Journal of Virological Methods 243 (2017) 109–112 Contents lists available at ScienceDirect Journal of Virological Methods j o ur na l ho me pa ge: www.elsevier.com/locate/jviromet Short communication Inactivation of rabies virus Guanghui Wu a,∗ , David Selden a , Anthony R. Fooks a,b , Ashley Banyard a a Wildlife Zoonoses and Vector-Borne Diseases Research Group, Animal and Plant Health Agency (APHA), Woodham Lane, New Haw, Addlestone, Surrey KT15 3NB, United Kingdom b Department of Clinical Infection, Microbiology and Immunology, Institute of Infection and Global Health, University of Liverpool, Liverpool, UK Article history: Received 22 November 2016 Received in revised form 27 January 2017 Accepted 2 February 2017 Available online 5 February 2017 Keywords: Rabies Virus Disinfectant Virkon S Isopropyl alcohol Paraformaldehyde a b s t r a c t Rabies virus is a notifiable pathogen that must be handled in high containment facilities where national and international guidelines apply. For the effective inactivation of rabies virus, a number of reagents were tested. Virkon S (1%) solution caused more than 4 log reduction of rabies virus in culture medium supplemented with 10% foetal calf serum within 1 min. Isopropyl alcohol (70%) treatment resulted in >3 log reduction of rabies virus within 20 s when applied at a ratio of 19:1, making it a suitable agent for surface decontamination whereas 70% ethanol was ineffective. Rabies virus (from 10 2.33 to 10 3 ffu/ml) was also inactivated when cell cultures were fixed with 3% or 4% paraformaldehyde for 30 min. Regardless of inactivation procedure, when taking inactivated virus preparations out of a biological containment envelope, proof of inocuity must be demonstrated to cover any possible error/deviation from procedure. Crown Copyright © 2017 Published by Elsevier B.V. All rights reserved. Rabies is a non-segmented negative strand enveloped RNA virus. It is considered to be one of the most deadly pathogens known with the case fatality rate approaching 100% after the onset of clinical disease (Fooks et al., 2014; Jackson, 2014). For these reasons, the laboratories that work with lyssaviruses must comply with relevant national biocontainment and biosafety regulations (OIE, 2013). Our current local requirements dictate that all work using live virus has to be performed in Microbiological Safety Cabinets (MBSCs) in either in class I or class III mode depending on the nature of pro- cedures undertaken. Solid waste materials (such as plastic ware) are treated with disinfectant, autoclaved and incinerated to ensure the complete destruction of infected materials. Liquid wastes are disinfected and emptied into an effluent treatment plant where the effluent is heated to 96 ◦ C for two hours then cooled prior to release. Alongside removal of solid and liquid wastes from a high contain- ment unit, the removal of items from MBSCs during procedures where live virus is present can also present disinfection challenges. Virkon S (manufactured by Du Pont) is a commonly used dis- infectant for all purposes in the laboratory. It contains potassium peroxymonosulfate (an oxidizing agent), sodium dodecylbenzene- sulfonate, sulfamic acid, and inorganic buffers. In house data showed that rabies virus was inactivated by Virkon within 24 min at 4 ◦ C (unpublished results). However, during standard laboratory ∗ Corresponding author. E-mail address: guanghui.wu@apha.gsi.gov.uk (G. Wu). working procedures, a 24 min contact time prohibits efficient work- ing practices. Alongside this, the use of Virkon S can be problematic as it represents a hazard itself to operators within the laboratory: it can produce acute oral, inhalation and dermal toxicity; serious eye damage. It is toxic to aquatic life following discharge into the environment. Furthermore, Virkon S is corrosive to metal surfaces following prolonged exposure which can reduce the life span of the MBSCs (http://virkon.com/en/products-applications/disinfectants/ ). For these reasons, alternative disinfectants were also assessed for their suitability in the inactivation of rabies virus. Published data indicated that rabies virus can be inactivated by a range of chemicals i.e. soap solutions (1%–20%), 43%–70% alco- hol, quaternary ammonium disinfectants in 1:500 dilution, 5–7% iodine solution (Kaplan et al., 1966), anionic and cationic deter- gents, lemon juice, vinegar, hydrochloric acid, sodium carbonate (Larghi et al., 1975), glutaraldehyde (Minamoto et al., 1988), and hydrogen peroxide (Abd-Elghaffar et al., 2016). One report stated that rabies was completely inactivated within 20 min with ace- tone at a concentration of >30%, within 2 min with 75% ethanol, within 6 h with 0.05% formaldehyde, and within 11 min with 10% formaldehyde (Pan et al., 2011). While rabies virus could be inac- tivated by reagents used for the fluorescent rabies antibody test (FAT) (White and Chappell, 1982), the prescribed conditions were not practical for routine use. A recent publication reiterated the fact that acetone fixation is not suitable for complete inactivation of rabies virions, instead only causing a reversible dehydration of the virus (Jarvis et al., 2016). Considering the available informa- http://dx.doi.org/10.1016/j.jviromet.2017.02.002 0166-0934/Crown Copyright © 2017 Published by Elsevier B.V. All rights reserved.