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 examined the presence of viable monkeypox virus (MPXV) in a room occupied by an infected male patient admitted to the National Center for Infectious Diseases (NCID), Singapore.

Study: Viable Monkeypox virus in a patient room environment. Image credit: Marina Demidiuk/Shutterstock


The MPXV continues to spread worldwide, with more than 16,000 MPXV cases and five deaths reported in 75 countries in five regions of the World Health Organization (WHO). Subsequently, the WHO declared the MPXV outbreak a global public health emergency in 2022. Studies have shown that contact with wildlife in endemic areas (sub-African tribes in forested areas) and close physical contact with infected individuals increase the risk of 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 aerosol-based transmission. Still, there is a lack of data on the modes of MPXV transmission, especially systemic studies investigating human-to-human MPXV transmission.

About the study

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

The researchers used the SASS and Coriolis air sampling samplers from the AIIR on days 7 and 8 of MPXV infection, with samplers located at 0.8 and 0.9 meters on the left and right sides of the patient, respectively. On days 13 and 21, the researchers placed an additional set of SASS and Coriolis air samplers 2.5 meters from the patient. They placed all the air samplers on a cart 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 for 1.5 hours at 100 L/minute. The samplers collected particulate matter (PM) samples in the 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 chamber, toilet and antechamber, using sterile nylon flocked swabs pre-moistened with universal VTM. Furthermore, the team used sterile vacuum socks to collect dust samples from linen, room and toilet floor. They sent all study samples to the Environmental Health Institute’s Biosafety Level-3 (BSL-3) lab in Singapore for testing. Finally, the researchers preprocessed these samples to extract MPXV deoxyribonucleic acid (DNA). They subjected viral DNA to a quantitative real-time polymerase chain reaction (PCR) to estimate the number of viral copies. They cultured viruses for selected MPXV DNA positive samples and observed a cytopathic effect (CPE).

Study findings

The researchers discovered MPXV DNA in the nasopharyngeal swab and the perianal lesions on day 5 of the infection when he was hospitalized. The frequency of his skin lesions was 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 was discharged from the hospital on Day 23 of MPXV infection.

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


The researchers found viable MPXV in nearly all air samples from the patient’s room, although not culturable, and extensive surface contamination of the patient’s chair, toilet seat, and dust from the patient’s bedding in the first week of infection, with a gradual decline later. While the detection of MPXV DNA over the sampling days showed that the virus persisted throughout the disease course, the recovery of viable virus from the chair and toilet seat correlated with the location of skin lesions. Similarly, viable MPXV in surface swabs and dust indicated a possibility of fomite-based transmission, especially in home environments.

The environmental contamination decreased from the second week of infection when the patient stopped developing new skin lesions. This finding highlighted the importance of disinfecting the surfaces of seats, toilets and floors and taking precautions when handling linens. The researchers found MPXV material only in particles >4 m, negating the possibility that MPXV was transmitted through breathing or talking in this case. This could be due to 12 HEPA filtered unidirectional 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 possible source of air pollution. It is likely that the inoculum dose and the host’s susceptibility to a particular transmission mode influence the further transmission of all viruses. Therefore, future studies should assess MPXV transmission dynamics, including the infectious dose required to induce the disease.

Previous studies suggested that pathogens capable of aerosol transfer should be associated with a high reproductive number (R0). Whooping cough transmitted via droplets, however, has a much higher R0 than tuberculosis-causing agents that are transmitted via aerosols. Given that and the exceptional ability of MPXV to mutate, studies should comprehensively evaluate all possible modes of MPXV transmission, especially in hospital settings, to reduce its spread.

*Important announcement

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

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