The SARS-CoV-2 viral load in COVID-19 patients is lower on face mask filters than on nasopharyngeal swabs.

Face masks and personal respirators are used to curb the transmission of SARS-CoV-2 in respiratory droplets; filters embedded in some personal protective equipment could be used as a non-invasive sample source for applications, including at-home testing, but information is needed about whether filters are suited to capture viral particles for SARS-CoV-2 detection. In this study, we generated inactivated virus-laden aerosols of 0.3-2 microns in diameter (0.9 µm mean diameter by mass) and dispersed the aerosolized viral particles onto electrostatic face mask filters. The limit of detection for inactivated coronaviruses SARS-CoV-2 and HCoV-NL63 extracted from filters was between 10 to 100 copies/filter for both viruses. Testing for SARS-CoV-2, using face mask filters and nasopharyngeal swabs collected from hospitalized COVID-19-patients, showed that filter samples offered reduced sensitivity (8.5% compared to nasopharyngeal swabs). The low concordance of SARS-CoV-2 detection between filters and nasopharyngeal swabs indicated that number of viral particles collected on the face mask filter was below the limit of detection for all patients but those with the highest viral loads. This indicated face masks are unsuitable to replace diagnostic nasopharyngeal swabs in COVID-19 diagnosis. The ability to detect nucleic acids on face mask filters may, however, find other uses worth future investigation.

The r131 gene of rat cytomegalovirus encodes a proinflammatory CC chemokine homolog which is essential for the production of infectious virus in the salivary glands.

Rat cytomegalovirus (RCMV) possesses two adjacent genes, r131 and r129, which have the potential to encode CC chemokine homologs. Interestingly, the amino acid sequences encoded by both genes show similarity to the sequence of the murine CMV (MCMV) MCK-2 protein, which is encoded by the m131/129 gene. In order to study the significance of the r131 gene in the pathogenesis of RCMV infection, we generated two different virus strains in which the r131 open reading frame is disrupted. Replication of these null mutant strains, designated RCMVdeltar131a and RCMVdeltar131b, was evaluated in vitro and in vivo. Both strains were found to replicate with a similar efficiency as wild-type (WT) RCMV in vitro. However, in contrast to WT virus, neither RCMVdeltar131a nor RCMVdeltar131b established a high-titer infection in the salivary glands of immunocompromised rats. Furthermore, in a local, rat footpad infection model, both recombinant viruses induced a significantly lower amount of paw swelling than did WT RCMV. Also, a higher number of infiltrating macrophages was observed in paws infected with WT RCMV than in those infected with the recombinants. Taken together, these results suggest that r131 (i) promotes inflammation at initial sites of inoculation and, subsequently, efficient virus dissemination to or infection of the salivary glands and (ii) might be involved in the persistence of virus infection, at least in the spleen. In addition, our data indicate that r131 represents the functional homolog of the MCMV m131/129 gene.

Mutational analysis of the R33-encoded G protein-coupled receptor of rat cytomegalovirus: identification of amino acid residues critical for cellular localization and ligand-independent signalling.

The rat cytomegalovirus (RCMV) R33 gene encodes a G protein-coupled receptor (GPCR), pR33, which possesses agonist-independent, constitutive signalling activity. To characterize this activity further, we generated a series of point and deletion mutants of pR33. Both expression of and signalling by the mutants was evaluated. Several point mutants were generated that contained modifications in the NRY motif. This motif, at aa 130-132 of pR33, is the counterpart of the common DRY motif of GPCRs, which is known to be involved in G protein coupling. We found that mutation of the asparagine residue within the NRY motif of pR33 (N(130)) to aspartic acid resulted in a mutant (N(130)D) with similar signalling characteristics to the wild-type (WT) protein, indicating that N(130) is not the determinant of constitutive activity of pR33. Interestingly, a mutant carrying an alanine at aa 130 (N(130)A) was severely impaired in G(q/11)-mediated, constitutive activation of phospholipase C, whereas it displayed similar levels of activity to pR33 in G(i/0)-mediated signalling. Another protein that contained a modified NRY motif, R(131)A, did not show constitutive activity, whereas mutants Y(132)F and Y(132)A displayed similar activities to the WT receptor. This indicated that residue R(131) is critical for pR33 function in vitro, whereas Y(132) is not. Finally, we identified two consecutive arginines within the C-terminal tails of both pR33 and its homologue from human CMV, pUL33, which are important for correct cell-surface expression of these receptors.