Viral load and infectivity of SARS-CoV-2 in paired respiratory and oral specimens from symptomatic, asymptomatic or post-symptomatic individuals

In the present study, we assessed the diagnostic sensitivity and determined the viral load and infectivity of SARS-CoV-2 in paired respiratory (nasopharyngeal and anterior nares) and oral samples (saliva and sublingual swab). Samples were collected from 77 individuals of which 75 were diagnosed with COVID-19 and classified as symptomatic (n=29), asymptomatic (n=31), or post-symptomatic (n=15). Specimens were collected at one time point from each individual, between day 1 to 23 after the initial COVID-19 diagnosis, and included self-collected saliva (S), or sublingual (SL) swab, and bilateral anterior nares (AN) swab, followed by healthcare provider collected nasopharyngeal (NP) swab. Sixty-three specimen sets were tested using five assay/platforms. The diagnostic sensitivity of each assay/platform and specimen type was determined. Of the 63 specimen sets, SARS-CoV-2 was detected in 62 NP specimens, 52 AN specimens, 59 saliva specimens, and 31 SL specimens by at least one platform. Infectious SARS-CoV-2 was isolated from 21 NP, 13 AN, 12 saliva, and one SL specimen out of 50 specimen sets. SARS-CoV-2 isolation was most successful up to 5 days after initial COVID-19 diagnosis using NP specimens from symptomatic patients (16 of 24 positives, 66.67%), followed by specimens from asymptomatic patients (5 of 17 positives, 29.41%), while it was not very successful with specimens from post-symptomatic patients. Benefits of self-collected saliva and AN specimens balance the loss of sensitivity relative to NP specimens. Therefore, saliva and AN specimens are acceptable alternatives for symptomatic SARS-CoV-2 diagnostic testing or surveillance with increased sampling frequency of asymptomatic individuals. ImportanceThe dynamics of infection with SARS-CoV-2 has a significant impact on virus infectivity and in the diagnostic sensitivity of molecular and classic virus detection tests. In the present study we determined the diagnostic sensitivity of paired respiratory (nasopharyngeal and anterior nares swabs) and oral secretions (saliva and sublingual swab) and assessed infectious virus shedding patterns by symptomatic, asymptomatic or post-symptomatic individuals. Understanding the diagnostic performance of these specimens and the patterns of infectious virus shedding in these bodily secretions provides critical information to control COVID-19, and may help to refine guidelines on isolation and quarantine of positive individuals and their close contacts identified through epidemiological investigations.

Age-related susceptibility of ferrets to SARS-CoV-2 infection

Susceptibility to SARS-CoV-2 and the outcome of COVID-19 have been linked to underlying health conditions and the age of affected individuals. Here we assessed the effect of age on SARS-CoV-2 infection using a ferret model. For this, young (6-month-old) and aged (18-to-39-month-old) ferrets were inoculated intranasally with various doses of SARS-CoV-2. By using infectious virus shedding in respiratory secretions and seroconversion, we estimated that the infectious dose of SARS-CoV-2 in aged animals is [~]32 plaque forming units (PFU) per animal while in young animals it was estimated to be [~]100 PFU. We showed that viral replication in the upper respiratory tract and shedding in respiratory secretions is enhanced in aged ferrets when compared to young animals. Similar to observations in humans, this was associated with higher expressions levels of two key viral entry factors - ACE2 and TMPRSS2 - in the upper respiratory tract of aged ferrets.

Functional evaluation of proteolytic activation for the SARS-CoV-2 variant B.1.1.7: role of the P681H mutation

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the agent causing the COVID-19 pandemic. SARS-CoV-2 B.1.1.7 (Alpha), a WHO variant of concern (VOC) first identified in the UK in late 2020, contains several mutations including P681H in the spike S1/S2 cleavage site, which is predicted to increase cleavage by furin, potentially impacting the viral cell entry. Here, we studied the role of the P681H mutation in B.1.1.7 cell entry. We performed assays using fluorogenic peptides mimicking the Wuhan-Hu-1 and B.1.1.7 S1/S2 sequence and observed no significant difference in furin cleavage. Functional assays using pseudoparticles harboring SARS-CoV-2 spikes and cell-to-cell fusion assays demonstrated no differences between Wuhan-Hu-1, B.1.1.7 or a P681H point mutant. Likewise, we observed no differences in viral growth between USA-WA1/2020 and a B.1.1.7 isolate in cell culture. Our findings suggest that while the B.1.1.7 P681H mutation may slightly increase S1/S2 cleavage this does not significantly impact viral entry or cell-cell spread. HighlightsO_LISARS-CoV-2 B.1.1.7 VOC has a P681H mutation in the spike that is predicted to enhance viral infection C_LIO_LIP681H does not significantly impact furin cleavage, viral entry or cell-cell spread C_LIO_LIOther mutations in the SARS-CoV-2 B.1.1.7 VOC may account for increased infection rates C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=134 SRC="FIGDIR/small/438731v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@c148d7org.highwire.dtl.DTLVardef@1954eeeorg.highwire.dtl.DTLVardef@171130dorg.highwire.dtl.DTLVardef@99bd45_HPS_FORMAT_FIGEXP M_FIG C_FIG