Explanation for Discordant Findings in Literature
When reviewing the four aforementioned studies (H&H versus S&S), we noticed that the media used to produce airborne droplets containing IAV differed. All the media contained some salts; however, those in H&H contained substantially more proteins than did those in S&S (Table 1). We thus hypothesize that the conflicting results between S&S and H&H were due to the varying protein content of the media used in these studies. Our results with model media confirm this hypothesis: the trends in viability v. RH in media with mainly salts (S&S and this study) resemble the illustration in Figure 4A, while those in media with both salts and proteins (H&H and this study) resemble Figure 4B.
We showed that viral decay is correlated with the concentration of salts in saline droplets, which is controlled by RH through evaporation (Figure 2). Previous studies based on experiments in bulk salt solutions at concentrations up to saturation failed to detect such a correlation [31]. The supersaturated conditions to which the virus is exposed in aerosols cannot be achieved in bulk media, and so the phenomenon observed here can only be demonstrated through methods that enable supersaturation. The combination of highly elevated salt concentrations at medium RH and salt crystallization at the ERH may explain the trends observed in the two media containing mainly salts (i.e., PBS and DMEM) and in S&S. The RH corresponding to the maximal decay rate may differ between media, possibly due to different ERHs for media of different compositions [27]. The minimum viabilities occurred between 40–70% RH, mostly around 50% in Schaffer et al. [14] and 58–60% in Shechmeister. [15] For comparison, we found minimum viabilities at 50% in PBS and 60% in DMEM.
As did H&H, we found that proteins had a “protective” effect for IAV under concentrated conditions at medium RH, although the reasons for it are still unknown. One possibility is that proteins in aerosol droplets could become enriched around viruses due to mutual hydrophobicity and provide some protection against concentrated salts. Although this hypothesis is highly speculative, it is supported by the fact that mucin glycoproteins usually serve as a barricade against potential pathogens including viruses and bacteria by specific or non-specific binding [28], [29].