In a recent study published in Emerging Infectious Diseases, researchers demonstrated the efficacy of two World Health Organization (WHO)‒recommended alcohol-based hand rub solutions to inactivate the monkeypox virus (MPXV).
Background
Implementing hygienic hand measures to preclude healthcare- and outbreak-triggering lethal viral infections is important. Thus, in 2009, the WHO proposed and implemented two alcohol-based formulations, I and II, for surgical and hygienic hand disinfection in healthcare settings. However, researchers have not assessed their inactivation efficacies against MPVX.
A concerning fact about MPXV that makes it a public health concern is that it is spreading among persons who have not traveled to disease-endemic areas. Its clinical and epidemiologic patterns are novel, unlike prior outbreaks. Also, it is remarkably more stable than other pox viruses. It is thus necessary to confirm which disinfectants and biocidal agents can effectively inactivate MPXV.
The study
In the present study, researchers obtained virus isolate MPXV-DUS_001 from a patient in Düsseldorf, Germany, who was infected early during the 2022 MPXV outbreak. They passaged this isolate twice on Vero 76 cells before experimental use.
First, the team cultured Vero 76 cells in Dulbecco modified Eagle (DME) medium and seeded them at a concentration of 0.33 × 106 cells/mL for MPXV preparation. Next, following a change of medium, they inoculated these cells with MPXV at a multiplicity of infection (MOI) of 0.01. The team incubated Vero 76 cells at 37°C until they observed a visible cytopathic effect (CPE). They moved on to harvesting MPXV-infected cells by scraping, then extensive vortexing and subsequent extraction of the infectious supernatant from cell debris by centrifugation. Finally, the researchers aliquoted and titrated virus suspensions per standard protocols and stored them at −80°C for future use. The researchers confirmed that MPXV-DUS_001 was MPXV clade II using panorthopoxvirus‒specific quantitative, real-time reverse transcription polymerase chain reaction (qRT-PCR).
The team adhered to the European guideline EN14476 to assess the virucidal activity of WHO formulations I and II. Since these formulations are 80% ethanol and 75% isopropanol-based, they also evaluated the MPXV susceptibility against ethanol and 2-propanol. The protocol mandated mixing eight parts of disinfectant with one part of bovine serum albumin and MPXV each to attain a final concentration of 0.3 g/L. The suspension was then vortexed and incubated for 30 seconds at room temperature.
They performed an endpoint dilution assay on Vero 76 cells and evaluated CPE microscopically after seven days. Next, they computed the 50% tissue culture infectious dose (ID50) per milliliter for all four disinfectants at final concentrations of 20%, 30%, 40%, 60%. and 80%. Finally, the researchers used a suspension test to examine the virucidal activity of the WHO formulations I and II against MPXV.
Study results
MPXV showed sensitivity to both WHO formulations but was also the most stable among all viruses tested, including modified vaccinia Ankara. Both formulations efficiently inactivated MPXV with reduction factors (RFs)>6.7 at concentrations of 60% and 80% volume/volume (v/v). Remarkably, the WHO formulation II remained effective against MPXV even at a dilution of 40% (vol/vol) with an RF of 6.6. Conversely, WHO formulation I at a similar concentration could not reduce MPXV titers.
The researchers noted that both ethanol and 2-propanol reduced MPXV titers to background levels with RFs >6.7 at >60% and >40% (v/v) concentrations, respectively. With RF=6.6, 40% ethanol (v/v) nearly inactivated MPXV completely. However, with RF=5.3, 30% v/v 2-propanol accomplished the same virucidal activity.
Conclusions
Overall, the study results showed that MPXV showed the highest stability to both WHO formulations compared with other (re)emerging orthopox family viruses. Infectious MPXV persists in a household environment for more than 15 days. Because of tight binding with fibrin matrixes of scab material, virions shed from lesions are even more resistant to desiccation than other enveloped viruses (e.g., influenza virus). The high stability of MPXV necessitates a comprehensive reevaluation of current hygiene measures. Thankfully, both tested WHO formulations effectively inactivated MPXV, supporting their use in healthcare systems and during MPXV outbreaks. To conclude, the study showed that the timely application of alcohol-based disinfectants could effectively minimize MPXV spread during the ongoing MPVX outbreak.
