Ultrastructural analysis of nasopharyngeal epithelial cells from patients with SARS-CoV-2 infection

Abstract

The nasal epithelium is an initial site for SARS-CoV-2 infection, responsible for the ongoing COVID-19 pandemic. However, the pathogenicity and morphological impact of SARS-CoV-2 on the nasopharynx cells from symptomatic patients with different viral loads remain poorly understood. Here, we investigated the ultrastructure of nasal cells obtained from individuals at distinct disease days and with high and low SARS-CoV-2 loads. Squamous and ciliated cells were the main cells observed in SARS-CoV-2 negative samples. We identified virus-like particles (VLPs) and replication organelles (RO)-like structures in the squamous cells from high viral load samples after 3- and 4-days of symptoms. Ultrastructural changes were found in those cells, such as the loss of microvilli and primary cilium, the increase of multivesicular bodies and autophagosomes, and signs of cell death. No ciliated cells were found in those samples. Squamous cells from low viral load sample after 5 days of symptoms showed few microvilli and no primary cilium. VLPs and RO-like structures were found in the ciliated cells only. No ultrastructural alterations were seen in the cells from low viral load individuals after 10- and 14-days of symptoms. Our results shed light on the ultrastructural effects of SARS-CoV-2 infection on the human nasopharyngeal cells.

Detection and identification of coronaviruses in human tissues using electron microscopy

“…A SARS-coronavirus specific antibody is applied to a thin section, followed by a secondary antibody conjugated to colloidal gold particles. The gold particles appear as electron dense black dots in areas where coronavirus is present or where the protein that the primary antibody targets is present, such as nucleocapsid inclusions (Figure 4a Respiratory system changes due to SARS-CoV-2 infection have been most thoroughly described (Ackermann et al, 2020;Martines et al, 2020;Santana et al, 2021;Saraiva et al, 2021). Type I pneumocyte degeneration, type II pneumocyte hyperplasia, and hyaline membrane formation were common ultrastructural features of SARS-CoV-2 infection (Martines et al, 2020;Santana et al, 2021).…”

“…Type I pneumocyte degeneration, type II pneumocyte hyperplasia, and hyaline membrane formation were common ultrastructural features of SARS‐CoV‐2 infection (Martines et al, 2020 ; Santana et al, 2021 ). A recent study of nasopharyngeal epithelial cells from SARS‐CoV‐2 infected individuals showed loss of microvilli and the primary cilium in squamous cells as well as an increase in the number of multivesicular bodies and autophagosomes when compared with specimens from SARS‐CoV‐2 negative individuals (Saraiva et al, 2021 ). Santana et al further noted disruption of the epithelial‐endothelial barrier, in addition to cellular debris and fibrin in the alveolar space and detachment of type II pneumocytes from the basement membrane (Santana et al, 2021 ).…”

“…An increasing number of articles have included descriptions of ultrastructural changes in SARS‐CoV‐2 positive autopsy tissues (Ackermann et al, 2020 ; Akilesh, Nast, et al, 2021 ; Deinhardt‐Emmer, Böttcher, et al, 2021 ; Deinhardt‐Emmer, Wittschieber, et al, 2021 ; Duarte‐Neto et al, 2021 ; Lüke et al, 2020 ; Rizzo et al, 2021 ; Santana et al, 2021 ; Saraiva et al, 2021 ). These articles do not necessarily include EM images of viral particles but describe possible ultrastructural alterations due to infection.…”

“…Comparison of infected and healthy tissues is necessary, and correlation with results of other diagnostic techniques is always recommended. Respiratory system changes due to SARS‐CoV‐2 infection have been most thoroughly described (Ackermann et al, 2020 ; Martines et al, 2020 ; Santana et al, 2021 ; Saraiva et al, 2021 ). Type I pneumocyte degeneration, type II pneumocyte hyperplasia, and hyaline membrane formation were common ultrastructural features of SARS‐CoV‐2 infection (Martines et al, 2020 ; Santana et al, 2021 ).…”

“…Grootemaat et al have elegantly used this technique to demonstrate antigens of coronavirus grown in tissue culture(Grootemaat et al, 2022). Despite these limitations, IEM can still be a valuable tool to confirm the presence of a virus in cells.5 | ULTRASTRUCTURAL CHANGES CAUSED BY SARS-COV-2 INFECTIONAn increasing number of articles have included descriptions of ultrastructural changes in SARS-CoV-2 positive autopsy tissues(Ackermann et al, 2020;Akilesh, Nast, et al, 2021;Deinhardt-Emmer, Böttcher, et al, 2021; Deinhardt-Emmer, Wittschieber, et al, 2021;Duarte-Neto et al, 2021;Lüke et al, 2020;Rizzo et al, 2021;Santana et al, 2021;Saraiva et al, 2021). These articles do not necessarily include EM images of viral particles but describe possible ultrastructural alterations due to infection.…”