools were similar in both groups. However, the number of stool samples positive for the inflammatory archaea MSS was higher in patients than in controls (47% vs 20%). Importantly, only IBD patients developed a significant anti-MSS IgG response. The prevalence of MSS is increased in IBD patients and is associated with an antigen-specific IgG response.

This study was designed to test the efficacy of an air treatment using ozone and relative humidity (RH) for the inactivation of airborne viruses. Four phages (φX174, PR772, MS2 and φ6) and one eukaryotic virus (murine norovirus MNV-1) were exposed to low ozone concentrations (1.23 ppm for phages and 0.23 ppm for MNV-1) and various levels of RH for 10 to 70 minutes. The inactivation of these viruses was then assessed to determine which of the tested conditions provided the greatest reduction in virus infectivity. An inactivation of at least two orders of magnitude for φX174, MS2 and MNV-1 was achieved with an ozone exposure of 40 minutes at 85% RH. For PR772 and φ6, exposure to the reference condition at 20% RH for 10 minutes yielded the same results. These findings suggest that ozone used at a low concentration is a powerful disinfectant for airborne viruses when combined with a high RH. Air treatment could therefore be implemented inside hospital rooms ventilated naturally.