ABSTRACT
The strong humoral immune response produced against the SARS-CoV-2 nucleocapsid (N) and spike (S) proteins has underpinned serological testing but the prevalence of antibody responses to other SARS-CoV-2 proteins, which may be of use as further serological markers, is still unclear. Cell-based serological screening platforms can fulfil a crucial niche in the identification of antibodies which recognise more complex folded epitopes or those incorporating post-translation modifications which may be undetectable by other methods used to investigate the antigenicity of the SARS-CoV-2 proteome. Here, we employed automated high content immunofluorescence microscopy (AHCIM) to assess the viability of such an approach as a method capable of assaying humoral immune responses against full length SARS-CoV-2 proteins in their native cellular state. We first demonstrate that AHCIM provides high sensitivity and specificity in the detection of SARS-CoV-2 N and S IgG. Assessing the prevalence of antibody responses to the SARS-CoV-2 structural membrane protein (M), we further find that 85% of COVID-19 patients within our sample set developed detectable M IgG responses (M sensitivity 85%, N sensitivity 93%, combined N + M sensitivity 95%). The identification of M as a serological marker of high prevalence may be of value in detecting additional COVID-19 cases during the era of mass SARS-CoV-2 vaccinations, where serological screening for SARS CoV-2 infections in vaccinated individuals is dependent on detection of antibodies against N. These findings highlight the advantages of using cell-based systems as serological screening platforms and raise the possibility of using M as a widespread serological marker alongside N and S.
Subject(s)
Severe Acute Respiratory Syndrome , COVID-19ABSTRACT
The antiviral restriction factor, tetherin, blocks the release of several different families of enveloped viruses, including the Coronaviridae. Tetherin is an interferon-induced protein that forms parallel homodimers between the host cell and viral particles, linking viruses to the surface of infected cells and inhibiting their release. We demonstrate that SARS-CoV-2 downregulates tetherin to aid its release from cells, and investigate potential proteins involved in this process. Loss of tetherin from cells caused an increase in SARS-CoV-2 viral titre. We find SARS-CoV-2 spike protein to be responsible for tetherin downregulation, rather than ORF7a as previously described for the 2002-2003 SARS-CoV. We instead find ORF7a to be responsible for Golgi fragmentation, and expression of ORF7a in cells recapitulates Golgi fragmentation observed in SARS-CoV-2 infected cells.
Subject(s)
Severe Acute Respiratory SyndromeABSTRACT
A novel coronavirus, SARS-CoV-2, has caused over 8538 million cases and over 1.8 1 million deaths worldwide since it occurred twelve months ago in Wuhan, China. Here we first analyzed 4,013 full-length SARS-CoV-2 genomes from different continents over a 14-week timespan since the outbreak in Wuhan, China. 2,954 unique nucleotide substitutions were identified with 31 of the 4,013 genomes remaining as ancestral type, and 952 (32.2%) mutations recurred in more than one genome. A viral genotype from the Seafood Market in Wuhan featured with two concurrent mutations was the dominant genotype (80.9%) of the pandemic. We also identified unique genotypic compositions from different geographic locations, and time-series viral genotypic dynamics in the early phase that reveal transmission routes and subsequent expansion. We also used the same approach to analyze 261,350 full-length SARS-CoV-2 genomes from the world over 12 months since the outbreak (i.e. all the available viral genomes in the GISAID database as of 25 December 2020). Our study indicates the viral genotypes can be utilized as molecular barcodes in combination with epidemiologic data to monitor the spreading routes of the pandemic and evaluate the effectiveness of control measures.