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The impact of variants of concern (VoC) on SARS-CoV-2 viral dynamics remains poorly understood and essentially relies on observational studies subject to various sorts of biases. In contrast, experimental models of infection constitute a powerful model to perform controlled comparisons of the viral dynamics observed with VoC and better quantify how VoC escape from the immune response. Here we used molecular and infectious viral load of 78 cynomolgus macaques to characterize in detail the effects of VoC on viral dynamics. We first developed a mathematical model that recapitulate the observed dynamics, and we found that the best model describing the data assumed a rapid antigen-dependent stimulation of the immune response leading to a rapid reduction of viral infectivity. When compared with the historical variant, all VoC except beta were associated with an escape from this immune response, and this effect was particularly sensitive for delta and omicron variant (p<10-6 for both). Interestingly, delta variant was associated with a 1.8-fold increased viral production rate (p=0.046), while conversely omicron variant was associated with a 14-fold reduction in viral production rate (p<10-6). During a natural infection, our models predict that delta variant is associated with a higher peak viral RNA than omicron variant (7.6 log10 copies/mL 95% CI 6.8 - 8 for delta; 5.6 log10 copies/mL 95% CI 4.8 - 6.3 for omicron) while having similar peak infectious titers (3.7 log10 PFU/mL 95% CI 2.4 - 4.6 for delta; 2.8 log10 PFU/mL 95% CI 1.9 - 3.8 for omicron). These results provide a detailed picture of the effects of VoC on total and infectious viral load and may help understand some differences observed in the patterns of viral transmission of these viruses.
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SARS-CoV-2 infection results in impaired interferon response in severe COVID-19 patients. However, how SARS-CoV-2 interferes with host immune response is incompletely understood. Here, we sequenced small RNAs from SARS-CoV-2-infected human cells and identified a micro-RNA (miRNA) encoded in a recently evolved region of the viral genome. We show that the virus-encoded miRNA produces two miRNA isoforms in infected cells by the enzyme Dicer and they are loaded into Argonaute proteins. Moreover, the predominant miRNA isoform targets the 3UTR of interferon-stimulated genes and represses their expression in a miRNA-like fashion. Finally, the two viral miRNA isoforms were detected in nasopharyngeal swabs from COVID-19 patients. We propose that SARS-CoV-2 employs a virus-encoded miRNA to hijack the host miRNA machinery and evade the interferon-mediated immune response.
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SARS-CoV-2 has infected almost 200 million humans and caused over 4 million deaths worldwide. Evaluating countermeasures and improving our understanding of COVID-19 pathophysiology require access to animal models that replicate the hallmarks of human disease. Mouse infection with SARS-CoV-2 is limited by poor affinity between the virus spike protein and its cellular receptor ACE2. We have developed by serial passages the MACo3 virus strain which efficiently replicates in the lungs of standard mouse strains and induces age-dependent lung lesions. Compared to other mouse-adapted strains and severe mouse models, infection with MACo3 results in mild to moderate disease and will be useful to investigate the role of host genetics and other factors modulating COVID-19 severity.
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Interferon restricts SARS-CoV-2 replication in cell culture, but only a handful of Interferon Stimulated Genes with antiviral activity against SARS-CoV-2 have been identified. Here, we describe a functional CRISPR/Cas9 screen aiming at identifying SARS-CoV-2 restriction factors. We identified DAXX, a scaffold protein residing in PML nuclear bodies known to limit the replication of DNA viruses and retroviruses, as a potent inhibitor of SARS-CoV-2 and SARS-CoV replication in human cells. Basal expression of DAXX was sufficient to limit the replication of SARS-CoV-2, and DAXX over-expression further restricted infection. In contrast with most of its previously described antiviral activities, DAXX-mediated restriction of SARS-CoV-2 was independent of the SUMOylation pathway. SARS-CoV-2 infection triggered the re-localization of DAXX to cytoplasmic sites and promoted its degradation. Mechanistically, this process was mediated by the viral papain-like protease (PLpro) and the proteasome. Together, these results demonstrate that DAXX restricts SARS-CoV-2, which in turn has evolved a mechanism to counteract its action.
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Receptor recognition is a major determinant of viral host range, infectivity and pathogenesis. Emergences have been associated with serendipitous events of adaptation upon encounters with novel hosts, and the high mutation rate of RNA viruses may explain their frequent host shifts. SARS-CoV-2 extensive circulation in humans results in the emergence of variants, including variants of concern (VOCs) with diverse mutations notably in the spike, and increased transmissibility or immune escape. Here we show that, unlike the initial and Delta variants, the three VOCs bearing the N501Y mutation can infect common laboratory mice. Contact transmission occurred from infected to naive mice through two passages. This host range expansion likely results from an increased binding of the spike to the mouse ACE2. Together with the observed contact transmission, it raises the possibility of wild rodent secondary reservoirs enabling the emergence of new variants.
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SARS-CoV-2 B.1.1.7 and B.1.351 variants emerged respectively in United Kingdom and South Africa and spread in many countries. Here, we isolated infectious B.1.1.7 and B.1.351 strains and examined their sensitivity to anti-SARS-CoV-2 antibodies present in sera and nasal swabs, in comparison with a D614G reference virus. We established a novel rapid neutralization assay, based on reporter cells that become GFP+ after overnight infection. B.1.1.7 was neutralized by 79/83 sera from convalescent patients collected up to 9 months post symptoms, almost similar to D614G. There was a mean 6-fold reduction in titers and even loss of activity against B.1.351 in 40% of convalescent sera after 9 months. Early sera from 19 vaccinated individuals were almost as potent against B.1.1.7 but less efficacious against B.1.351, when compared to D614G. Nasal swabs from vaccine recipients were not neutralizing, except in individuals who were diagnosed COVID-19+ before vaccination. Thus, faster-spreading variants acquired a partial resistance to humoral immunity generated by natural infection or vaccination, mostly visible in individuals with low antibody levels.
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BackgroundChildren have a lower rate of COVID-19, potentially related to cross-protective immunity conferred by seasonal coronaviruses (HCoVs). We tested if prior infections with seasonal coronaviruses impacted SARS-CoV-2 infections and related Multisystem Inflammatory Syndrome (MIS). MethodsThis cross-sectional observational study in Paris hospitals enrolled 739 pauci or asymptomatic children (HOS group) plus 36 children with suspected MIS (MIS group). Prevalence, antigen specificity and neutralizing capability of SARS-CoV-2 antibodies were tested. Antibody frequency and titres against Nucleocapsid (N) and Spike (S) of the four seasonal coronaviruses (NL63, HKU1, 229E, OC43) were measured in a subset of seropositive patients (54 SARS-CoV-2 (HOS-P subgroup) and 15 MIS (MIS-P subgroup)), and in 118 matched SARS-CoV-2 seronegative patients (CTL subgroup). FindingsSARS-CoV-2 mean prevalence rate in HOSP children was 11.7% from April 1 to June 1. Neutralizing antibodies were found in 55{middle dot}6% of seropositive children, and their relative frequency increased with time (up to 100 % by mid-May). A majority of MIS children (25/36) were SARS-CoV-2 seropositive, of which all tested (n=15) had neutralizing antibodies. On average, seropositive MIS children had higher N and S1 SARS-CoV-2 titres as compared to HOS children. Patients from HOS-P, MIS-P, and CTL subgroups had a similar prevalence of antibodies against the four seasonal HCoVs (66{middle dot}9 -100%). The level of anti-SARS-CoV-2 antibodies was not significantly different in children who had prior seasonal coronavirus infection. InterpretationPrior infection with HCoVs does not prevent SARS-CoV-2 infection and related MIS in children. Children develop neutralizing antibodies after SARS-CoV-2 infection. Evidence before this studyChildren seem to be less likely affected by SARS-CoV-2 infection and clinical course of COVID-19 is less severe than in adults. As those asymptomatic or mildly symptomatic children are underdiagnosed and their viral loads are comparable to those of adults, they may act as an asymptomatic reservoir for the spread of the virus. One explanation of the difference between the adult and the pediatric infectious profile might be that infection with seasonal human coronaviruses, which is very frequent from a very young age, could lead to cross protective immunity. We searched in PubMed, MedRxiv and BioRxiv for publications from inception to June 15, 2020, using the terms "COVID-19, SARS-CoV-2, children, serology, Kawasaki, Corona Virus". Added value of this studySARS-CoV-2 mean prevalence rate was 11.7% from April 1 to June 1 and neutralizing antibodies were found in 55% of the tested seropositive children. Among patients with a Multisystem Inflammatory Syndrome, Kawasaki-like disease, 70% were SARS-CoV-2 seropositive and had neutralizing antibodies. COVID-19 and MIS attack rates, and anti-SARS-CoV-2 antibodies titres were not significantly impacted by prior seasonal coronavirus infection. Implications of all the available evidencePrior infection by seasonal coronaviruses does not prevent SARS-CoV-2 infection and associated Multisystem Inflammatory Syndrome in children As antibodies against seasonal coronaviruses are very frequent and as these viruses circulate efficiently in human populations every winter, our results question to what extent the concept of herd immunity based on circulating antibodies can be applied to seasonal coronaviruses and possibly SARS-CoV-2.
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Following the emergence of coronavirus disease (COVID-19) in Wuhan, China in December 2019, specific COVID-19 surveillance was launched in France on January 10, 2020. Two weeks later, the first three imported cases of COVID-19 into Europe were diagnosed in France. We sequenced 97 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes from samples collected between January 24 and March 24, 2020 from infected patients in France. Phylogenetic analysis identified several early independent SARS-CoV-2 introductions without local transmission, highlighting the efficacy of the measures taken to prevent virus spread from symptomatic cases. In parallel, our genomic data reveals the later predominant circulation of a major clade in many French regions, and implies local circulation of the virus in undocumented infections prior to the wave of COVID-19 cases. This study emphasizes the importance of continuous and geographically broad genomic sequencing and calls for further efforts with inclusion of asymptomatic infections.
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It is of paramount importance to evaluate the prevalence of both asymptomatic and symptomatic cases of SARS-CoV-2 infection and their antibody response profile. Here, we performed a pilot study to assess the levels of anti-SARS-CoV-2 antibodies in samples taken from 491 pre-epidemic individuals, 51 patients from Hopital Bichat (Paris), 209 pauci-symptomatic individuals in the French Oise region and 200 contemporary Oise blood donors. Two in-house ELISA assays, that recognize the full-length nucleoprotein (N) or trimeric Spike (S) ectodomain were implemented. We also developed two novel assays: the S-Flow assay, which is based on the recognition of S at the cell surface by flow-cytometry, and the LIPS assay that recognizes diverse antigens (including S1 or N C-terminal domain) by immunoprecipitation. Overall, the results obtained with the four assays were similar, with differences in sensitivity that can be attributed to the technique and the antigen in use. High antibody titers were associated with neutralisation activity, assessed using infectious SARS-CoV-2 or lentiviral-S pseudotypes. In hospitalized patients, seroconversion and neutralisation occurred on 5-14 days post symptom onset, confirming previous studies. Seropositivity was detected in 29% of pauci-symptomatic individuals within 15 days post-symptoms and 3 % of blood of healthy donors collected in the area of a cluster of COVID cases. Altogether, our assays allow for a broad evaluation of SARS-CoV2 seroprevalence and antibody profiling in different population subsets.