Inactivation of Venezuelan Equine Encephalitis Virus Genome Using Two Methods

• Published in: Viruses Vol. 14; no. 2; p. 272
• Main Authors: Behnia, Mahgol, Baer, Alan, Skidmore, Andrew M., Lehman, Caitlin W., Bracci, Nicole, Kehn-Hall, Kylene, Bradfute, Steven B.
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 28.01.2022; MDPI
• Subjects: Animals; cDNA synthesis; Cells, Cultured; Chlorocebus aethiops; Communication; Cytopathogenic Effect, Viral; Disease; DNA, Complementary - biosynthesis; Encephalitis; Encephalitis Virus, Eastern Equine - genetics; Encephalitis Virus, Eastern Equine - physiology; Encephalitis Virus, Venezuelan Equine - genetics; Encephalitis Virus, Venezuelan Equine - physiology; Encephalitis Virus, Western Equine - genetics; Encephalitis Virus, Western Equine - physiology; Genome, Viral; Genomes; Genomics; Infections; Laboratories; Mortality; Ribonucleases - metabolism; RNA; RNA fragmentation; RNA viruses; RNA, Viral - chemistry; RNA, Viral - physiology; Strains (organisms); Transcription; Transfection; Trinidad and Tobago; Vaccines; Venezuelan equine encephalitis; Venezuelan equine encephalitis virus; Vero Cells; viral genome; viral genome inactivation; Viral infections; Virions; Virus Inactivation; Viruses; Western equine encephalitis; United States > US; Trinidad and Tobago; viral genome inactivation; encephalitis; Venezuelan equine encephalitis virus; RNA fragmentation; cDNA synthesis
• DOI: 10.3390/v14020272
• ISSN: 1999-4915

Venezuelan equine encephalitis virus (VEEV) is an Alphavirus in the Togaviridae family of positive-strand RNA viruses. The viral genome of positive-strand RNA viruses is infectious, as it produces infectious virus upon introduction into a cell. VEEV is a select agent and samples containing viral RNA are subject to additional regulations due to their infectious nature. Therefore, RNA isolated from cells infected with BSL-3 select agent strains of VEEV or other positive-strand viruses must be inactivated before removal from high-containment laboratories. In this study, we tested the inactivation of the viral genome after RNA fragmentation or cDNA synthesis, using the Trinidad Donkey and TC-83 strains of VEEV. We successfully inactivated VEEV genomic RNA utilizing these two protocols. Our cDNA synthesis method also inactivated the genomic RNA of eastern and western equine encephalitis viruses (EEEV and WEEV). We also tested whether the purified VEEV genomic RNA can produce infectious virions in the absence of transfection. Our result showed the inability of the viral genome to cause infection without being transfected into the cells. Overall, this work introduces RNA fragmentation and cDNA synthesis as reliable methods for the inactivation of samples containing the genomes of positive-strand RNA viruses.