Comparative characterization of Crimean-Congo hemorrhagic fever virus cell culture systems with application to propagation and titration methods

Virology Journal

Research

Peter Marszal¹  Hongzhao Li¹  Greg Smith¹  Melissa Goolia¹  Bradley S. Pickering² 

[1] National Centre for Foreign Animal Disease, Canadian Food Inspection Agency, Winnipeg, MB, Canada;National Centre for Foreign Animal Disease, Canadian Food Inspection Agency, Winnipeg, MB, Canada;Department of Medical Microbiology and Infectious Diseases, College of Medicine, Faculty of Health Sciences, University of Manitoba, Winnipeg, MB, Canada;Department of Veterinary Microbiology and Preventive Medicine, College of Veterinary Medicine, Iowa State University, Ames, IA, USA;

关键词: Crimean-Congo hemorrhagic fever virus;    Cell line;    Cell culture;    Titration;    SW-13;    Vero E6;    HuH-7;    BSR-T7/5;    Plaque assay;    TCID50;   

DOI  :  10.1186/s12985-023-02089-w

 received in 2022-12-07, accepted in 2023-06-02,  发布年份 2023

来源: Springer

【 摘 要 】

Crimean-Congo hemorrhagic fever orthonairovirus (CCHFV) is a biosafety level 4 and World Health Organization top priority pathogen. Infection leads to an often fatal hemorrhagic fever disease in humans. The tick-borne virus is endemic in countries across Asia, Europe and Africa, with signs of spreading into new regions. Despite the severity of disease and the potential of CCHFV geographic expansion to cause widespread outbreaks, no approved vaccine or treatment is currently available. Critical for basic research and the development of diagnostics or medical countermeasures, CCHFV viral stocks are commonly produced in Vero E6 and SW-13 cell lines. While a variety of in-house methods are being used across different laboratories, there has been no clear, specific consensus on a standard, optimal system for CCHFV growth and titration. In this study, we perform a systematic, side-by-side characterization of Vero E6 and SW-13 cell lines concerning the replication kinetics of CCHFV under different culture conditions. SW-13 cells are typically cultured in a CO2-free condition (SW-13 CO2−) according to the American Type Culture Collection. However, we identify a CO2-compatible culture condition (SW-13 CO2+) that demonstrates the highest viral load (RNA concentration) and titer (infectious virus concentration) in the culture supernatants, in comparison to SW-13 CO2− and Vero E6 cultures. This optimal viral propagation system also leads to the development of two titration methods: an immunostaining-based plaque assay using a commercial CCHFV antibody and a colorimetric readout, and an antibody staining-free, cytopathic effect-based median tissue culture infectious dose assay using a simple excel calculator. These are anticipated to serve as a basis for a reproducible, standardized and user-friendly platform for CCHFV propagation and titration.

【 授权许可】

CC BY   
© Crown 2023

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