Study of a cold-adapted attenuated SARS-CoV-2 mutant using transmission, cryo-electron and atomic force microscopy

One of the traditional approaches to the creation of live attenuated vaccines is cold adaptation of the virus to produce temperature-sensitive (ts) mutants. In this work, we investigated the mor phological features and antigenic properties of the attenuated ts mutant F-F3 SARS-CoV-2 in comparison with the parent strain FEB2 (Omicron BA.5.2). Transmission electron microscopy of the virus inactivated by ultraviolet radiation revealed no significant differences in the morphol ogy of negatively contrasted virus particles and S-spikes: a characteristic “crown” consisting of spikes in the native prefusion conformation was found around the virions. Cryoelectron micros copy of the parent strain confirmed the presence of S-spikes in the prefusion conformation, while the ts mutant was not studied by this method due to insufficient virion concentration. In cubation with immune sera against an omicron-like strain revealed the formation of immune complexes both in the case of the parent strain and the ts mutant. Atomic force microscopy sug gested the presence of single virions in the preparation, but did not reveal a characteristic corona around them. This may be due to the fragility of S-spikes destroyed during sample preparation, or to the masking effect of serum albumin aggregates from the culture cell medium.

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