Analysis of samples from three types of salivary gland obtained during a minimally invasive autopsy procedure performed on patients who died from complications of COVID-19 at Hospital das Clínicas, FM-USP’s hospital complex, showed that tissues specializing in producing and secreting saliva serve as reservoirs for the novel coronavirus.
The discovery helps explain why the virus is so abundant in saliva and has enabled scientists to develop saliva-based diagnostic tests for COVID-19.
“This is the first report of a respiratory virus’s capacity to infect and replicate in salivary glands,” said first author Bruno Fernandes Matuck, a PhD candidate at USP’s dental school.
“Until now, it was thought that only viruses that cause highly prevalent diseases such as herpes used salivary glands as reservoirs. The discovery may help explain why SARS-CoV-2 is so infectious,” he said.
A previous study by the same researchers had already demonstrated the presence of RNA from SARS-CoV-2 in the periodontal tissue of patients who died from COVID-19.
SARS-CoV-2 is highly infectious compared to other respiratory viruses, the researchers said, so they hypothesized that it may replicate in salivary gland cells and be present in saliva without coming into contact with nasal and lung secretions.
Prior research detected ACE2 receptors in salivary gland ducts. The spike protein in SARS-CoV-2 binds to ACE2 to invade and infect cells.
More recently, other researchers have conducted studies in animals showing that other receptors besides ACE2 such as transmembrane serine protease 2 (TMPRSS2) and furin, both of which are present in salivary glands, are targets of SARS-CoV-2.
To test this hypothesis in human beings, the researchers performed ultrasound-guided autopsies on 24 patients who died of COVID-19, with a mean age of 53, to extract tissue samples from the parotid, submandibular, and minor salivary glands.
The tissue samples were submitted to molecular analysis (RT-PCR), which detected the presence of the virus in more than two-thirds of the samples.
Immunohistochemistry, which is a form of immunostaining in which antibodies bind to the antigen in the tissue sample, a dye is activated, and the antigen can be seen under a microscope, also demonstrated the presence of the virus in the tissue.
Finally, examination under an electron microscope detected not just the presence of the virus but also its replication in cells and the type of organelle it uses to replicate.
“We observed several viruses clustering in salivary gland cells, which showed that they were replicating there. They weren’t in these cells passively,” said Matuck.
The researchers now plan to see whether the mouth can be a direct point of entry for SARS-CoV-2 given that ACE2 and TMPRSS2 are found in various parts of the oral cavity as well as in gum tissue and oral mucosa.
In addition, the mouth has a larger contact area than the nasal cavity, which is widely considered the main way into the body for the virus, the researchers said.
“We’re going to partner with researchers at the University of North Carolina in the United States to map the distribution of these receptors in the mouth and quantify viral replication in oral tissues,” said principal investigator Luiz Fernando Ferraz da Silva, a professor at FM-USP.
“The mouth could be a viable medium for the virus to enter the body directly,” said Matuck.
Another idea is to find out whether older people have more ACE2 receptors in their mouths than younger people, given the decrease in salivary secretion with age, the researchers said.
Nevertheless, the researchers said, they found a high viral load even in older patients, who have less salivary tissue.
“These patients had almost no salivary tissue, almost only fatty tissue. Even so, viral load was relatively high,” Matuck said.
The study, “Salivary Glands Are a Target for SARS-CoV-2: A Source for Saliva Contamination,” was published by the Journal of Pathology.