Susceptibility of monkeypox virus aerosol suspensions in a rotating chamber

Abstract

Viral aerosols can have a major impact on public health and on the dynamics of infection. Once aerosolized, viruses are subjected to various stress factors and their integrity and potential of infectivity can be altered. Empirical characterization is needed in order to predict more accurately the fate of these bioaerosols both for short term and long term suspension in the air. Here the susceptibility to aerosolization of the monkeypox virus (MPXV), associated with emerging zoonotic diseases, was studied using a 10.7 L rotating chamber. This chamber was built to fit inside a Class three biological safety cabinet, specifically for studying airborne biosafety level three (BSL3) microorganisms. Airborne viruses were detected by culture and quantitative polymerase chain reaction (qPCR) after up to 90 h of aging. Viral concentrations detected dropped by two logs for culture analysis and by one log for qPCR analysis within the first 18 h of aging; viral concentrations were stable between 18 and 90 h, suggesting a potential for the MPXV to retain infectivity in aerosols for more than 90 h. The rotating chamber used in this study maintained viral particles airborne successfully for prolonged periods and could be used to study the susceptibility of other BSL3 microorganisms.

Fig. 1

Rotating chamber. (A) Aluminum cylinder has an interior diameter of 17.8 cm, an outer diameter of 20.3 cm and is 45.7 cm long, total volume is 10.7 L; (B) end caps are composed of three pieces; a 2.5 cm thick aluminum disc (white) mounted on the outer ring of a double sealed ball bearing (dark gray), the extended inner ring of the ball bearing holds a 5.1 cm diameter aluminum rod (light gray) with 3 holes giving access to the interior of the chamber, the chamber is supported by the aluminum rods which stay fixed in position during chamber rotation; (C) Collison three-jet nebulizer; (D) AGI-4 sampler; (E) ball valves controlling access to the interior of the chamber through the end caps; (F) capsule HEPA filters; (G) thermocouple probe with USB connection for recording temperature and relative humidity inside the rotating chamber; (H) electric motor with speed controller providing rotation with a V-belt strapped around the chamber.

Fig. 2

Aerodynamic distribution of airborne particles inside the rotating chamber after nebulization of sterile media with a Collison three-jet nebulizer. Samples (20 s) were taken with the APS immediately after nebulization was stopped (dashed line) and after 18 h of aging in the rotating chamber (solid line). Graph (A) shows the total mass versus the particle size; the transecting lines indicate the mass median aerodynamic diameter (MMAD). Graph (B) shows the data from the same two samples with the total count versus the particle size, the count median aerodynamic diameter (CMAD) is indicated by the transecting lines. The geometric standard deviation is presented in both graphs within brackets.

Fig. 3

Aerosol sample analysis by qPCR (full circles) and by plaque assay (white circles). Decay rate is rapid in the first hours and slows down with time. Error bars are standard deviations. Significant differences (one way ANOVA, Holm–Sidak method; P < 0.05) are indicated by asterisk (*). Differences are significant when compared to the first time point at 18, 48 and 90 h for qPCR and culture analyses.

Fig. 4

Proportion of genomes detected associated with infective viral particles enumerated by plaque assay. Asterisk (*) indicates a significant difference as opposed to the samples taken at 0.03 h (t-test; P < 0.05). There is no significant difference in the proportion of infective viruses taken at time points 0.03 and 0.5 h. There are also no significant differences observed when comparing time points 0.5, 3, 18, 48 and 90 h among each other.

Fig. 5

Aerosol sample analysis by qPCR (A; full circles) and plaque assay (B; white circles). The concentration of the aged aerosol in genomes per liter of air (G_aged), or in pfu/L of air (C_aged), was divided by the concentrations detected after the initial aerosolization (t = 0.03 h; G₀ for qPCR and C₀ for culture) to determine the remaining fractions. Error bars are standard deviations, and each data point is the mean of three repeats; time points 0.5 and 18 h include four repeats. Significant differences (One way ANOVA, Holm–Sidak method; P < 0.05) are indicated by asterisk (*).