UVC inactivation of pathogenic samples suitable for cryo-EM analysis

Jamie S Depelteau; Ludovic Renault; Nynke Althof; C Keith Cassidy; Luiza M Mendonça; Grant J Jensen; Guenter P Resch; Ariane Briegel

doi:10.1038/s42003-021-02962-w

UVC inactivation of pathogenic samples suitable for cryo-EM analysis

Jamie S Depelteau, Ludovic Renault, Nynke Althof, C Keith Cassidy, Luiza M Mendonça, Grant J Jensen, Guenter P Resch, Ariane Briegel

Research output: Contribution to journal › Article › Academic › peer-review

Abstract

Cryo-electron microscopy has become an essential tool to understand structure and function of biological samples. Especially for pathogens, such as disease-causing bacteria and viruses, insights gained by cryo-EM can aid in developing cures. However, due to the biosafety restrictions of pathogens, samples are often treated by chemical fixation to render the pathogen inert, affecting the ultrastructure of the sample. Alternatively, researchers use in vitro or ex vivo models, which are non-pathogenic but lack the complexity of the pathogen of interest. Here we show that ultraviolet-C (UVC) radiation applied at cryogenic temperatures can be used to eliminate or dramatically reduce the infectivity of Vibrio cholerae and the bacterial virus, the ICP1 bacteriophage. We show no discernable structural impact of this treatment of either sample using two cryo-EM methods: cryo-electron tomography followed by sub-tomogram averaging, and single particle analysis (SPA). Additionally, we applied the UVC irradiation to the protein apoferritin (ApoF), which is a widely used test sample for high-resolution SPA studies. The UVC-treated ApoF sample resulted in a 2.1 Å structure indistinguishable from an untreated published map. This research demonstrates that UVC treatment is an effective and inexpensive addition to the cryo-EM sample preparation toolbox.

Original language	English
Article number	29
Pages (from-to)	1-8
Journal	Communications Biology
Volume	5
Issue number	1
DOIs	https://doi.org/10.1038/s42003-021-02962-w
Publication status	Published - Dec 2022
Externally published	Yes

Bibliographical note

Funding Information:
We are grateful to Davi Ortega for helpful discussions. We also wish to thank Weng Yang and Willem Noteborn of NeCEN for assistance during data collection, and Gert Koning (Department of Fine Mechanicsl) for assistance with the construction of the prototype UVC inactivation box. This work was funded in part by the Building Blocks of Life Grant 737.016.004 from the Netherlands Organization for Scientific Research (A.B.) and Instruct-ULTRA (Horizon 2020 Coordination and Support action Number ID: 731005 (A.B. and NeCEN). Funding was also provided to C.K.C. by UK Biotechnology and Biological Sciences Research Council grant number BB/S003339/1. G.J.J. and L.M.M. was funded by NIH grant AI150464.

Publisher Copyright:
© 2022, The Author(s).

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Cite this

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title = "UVC inactivation of pathogenic samples suitable for cryo-EM analysis",

abstract = "Cryo-electron microscopy has become an essential tool to understand structure and function of biological samples. Especially for pathogens, such as disease-causing bacteria and viruses, insights gained by cryo-EM can aid in developing cures. However, due to the biosafety restrictions of pathogens, samples are often treated by chemical fixation to render the pathogen inert, affecting the ultrastructure of the sample. Alternatively, researchers use in vitro or ex vivo models, which are non-pathogenic but lack the complexity of the pathogen of interest. Here we show that ultraviolet-C (UVC) radiation applied at cryogenic temperatures can be used to eliminate or dramatically reduce the infectivity of Vibrio cholerae and the bacterial virus, the ICP1 bacteriophage. We show no discernable structural impact of this treatment of either sample using two cryo-EM methods: cryo-electron tomography followed by sub-tomogram averaging, and single particle analysis (SPA). Additionally, we applied the UVC irradiation to the protein apoferritin (ApoF), which is a widely used test sample for high-resolution SPA studies. The UVC-treated ApoF sample resulted in a 2.1 {\AA} structure indistinguishable from an untreated published map. This research demonstrates that UVC treatment is an effective and inexpensive addition to the cryo-EM sample preparation toolbox.",

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note = "Funding Information: We are grateful to Davi Ortega for helpful discussions. We also wish to thank Weng Yang and Willem Noteborn of NeCEN for assistance during data collection, and Gert Koning (Department of Fine Mechanicsl) for assistance with the construction of the prototype UVC inactivation box. This work was funded in part by the Building Blocks of Life Grant 737.016.004 from the Netherlands Organization for Scientific Research (A.B.) and Instruct-ULTRA (Horizon 2020 Coordination and Support action Number ID: 731005 (A.B. and NeCEN). Funding was also provided to C.K.C. by UK Biotechnology and Biological Sciences Research Council grant number BB/S003339/1. G.J.J. and L.M.M. was funded by NIH grant AI150464. Publisher Copyright: {\textcopyright} 2022, The Author(s).",

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AU - Jensen, Grant J

AU - Resch, Guenter P

AU - Briegel, Ariane

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UVC inactivation of pathogenic samples suitable for cryo-EM analysis

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