tetano
Editor, Senior Moderator
Med Microbiol Immunol
. 2021 Jul 1.
doi: 10.1007/s00430-021-00716-3. Online ahead of print.
Evaluation of stability and inactivation methods of SARS-CoV-2 in context of laboratory settings
Marek Widera[SUP] 1 [/SUP], Sandra Westhaus[SUP] 2 [/SUP], Holger F Rabenau[SUP] 2 [/SUP], Sebastian Hoehl[SUP] 2 [/SUP], Denisa Bojkova[SUP] 2 [/SUP], Jindrich Cinatl Jr[SUP] 2 [/SUP], Sandra Ciesek[SUP] 2 3 4 [/SUP]
Affiliations
Abstract
The novel coronavirus SARS-CoV-2 is the causative agent of the acute respiratory disease COVID-19, which has become a global concern due to its rapid spread. Laboratory work with SARS-CoV-2 in a laboratory setting was rated to biosafety level 3 (BSL-3) biocontainment level. However, certain research applications in particular in molecular biology require incomplete denaturation of the proteins, which might cause safety issues handling contaminated samples. In this study, we evaluated lysis buffers that are commonly used in molecular biological laboratories for their ability to inactivate SARS-CoV-2. In addition, viral stability in cell culture media at 4 °C and on display glass and plastic surfaces used in laboratory environment was analyzed. Furthermore, we evaluated chemical and non-chemical inactivation methods including heat inactivation, UV-C light, addition of ethanol, acetone-methanol, and PFA, which might be used as a subsequent inactivation step in the case of insufficient inactivation. We infected susceptible Caco-2 and Vero cells with pre-treated SARS-CoV-2 and determined the tissue culture infection dose 50 (TCID[SUB]50[/SUB]) using crystal violet staining and microscopy. In addition, lysates of infected cells and virus containing supernatant were subjected to RT-qPCR analysis. We have found that guanidine thiocyanate and most of the tested detergent containing lysis buffers were effective in inactivation of SARS-CoV-2, however, the M-PER lysis buffer containing a proprietary detergent failed to inactivate the virus. In conclusion, careful evaluation of the used inactivation methods is required especially for non-denaturing buffers. Additional inactivation steps might be necessary before removal of lysed viral samples from BSL-3.
Keywords: Bio safety; COVID-19; Corona virus; Inactivation; SARS-CoV-2; Stability.
. 2021 Jul 1.
doi: 10.1007/s00430-021-00716-3. Online ahead of print.
Evaluation of stability and inactivation methods of SARS-CoV-2 in context of laboratory settings
Marek Widera[SUP] 1 [/SUP], Sandra Westhaus[SUP] 2 [/SUP], Holger F Rabenau[SUP] 2 [/SUP], Sebastian Hoehl[SUP] 2 [/SUP], Denisa Bojkova[SUP] 2 [/SUP], Jindrich Cinatl Jr[SUP] 2 [/SUP], Sandra Ciesek[SUP] 2 3 4 [/SUP]
Affiliations
- PMID: 34196781
- DOI: 10.1007/s00430-021-00716-3
Abstract
The novel coronavirus SARS-CoV-2 is the causative agent of the acute respiratory disease COVID-19, which has become a global concern due to its rapid spread. Laboratory work with SARS-CoV-2 in a laboratory setting was rated to biosafety level 3 (BSL-3) biocontainment level. However, certain research applications in particular in molecular biology require incomplete denaturation of the proteins, which might cause safety issues handling contaminated samples. In this study, we evaluated lysis buffers that are commonly used in molecular biological laboratories for their ability to inactivate SARS-CoV-2. In addition, viral stability in cell culture media at 4 °C and on display glass and plastic surfaces used in laboratory environment was analyzed. Furthermore, we evaluated chemical and non-chemical inactivation methods including heat inactivation, UV-C light, addition of ethanol, acetone-methanol, and PFA, which might be used as a subsequent inactivation step in the case of insufficient inactivation. We infected susceptible Caco-2 and Vero cells with pre-treated SARS-CoV-2 and determined the tissue culture infection dose 50 (TCID[SUB]50[/SUB]) using crystal violet staining and microscopy. In addition, lysates of infected cells and virus containing supernatant were subjected to RT-qPCR analysis. We have found that guanidine thiocyanate and most of the tested detergent containing lysis buffers were effective in inactivation of SARS-CoV-2, however, the M-PER lysis buffer containing a proprietary detergent failed to inactivate the virus. In conclusion, careful evaluation of the used inactivation methods is required especially for non-denaturing buffers. Additional inactivation steps might be necessary before removal of lysed viral samples from BSL-3.
Keywords: Bio safety; COVID-19; Corona virus; Inactivation; SARS-CoV-2; Stability.