SARS-CoV-2: Ultrastructural Characterization of Morphogenesis in an In Vitro System

Abstract

The pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has impacted public health and the world economy and fueled a worldwide race to approve therapeutic and prophylactic agents, but so far there are no specific antiviral drugs. Understanding the biology of the virus is the first step in structuring strategies to combat it, and in this context several studies have been conducted with the aim of understanding the replication mechanism of SARS-CoV-2 in vitro systems. In this work, studies using transmission and scanning electron microscopy and 3D electron microscopy modeling were performed with the goal of characterizing the morphogenesis of SARS-CoV-2 in Vero-E6 cells. Several ultrastructural changes were observed—such as syncytia formation, cytoplasmic membrane projections, lipid droplets accumulation, proliferation of double-membrane vesicles derived from the rough endoplasmic reticulum, and alteration of mitochondria. The entry of the virus into cells occurred through endocytosis. Viral particles were observed attached to the cell membrane and in various cellular compartments, and extrusion of viral progeny took place by exocytosis. These findings allow us to infer that Vero-E6 cells are highly susceptible to SARS-CoV-2 infection as described in the literature and their replication cycle is similar to that described with SARS-CoV and MERS-CoV in vitro models.

Ultrastructural analysis and three-dimensional reconstruction of cellular structures involved in SARS-CoV-2 spread

“…Filopodia growth induced by SARS-CoV-2 infection in cells has been described in previous studies (Barreto-Vieira et al 2022). These nanostructures were slightly finer and more homogeneous in the control cells (104 ± 22 nm) compared to the infected cells (120 ± 47 nm).…”

“…Several studies have already characterized cell alterations at the ultrastructural level during SARS-CoV-2 infection. The accumulation of cytoplasmic lipid droplets, the thickening of the rough endoplasmic reticulum (RER), and its associated proliferation of double-membrane vesicles (DMV), the mitochondrial alterations, and the formation of filopodia as membrane projections have been well described (Goldsmith et al 2004;Qinfen et al 2004;Tseng et al 2005;Snijder et al 2006;Knoops et al 2008;Klein et al 2020;Barreto-Vieira et al 2022). Nevertheless, cytoskeleton alterations have been poorly assessed.…”

“…Ultrastructural analyses of morphogenesis in SARS-CoV-2-infected cells point out filopodia formation as plasma membrane projections (Ng et al 2004;Caldas et al 2020;Bideau et al 2021;Barreto-Vieira et al 2022). However, uncertainty remains about the involvement of the cellular cytoskeleton in SARS-CoV-2 infection.…”

Newly Emerged Antiviral Strategies for SARS-CoV-2: From Deciphering Viral Protein Structural Function to the Development of Vaccines, Antibodies, and Small Molecules

“…Membrane (M): The M protein is one of the structural proteins. It comprises the virus envelope and contributes to viral morphogenesis [ 7 , 16 ]. The similarity of genomic sequences encoding the M proteins between SARS-CoV-1 and SARS-CoV-2 is significantly high [ 17 , 18 , 19 ].…”

Estimation of virus-mediated cell fusion rate of SARS-CoV-2

“…One possible contributor to both damage and disease severity is the formation of multinucleated cells known as syncytia. SARS-CoV-2 is known to have the ability to form syncytia ( Braga et al, 2021 ; Barreto-Vieira et al, 2022 ; Hoffmann et al, 2020 ) that can contribute to disease severity in a number of ways. The formation of syncytia causes physical tissue damage that results in more severe clinical symptoms ( Bussani et al, 2020 ).…”