Viable monkeypox virus found in nearly all environmental samples from patient's room in first week of infection

In a recent study posted to the medRxiv* pre-print server, researchers investigated the presence of viable monkeypox virus (MPXV) in a room occupied by an infected male patient admitted to the National Centre for Infectious Diseases (NCID), Singapore.

Background

The MPXV continues to spread globally, with over 16,000 MPXV cases and five deaths reported in 75 countries in five World Health Organization (WHO) regions. Subsequently, the WHO has declared the 2022 MPXV outbreak a global public health emergency. Studies have found that contact with wild animals in endemic areas (sub-African tribes in forested areas) and close physical contact with infected individuals increase the risk of contracting MPXV infection.

Epidemiological studies have shown that direct physical contact prolonged throat positivity for MPXV even after the resolution of skin lesions, raising concerns about its aerosol-based transmissions. Yet, there is a lack of data about the modes of MPXV transmission, especially systemic studies investigating human-to-human MPXV transmission.

About the study

In the present study, researchers longitudinally sampled the air, surfaces, water, and dust in an airborne infection isolation room (AIIR) occupied by an MPXV patient who presented with skin lesions and fever. The room was cleaned daily with 10,000 parts-per-million (ppm) bleach and had 12 uni-directional high-efficiency particulate air (HEPA) filter changes per hour. The team performed environmental sampling on infection days 7, 8, 13, and 21. Additionally, they conducted air sampling on day 15 of infection using four NIOSH and two SASS samplers.

The researchers used the SASS and Coriolis samplers for air sampling of the AIIR on days 7 and 8 of the MPXV infection, with samplers set at 0.8 and 0.9 meters on the patient's left and right sides, respectively. On days 13 and 21, the researchers placed an additional set of SASS and Coriolis air samplers at 2.5 meters from the patient. They placed all the air samplers on a trolley at 1.2 meters from the ground. While the SAAS sampler ran for two hours at a flow rate of 300 liters (l)/minute, the Coriolis sampler ran at 100 l/minute for 1.5 hours. The samplers collected particulate matter (PM) samples in sizes PM1, PM 2.5, PM4, and PM 10 in the viral transport medium (VTM).

The researchers collected all surface samples from the AIIR, including room, toilet, and anteroom, using sterile nylon flocked swabs pre-moistened with universal VTM. Further, the team used sterile-vacuum socks to collect dust samples from linen, room, and toilet floor. They sent all the study samples for testing to the biosafety level-3 (BSL-3) laboratory at the Environmental Health Institute in Singapore. Finally, the researchers pre-processed these samples to extract MPXV deoxyribonucleic acid (DNA). They subjected viral DNA to quantitative real-time polymerase chain reaction (PCR) to estimate the viral copy number. They cultured viruses for selected MPXV DNA positive samples and observed cytopathic effect (CPE).

Study findings

The researchers detected MPXV DNA in the patient's nasopharyngeal swab and peri-anal lesions on day 5 of infection when he got admitted to the hospital. The frequency of his skin lesions was the highest on the buttocks, followed by the back and extremities (23 vs. 15 vs. four), but they resolved after eight days of infection. The patient got discharged from the hospital on day 23 of the MPXV infection.

The team collected 179 environmental samples, with 56, 100, 16, and seven air, surface, dust, and water samples. The viral contamination in the air persisted for 21 days, with a peak viral load of 1.25 x 10⁴ copies/swab on day 8 of air sampling. Dust samples had MPXV DNA until day 21, with the highest viral load on day 7 in toilet floor dust samples, equivalent to 5.94 x 10⁷ virus copies/sample. It also declined to the lowest contamination level by day 21 of infection. The team noted that water samples collected from the Sink P-traps were positive for MPXV DNA until day 13.

Conclusions

The researchers recovered viable MPXV in nearly all air samples of the patient's room, though not culturable, and extensive surface contamination of the patient's chair, toilet seat, and dust from bed linen in the first week of infection, with gradual decline later. While the detection of MPXV DNA across sampling days showed continued viral shedding throughout the disease course, the recovery of viable virus from the chair and toilet seat correlated with the location of skin lesions. Likewise, viable MPXV in surface swabs and dust indicated a possibility of fomite-based transmission, especially in home settings.

The environmental contamination reduced from the second week of infection when the patient stopped developing new skin lesions. This finding highlighted the importance of disinfection of the surfaces of chairs, toilets, and floors and taking precautions when handling linens. The researchers found MPXV material only in particles of >4 μm sizes, which nullified the possibility that MPXV was transmitted via breathing or talking in this case. It could be due to 12 HEPA-filtered uni-directional air changes per hour or high ventilation rates. Therefore, future studies should examine direct breath samples in the environment with typical air conditions for a better understanding of the respiratory source of MPXV transmission.

Nevertheless, the presence of live MPXV in dust samples suggested lesion shedding as the potential source of air contamination. Likely, the inoculum dose and host susceptibility for a particular transmission mode affect the onward transmission of all viruses. Therefore, future studies should assess the MPXV transmission dynamics, including the infectious dose required to cause the disease.

Previous studies suggested that pathogens capable of aerosol transmission should be associated with a high reproductive number (R₀). However, pertussis that transmits via droplets has a much higher R₀ than tuberculosis-causal pathogen transmitted via aerosols. Considering that and the exceptional abilities of MPXV to mutate, studies should extensively evaluate all the possible modes of MPXV transmission, especially in hospital settings, to mitigate its spread.

*Important notice

medRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.

Written by

Neha Mathur

Neha is a digital marketing professional based in Gurugram, India. She has a Master’s degree from the University of Rajasthan with a specialization in Biotechnology in 2008. She has experience in pre-clinical research as part of her research project in The Department of Toxicology at the prestigious Central Drug Research Institute (CDRI), Lucknow, India. She also holds a certification in C++ programming.