In the past, assessment of mask/respirator efficacy by certification entities such as NIOSH has been limited to filter material testing and mannequin head models [22
]. However, the emergence of new pathogens such as severe acute respiratory syndrome (SARS) and most recently the H1N1 virus have called into question the applicability of these findings to real-world practice [23
]. Two recent field studies have attempted to address this question. Loeb et al enrolled 446 nurses in 8 tertiary care centers who were randomly assigned to wear a medical mask or a fit-tested N95 respirator [7
]. No differences between the 2 groups could be detected. Similar results were observed by Ang et al [8
] during the 2009 H1N1 pandemic in Singapore. Groups of persons wearing N95 respirators and surgical masks experienced similar rates of acute respiratory illness. Several problems are inherent to these field studies. Individual exposure risks of health-care workers are not limited to patient contacts in the hospital but are affected by exposures outside the work setting. These risks may be highly variable and difficult to account for in field studies. Furthermore, compliance to other infection control activities such as hand hygiene, isolation/cohorting of patients, or triage may greatly influence individual exposure risks. This was demonstrated during the 2002–2004 SARS outbreaks [24
We have developed a novel approach to test the efficacy of face masks and respirators. In it, human subjects were exposed to live viruses in defined concentrations and particle sizes challenging the barrier precautions by using small, filter- and face-leak–penetrating particles. Safety of the participants is assured by using attenuated viral pathogens in the form of Food and Drug Administration (FDA)–approved vaccine strains, such as LAIV. This novel setup allows a cost-efficient, timely, and accurate way to study infection routes and the efficacy of barrier precautions in controlling airborne and droplet viral transmission in humans.
The most surprising finding of this study is very high rates of transocular transmission in subjects exposed to aerosols containing LAIV. Nasal washes from 3 of 4 participants were positive for LAIV immediately following exposure. This strongly suggests that LAIV is reaching the nasopharynx by way of the nasolacrimal duct. It should also be noted that the lag time between exposure to aerosolized LAIV and detection in nasal washes was less than 30 min, indicating a fast transfer rate of virus to the nasopharynx. Indirect evidence of transocular transmission has previously been reported for the common cold and respiratory syncytial virus (RSV). Winther et al showed that virus can be detected in the nasopharynx after eye inoculation with a rhinovirus suspension (human rhinovirus [HRV] 39; 300 median tissue culture infective dose; first sample taken 24 hour after inoculation) [25
]. Gala et al found a decrease in RSV infections due to disposable eye-nose goggles on an infant ward, indicating an effect of eye protection [26
]. However, this is the first study presenting direct evidence of transocular delivery of influenza viruses in airborne form.
In contrast to our findings using LAIV, we could not detect a common cold virus (HRV 39) in nasal washes in 10 subjects after eye exposure [27
]. We used the same testing environment as in the present study, and the only difference was a much lower exposure dosage for the common cold virus (human infective dose 100 for HRV without protection: 560 PFU total exposure) compared with LAIV (106.5–7.5
FFU total exposure). This raises the question if factors such as specific virus characteristics (enveloped vs nonenveloped) or exposure amounts may influence transocular delivery.
Efficacy testing of the 2 respiratory barrier types against the mechanically generated aerosol revealed the superiority of a fit-tested N95 respirator over a surgical mask; the surgical mask did not provide protection from LAIV transmission whether or not eye protection was also employed. This is in contrast to the recent findings from field studies [7
]. The addition of eye protection to the fit-tested N95 respirator increased protection from LAIV transmission, suggesting a combination of transocular and direct respiratory transmission. However, due to the small sample size, this trend remains to be confirmed.
Human exposure studies using viral pathogens raise concerns regarding the safety of participants. To minimize the risk of adverse events, we decided to use seasonal LAIV approved by the FDA for intranasal spray application. None of the study subjects reported any influenza symptoms or adverse events.
Another risk associated with live virus studies is cross-contamination of samples. Close monitoring, guidance of the participants, and thorough cleaning of the testing environment after the exposure sessions assured compliance with the removal of contaminated clothing and disinfection steps. In addition, our previous study with a common cold virus using the same stringent protocol did not show any evidence of cross-contamination [27
In a recent study, Lindsley et al described the overall distribution of airborne influenza virus in an urgent care medical center [28
]. They found that about 42% of influenza A RNA in particles were ≤4.1 μm, indicating potential transmission by aerosols, as opposed to larger droplets. This finding affirms the importance of studying influenza transmission in aerosols, as we have done in this study. Interestingly, influenza A was detected in concentrations ranging from 0.1 pg RNA/m3
to 75.4 pg RNA/m3
, confirming the presence of viral RNA in a patient care setting. In our study, subjects were exposed to 0.01 pg RNA/m3
, at least 10-fold lower than concentrations of viral RNA found by Lindsley et al. Our decision to expose study subjects to a complete vaccine dosage did not result in influenza virus exposures higher than those likely encountered in clinical settings.
Our study has several limitations. Potential differences between the vaccine strains, wild-type seasonal influenza, and the pandemic H1N1 strain should be considered. To date, only 1 case of person-to-person LAIV transmission following vaccination has been reported, suggesting that this influenza strain may have different transmission characteristics than wild-type influenza strains [29
]. There are similar concerns regarding the contents of bioaerosols. Proteins and other substances expelled with virus particles may change their survival and infectivity properties. However, our study detected influenza present in the nasopharynx immediately following exposure, and we did not measure viral infection or growth in the nasopharynx of study subjects. Thus, the applicability of our findings to influenza virus transmission are not likely influenced by differences in transmission dynamics or growth kinetics between wild-type strains and the cold-adapted, attenuated vaccine strains of this study.
Because we primarily used RT-PCR detection methods in our analysis, we wished to confirm the viability of LAIV vaccine strains used. Therefore, we performed viral cultures in a selection of nasal wash specimens, with the caveat that identification of virus in the samples does not imply infection. We detected LAIV by culture in 2 of 4 samples. Our findings are consistent with those of other groups that showed greatly enhanced influenza detection sensitivity of RT-PCR techniques [30
Our limited sample size did not allow calculation of P
values for the group count outcomes. However, exact 95% confidence intervals were calculated to establish an estimate of certainty for the observed results. We also compared the quantitative RT-PCR results revealing significant differences between no protection, surgical masks plus eye protection, and N95 respirator groups. Wearing the surgical mask with or without eye protection did not reduce virus transmission. We were also limited to 2 brands of masks and respirators. In particular, surgical masks have shown filter efficiencies ranging from 10% to 90% in sodium chloride tests [31
]. We tested only 1 N95 respirator, fit-tested to the individual participant as required by the CDC. The effect of fit-testing on respirator efficacy may be an important variable to measure in future studies incorporating larger sample sizes. During the H1N1 pandemic the efficacy of surgical masks compared with N95 respirators for prevention of aerosol transmission of influenza was widely debated, resulting in differing recommendations [32
]. Recently, the CDC has updated its recommendations for the 2010/2011 influenza season, requiring surgical masks for all patient care activities except aerosol-producing procedures. The latter require fit-tested N95 respirators [34
]. However, evidence regarding the efficacy of masks or respirators against influenza is only beginning to emerge. Our model provides a novel approach to evaluate the preventive qualities of these barrier precautions in a controlled testing environment, allowing the manipulation of key variables such as viral load, particle size, temperature, and humidity during human exposure.
According to our results, the eyes could be an entry route for influenza, allowing viral particles easy and fast access to the upper respiratory tract. The type of surgical mask tested was inferior to a fit-tested N95 respirator in preventing aerosol delivery; however, none of the tested barrier precautions provided complete protection, including a CDC-recommended fit-tested N95 respirator and the addition of eye protection.
The conclusions drawn from this study are influenced by the current lack of understanding regarding the viral particle load necessary to infect an individual with influenza, and the dispersal pattern of influenza produced by affected patients. Mechanically produced aerosol represents a high-risk exposure situation using small aerosol particles, and will need to be compared with concentrations and size distributions that represent human-generated aerosols to allow an understanding of actual risk. However, our study introduces a novel testing approach and points to the potential need for combining effective respirator types with eye protection to successfully interrupt transmission of influenza in aerosol form. These outcomes may provide at least first insights in the transocular transmission dynamics of influenza and the efficacy of the barrier precautions tested.