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Art of the Kill: Designing and Testing Viral Inactivation Procedures for Highly Pathogenic Negative Sense RNA Viruses
Judith Olejnik1,2, Adam J Hume1,2, Stephen J Ross1,2,3
1Department of Virology, Immunology and Microbiology, Chobanian & Avedisian School of Medicine, Boston University, Boston, MA 02118, USA.
Abstract:
The study of highly pathogenic viruses handled under BSL-4 conditions and classified as Select Agents frequently involves the transfer of inactivated materials to lower containment levels for downstream analyses. Adhering to Select Agent and BSL-4 safety regulations requires validation or verification of the inactivation procedures, which comes with an array of challenges for each method. This includes the use of cytotoxic reagents for chemical inactivation and defining the precise inactivation parameters for physical inactivation. Here, we provide a workflow for various inactivation methods using Ebola, Nipah, and Lassa viruses as our examples. We choose three distinct inactivation methods (TRIzol/TRIzol LS, aldehyde fixation using different fixatives, and heat) to highlight the challenges of each method and provide possible solutions. We show that, whereas published chemical inactivation methods are highly reliable, the parameters for heat inactivation must be clearly defined to ensure complete inactivation. In addition to the inactivation data, we also provide examples and templates for the documentation required for approval and use of inactivation SOPs, including an inactivation report, the procedure sections of developed SOPs, and an electronic inactivation certificate that accompanies inactivated samples. The provided information can be used as a roadmap for similar studies at high and maximum containment laboratories.
Insights
This study validates inactivation methods for highly pathogenic viruses like Ebola, Nipah, and Lassa, crucial for safe sample transfer from BSL-4 labs. It emphasizes clear parameter definition for heat inactivation and provides documentation templates for regulatory compliance.
Area of Science:
- Virology
- Biosafety
- Laboratory Science
Background:
- Handling highly pathogenic viruses under Biosafety Level 4 (BSL-4) conditions requires validated inactivation methods for safe transfer of materials for downstream analysis.
- Select Agent regulations and BSL-4 safety protocols mandate rigorous validation of inactivation procedures, presenting challenges with cytotoxic reagents and precise parameter definition.
Purpose of the Study:
- To provide a workflow and practical solutions for validating virus inactivation methods for highly pathogenic viruses.
- To address challenges associated with chemical and physical inactivation methods for viruses like Ebola, Nipah, and Lassa.
- To offer templates for documentation required for inactivation Standard Operating Procedures (SOPs) and sample tracking.
Main Methods:
- Evaluation of three distinct inactivation methods: chemical inactivation (TRIzol/TRIzol LS), aldehyde fixation, and heat inactivation.
- Application of methods to model viruses including Ebola, Nipah, and Lassa viruses.
- Development and presentation of documentation templates for inactivation SOPs and reports.
Main Results:
- Published chemical inactivation methods demonstrated high reliability for virus inactivation.
- Heat inactivation requires clearly defined parameters to ensure complete inactivation of viruses.
- Successful demonstration of a workflow for inactivation validation and documentation.
Conclusions:
- Validated inactivation protocols are essential for the safe transfer of viral materials from high-containment laboratories.
- Standardized documentation, including inactivation reports and certificates, facilitates regulatory compliance and sample traceability.
- The provided workflow serves as a guide for laboratories working with high-containment viruses and inactivation procedures.
