Aerosol Particle Size Influences the Infectious Dose and Disease Severity in a Golden Syrian Hamster Model of Inhalational COVID-19.

Background: Significant evidence suggests that SARS-CoV-2 can be transmitted via respiratory aerosols, which are known to vary as a function of respiratory activity. Most animal models examine disease presentation following inhalation of small-particle aerosols similar to those generated during quiet breathing or speaking. However, despite evidence that particle size can influence dose-infectivity relationships and disease presentation for other microorganisms, no studies have examined the infectivity of SARS-CoV-2 contained in larger particle aerosols similar to those produced during coughing, singing, or talking. Therefore, the aim of the present study was to assess the influence of aerodynamic diameter on the infectivity and virulence of aerosols containing SARS-CoV-2 in a hamster model of inhalational COVID-19. Methods: Dose-response relationships were assessed for two different aerosol particle size distributions, with mass median aerodynamic diameters (MMADs) of 1.3 and 5.2 µm in groups of Syrian hamsters exposed to aerosols containing SARS-CoV-2. Results: Disease was characterized by viral shedding in oropharyngeal swabs, increased respiratory rate, decreased activity, and decreased weight gain. Aerosol particle size significantly influenced the median doses to induce seroconversion and viral shedding, with both increasing ∼30-fold when the MMAD was increased. In addition, disease presentation was dose-dependent, with seroconversion and viral shedding occurring at lower doses than symptomatic disease characterized by increased respiratory rate and decreased activity. Conclusions: These results suggest that aerosol particle size may be an important factor influencing the risk of COVID-19 transmission and needs to be considered when developing animal models of disease. This result agrees with numerous previous studies with other microorganisms and animal species, suggesting that it would be generally translatable across different species. However, it should be noted that the absolute magnitude of the observed shifts in the median doses obtained with the specific particle sizes utilized herein may not be directly applicable to other species.

Comparison of Dose-Response Relationships for Two Isolates of SARS-CoV-2 in a Nonhuman Primate Model of Inhalational COVID-19.

Background: As the COVID-19 pandemic has progressed, numerous variants of SARS-CoV-2 have arisen, with several displaying increased transmissibility. Methods: The present study compared dose-response relationships and disease presentation in nonhuman primates infected with aerosols containing an isolate of the Gamma variant of SARS-CoV-2 to the results of our previous study with the earlier WA-1 isolate of SARS-CoV-2. Results: Disease in Gamma-infected animals was mild, characterized by dose-dependent fever and oronasal shedding of virus. Differences were observed in shedding in the upper respiratory tract between Gamma- and WA-1-infected animals that have the potential to influence disease transmission. Specifically, the estimated median doses for shedding of viral RNA or infectious virus in nasal swabs were approximately 10-fold lower for the Gamma variant than the WA-1 isolate. Given that the median doses for fever were similar, this suggests that there is a greater difference between the median doses for viral shedding and fever for Gamma than for WA-1 and potentially an increased range of doses for Gamma over which asymptomatic shedding and disease transmission are possible. Conclusions: These results complement those of previous studies, which suggested that differences in exposure dose may help to explain the range of clinical disease presentations observed in individuals with COVID-19, highlighting the importance of public health measures designed to limit exposure dose, such as masking and social distancing. The dose-response data provided by this study are important to inform disease transmission and hazard modeling, as well as to inform dose selection in future studies examining the efficacy of therapeutics and vaccines in animal models of inhalational COVID-19.

SARS-CoV-2 inactivation by ultraviolet radiation and visible light is dependent on wavelength and sample matrix.

Numerous studies have demonstrated that SARS-CoV-2 can be inactivated by ultraviolet (UV) radiation. However, there are few data available on the relative efficacy of different wavelengths of UV radiation and visible light, which complicates assessments of UV decontamination interventions. The present study evaluated the effects of monochromatic radiation at 16 wavelengths from 222 nm through 488 nm on SARS-CoV-2 in liquid aliquots and dried droplets of water and simulated saliva. The data were used to generate a set of action spectra which quantify the susceptibility of SARS-CoV-2 to genome damage and inactivation across the tested wavelengths. UVC wavelengths (≤280 nm) were most effective for inactivating SARS-CoV-2, although inactivation rates were dependent on sample type. Results from this study suggest that UV radiation can effectively inactivate SARS-CoV-2 in liquids and dried droplets, and provide a foundation for understanding the factors which affect the efficacy of different wavelengths in real-world settings.

Seroconversion and fever are dose-dependent in a nonhuman primate model of inhalational COVID-19.

While evidence exists supporting the potential for aerosol transmission of SARS-CoV-2, the infectious dose by inhalation remains unknown. In the present study, the probability of infection following inhalation of SARS-CoV-2 was dose-dependent in a nonhuman primate model of inhalational COVID-19. The median infectious dose, assessed by seroconversion, was 52 TCID50 (95% CI: 23-363 TCID50), and was significantly lower than the median dose for fever (256 TCID50, 95% CI: 102-603 TCID50), resulting in a group of animals that developed an immune response post-exposure but did not develop fever or other clinical signs of infection. In a subset of these animals, virus was detected in nasopharyngeal and/or oropharyngeal swabs, suggesting that infected animals without signs of disease are able to shed virus and may be infectious, which is consistent with reports of asymptomatic spread in human cases of COVID-19. These results suggest that differences in exposure dose may be a factor influencing disease presentation in humans, and reinforce the importance of public health measures that limit exposure dose, such as social distancing, masking, and increased ventilation. The dose-response data provided by this study are important to inform disease transmission and hazard modeling, and, ultimately, mitigation strategies. Additionally, these data will be useful to inform dose selection in future studies examining the efficacy of therapeutics and vaccines against inhalational COVID-19, and as a baseline in healthy, young adult animals for assessment of the importance of other factors, such as age, comorbidities, and viral variant, on the infectious dose and disease presentation.

The Stability of an Isolate of the SARS-CoV-2 B.1.1.7 Lineage in Aerosols Is Similar to 3 Earlier Isolates.

BACKGROUND: Our laboratory previously examined the influence of environmental conditions on the stability of an early isolate of SARS-CoV-2 (hCoV-19/USA/WA-1/2020) in aerosols generated from culture medium or simulated saliva. However, genetic differences have emerged among SARS-CoV-2 lineages, and it is possible that these differences may affect environmental stability and the potential for aerosol transmission. METHODS: The influence of temperature, relative humidity, and simulated sunlight on the decay of 4 SARS-CoV-2 isolates in aerosols, including 1 belonging to the recently emerged B.1.1.7 lineage, were compared in a rotating drum chamber. Aerosols were generated from simulated respiratory tract lining fluid to represent aerosols originating from the deep lung. RESULTS: No differences in the stability of the isolates were observed in the absence of simulated sunlight at either 20°C or 40°C. However, a small but statistically significant difference in the stability was observed between some isolates in simulated sunlight at 20°C and 20% relative humidity. CONCLUSIONS: The stability of SARS-CoV-2 in aerosols does not vary greatly among currently circulating lineages, including B.1.1.7, suggesting that the increased transmissibility associated with recent SARS-CoV-2 lineages is not due to enhanced survival in the environment.

The use of an Ebola virus reporter cell line in a semi-automated microtitration assay.

A variety of methods have been developed for quantification of infectious Ebola virus in clinical or laboratory samples, but existing methods often require extensive operator involvement, manual assay scoring, or the use of custom reagents. In this study, we utilize a recently developed Ebola-specific reporter cell line that expresses ZsGreen in response to Ebola virus infection, in conjunction with semi-automated processing and quantification techniques, to develop an unbiased, high-throughput microtitration assay for quantification of infectious Ebola virus in vitro. This assay was found to have equivalent sensitivity to a standardized plaque assay for quantifying viral titers. However, the new assay could be implemented with fewer reagents and processing steps, reduced subjectivity, and higher throughput. This assay may be useful for a variety of applications, particularly studies that require the detection or quantification of infectious Ebola virus in large numbers of samples.

SARS-CoV-2 is rapidly inactivated at high temperature.

In the absence of a vaccine, preventing the spread of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the primary means to reduce the impact of the 2019 coronavirus disease (COVID-19). Multiple studies have reported the presence of SARS-CoV-2 genetic material on surfaces suggesting that fomite transmission of SARS-CoV-2 is feasible. High temperature inactivation of virus has been previously suggested, but not shown. In the present study, we investigated the environmental stability of SARS-CoV-2 in a clinically relevant matrix dried onto stainless steel at a high temperature. The results show that at 54.5 °C, the virus half-life was 10.8 ± 3.0 min and the time for a 90% decrease in infectivity was 35.4 ± 9.0 min. These findings suggest that in instances where the environment can reach temperatures of at least 54.5 °C, such as in vehicle interior cabins when parked in warmer ambient air, that the potential for exposure to infectious virus on surfaces could be decreased substantially in under an hour.

Comparison of the performance of aerosol sampling devices for measuring infectious SARS-CoV-2 aerosols.

To assess the risk of aerosol transmission of SARS-CoV-2, measurements of the airborne viral concentrations in proximity to infected individuals, the persistence of the virus in aerosols, and the dose of the virus needed to cause infection following inhalation are required. For studies aimed at quantifying these parameters, an aerosol sampling device needs to be employed. A number of recent studies have reported the detection of both genetic material and infectious SARS-CoV-2 virus in air samples collected in clinical settings. Previous studies have demonstrated that the efficiency of different samplers for collection and preservation of the infectivity of microorganisms can vary as a function of the specific microorganism. In the present study, the performance of eight common low-flow aerosol sampling devices were compared for their ability to collect and preserve the infectivity of airborne SARS-CoV-2 contained in small particle aerosols. The influence of sampling duration on recovery of infectious virus was also evaluated. Similar concentrations of infectious SARS-CoV-2 were measured in aerosols for the majority of the samplers tested, with the exception of the midget impingers, which measured significantly lower concentrations of SARS-CoV-2. Additionally, in three of the four impingers tested, additional clean airflow through the device following collection of infectious virus resulted in a decrease of the infectious concentration of virus over time, suggesting that virus was being inactivated and these devices may not be suitable for sampling for long durations. Further, RNA copies in the samples over time did not correspond with the losses of infectious SARS-CoV-2 observed in the impingers samples. These data can be utilized to inform interpretation of current studies on the SARS-CoV-2 viral loads in air samples, as well as inform sampling device selection in future studies.

The influence of temperature, humidity, and simulated sunlight on the infectivity of SARS-CoV-2 in aerosols.

Recent evidence suggests that respiratory aerosols may play a role in the spread of SARS-CoV-2 during the ongoing COVID-19 pandemic. Our laboratory has previously demonstrated that simulated sunlight inactivated SARS-CoV-2 in aerosols and on surfaces. In the present study, we extend these findings to include the persistence of SARS-CoV-2 in aerosols across a range of temperature, humidity, and simulated sunlight levels using an environmentally controlled rotating drum aerosol chamber. The results demonstrate that temperature, simulated sunlight, and humidity are all significant factors influencing the persistence of infectious SARS-CoV-2 in aerosols, but that simulated sunlight and temperature have a greater influence on decay than humidity across the range of conditions tested. The time needed for a 90% decrease in infectious virus ranged from 4.8 min at 40 °C, 20% relative humidity, and high intensity simulated sunlight representative of noon on a clear day on the summer solstice at 4°N latitude, to greater than two hours under conditions representative of those expected indoors or at night. These results suggest that the persistence of infectious SARS-CoV-2 in naturally occurring aerosols may be affected by environmental conditions, and that aerosolized virus could remain infectious for extended periods of time under some environmental conditions. The present study provides a comprehensive dataset on the influence of environmental parameters on the survival of SARS-CoV-2 in aerosols that can be utilized, along with data on viral shedding from infected individuals and the inhalational infectious dose, to inform future modeling and risk assessment efforts.

Increasing Temperature and Relative Humidity Accelerates Inactivation of SARS-CoV-2 on Surfaces.

Coronavirus disease 2019 (COVID-19) was first identified in China in late 2019 and is caused by newly identified severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Previous studies had reported the stability of SARS-CoV-2 in cell culture media and deposited onto surfaces under a limited set of environmental conditions. Here, we broadly investigated the effects of relative humidity, temperature, and droplet size on the stability of SARS-CoV-2 in a simulated clinically relevant matrix dried on nonporous surfaces. The results show that SARS-CoV-2 decayed more rapidly when either humidity or temperature was increased but that droplet volume (1 to 50 µl) and surface type (stainless steel, plastic, or nitrile glove) did not significantly impact decay rate. At room temperature (24°C), virus half-life ranged from 6.3 to 18.6 h depending on the relative humidity but was reduced to 1.0 to 8.9 h when the temperature was increased to 35°C. These findings suggest that a potential for fomite transmission may persist for hours to days in indoor environments and have implications for assessment of the risk posed by surface contamination in indoor environments.IMPORTANCE Mitigating the transmission of SARS-CoV-2 in clinical settings and public spaces is critically important to reduce the number of COVID-19 cases while effective vaccines and therapeutics are under development. SARS-CoV-2 transmission is thought to primarily occur through direct person-to-person transfer of infectious respiratory droplets or through aerosol-generating medical procedures. However, contact with contaminated surfaces may also play a significant role. In this context, understanding the factors contributing to SARS-CoV-2 persistence on surfaces will enable a more accurate estimation of the risk of contact transmission and inform mitigation strategies. To this end, we have developed a simple mathematical model that can be used to estimate virus decay on nonporous surfaces under a range of conditions and which may be utilized operationally to identify indoor environments in which the virus is most persistent.

Airborne SARS-CoV-2 Is Rapidly Inactivated by Simulated Sunlight.

Aerosols represent a potential transmission route of COVID-19. This study examined effect of simulated sunlight, relative humidity, and suspension matrix on stability of SARS-CoV-2 in aerosols. Simulated sunlight and matrix significantly affected decay rate of the virus. Relative humidity alone did not affect the decay rate; however, minor interactions between relative humidity and other factors were observed. Mean decay rates (± SD) in simulated saliva, under simulated sunlight levels representative of late winter/early fall and summer were 0.121 ±â€…0.017 min-1 (90% loss, 19 minutes) and 0.306 ±â€…0.097 min-1 (90% loss, 8 minutes), respectively. Mean decay rate without simulated sunlight across all relative humidity levels was 0.008 ±â€…0.011 min-1 (90% loss, 286 minutes). These results suggest that the potential for aerosol transmission of SARS-CoV-2 may be dependent on environmental conditions, particularly sunlight. These data may be useful to inform mitigation strategies to minimize the potential for aerosol transmission.

Simulated Sunlight Rapidly Inactivates SARS-CoV-2 on Surfaces.

Previous studies have demonstrated that SARS-CoV-2 is stable on surfaces for extended periods under indoor conditions. In the present study, simulated sunlight rapidly inactivated SARS-CoV-2 suspended in either simulated saliva or culture media and dried on stainless steel coupons. Ninety percent of infectious virus was inactivated every 6.8 minutes in simulated saliva and every 14.3 minutes in culture media when exposed to simulated sunlight representative of the summer solstice at 40°N latitude at sea level on a clear day. Significant inactivation also occurred, albeit at a slower rate, under lower simulated sunlight levels. The present study provides the first evidence that sunlight may rapidly inactivate SARS-CoV-2 on surfaces, suggesting that persistence, and subsequently exposure risk, may vary significantly between indoor and outdoor environments. Additionally, these data indicate that natural sunlight may be effective as a disinfectant for contaminated nonporous materials.

The Influence of Simulated Sunlight on the Inactivation of Influenza Virus in Aerosols.

BACKGROUND: Environmental parameters, including sunlight levels, are known to affect the survival of many microorganisms in aerosols. However, the impact of sunlight on the survival of influenza virus in aerosols has not been previously quantified. METHODS: The present study examined the influence of simulated sunlight on the survival of influenza virus in aerosols at both 20% and 70% relative humidity using an environmentally controlled rotating drum aerosol chamber. RESULTS: Measured decay rates were dependent on the level of simulated sunlight, but they were not significantly different between the 2 relative humidity levels tested. In darkness, the average decay constant was 0.02 ± 0.06 min-1, equivalent to a half-life of 31.6 minutes. However, at full intensity simulated sunlight, the mean decay constant was 0.29 ± 0.09 min-1, equivalent to a half-life of approximately 2.4 minutes. CONCLUSIONS: These results are consistent with epidemiological findings that sunlight levels are inversely correlated with influenza transmission, and they can be used to better understand the potential for the virus to spread under varied environmental conditions.

Two-Center Evaluation of Disinfectant Efficacy against Ebola Virus in Clinical and Laboratory Matrices.

Ebola virus (EBOV) in body fluids poses risk for virus transmission. However, there are limited experimental data for such matrices on the disinfectant efficacy against EBOV. We evaluated the effectiveness of disinfectants against EBOV in blood on surfaces. Only 5% peracetic acid consistently reduced EBOV titers in dried blood to the assay limit of quantification.

Interferon-ß and Interferon-γ Are Weak Inhibitors of Ebola Virus in Cell-Based Assays.

Previous studies have demonstrated little efficacy of interferons (IFNs) in animal models of Ebola virus disease. However, these studies were limited to a small number of type I IFNs and, during the most recent outbreak of Ebola virus, questions regarding the suitability of the animal models to evaluate IFNs were raised. To address the potential that anti-Ebola virus activity was overlooked, type I and type II IFNs (α-2a, α-2b, -ß, -γ, and -universal) were tested in a variety of cell types (Vero E6, Huh 7 cells, and human macrophages). IFNs are weak inhibitors of Ebola virus Makona in these cell lines.