The SARS-CoV-2 Omicron sub-variant BA.2.86 is attenuated in hamsters (preprint)

SARS-CoV-2 variants have emerged throughout the COVID-19 pandemic. There is a need to risk-assess newly emerged variants in near "real-time" to estimate their potential threat to public health. The recently emerged Omicron sub-variant BA.2.86 raised concerns as it carries a high number of mutations compared to its predecessors. Here, we assessed the virulence of BA.2.86 in hamsters. We compared the pathogenesis of BA.2.86 and BA.2.75, as the latter is one of the most virulent Omicron sub-variants in this animal model. Using digital pathology pipelines, we quantified the extent of pulmonary lesions measuring T cell and macrophage infiltrates, in addition to alveolar epithelial hyperplasia. We also assessed body weight loss, clinical symptoms, virus load in oropharyngeal swabs, and virus replication in the respiratory tract. Our data show that BA.2.86 displays an attenuated phenotype in hamsters, suggesting that it poses no greater risk to public health than its parental Omicron sub-variants. Article summary lineThe newly emerged Omicron sub-variant BA.2.86 is attenuated in hamsters.

The P681H mutation in the Spike glycoprotein escapes IFITM restriction and is necessary for type I interferon resistance in the SARS-CoV-2 alpha variant (preprint)

The appearance of new dominant variants of concern (VOCs) of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) threatens the global response to the COVID-19 pandemic. Of these, the alpha variant (also known as B.1.1.7) that appeared initially in the UK became the dominant variant in much of Europe and North America in the first half of 2021. The Spike (S) glycoprotein of alpha acquired seven mutations and two deletions compared to the ancestral virus, including the P681H mutation in the polybasic cleavage site that has been suggested to enhance S cleavage. Here, we show that the alpha S protein confers a level of resistance to the effects of interferon-b (IFNb) in human lung epithelial cells. This correlates with resistance to an entry restriction mediated by interferon-induced transmembrane protein 2 (IFITM2) and a pronounced infection enhancement by IFITM3. Furthermore, the P681H mutation is essential for resistance to IFNb and context-dependent resistance to IFITMs in the alpha S. However, while this appears to confer changes in sensitivity to endosomal protease inhibition consistent with enhanced cell-surface entry, its reversion does not reduce cleaved S incorporation into particles, indicating a role downstream of furin cleavage. Overall, we suggest that, in addition to adaptive immune escape, mutations associated with VOCs may well also confer replication and/or transmission advantage through adaptation to resist innate immune mechanisms.

Enhanced innate immune suppression by SARS-CoV-2 Omicron subvariants BA.4 and BA.5 (preprint)

SARS-CoV-2 adaptation to its human host is evidenced by the emergence of new viral lineages with distinct genotypic and phenotypic characteristics, termed variants of concern (VOCs). Particular VOCs have become sequentially dominant globally (Alpha, Delta, Omicron) with each evolving independently from the ancestral Wuhan strain. Omicron is notable for its large number of Spike mutations found to promote immune escape and re-infection. Most recently, Omicron BA.4 and BA.5 subvariants have emerged with increasing levels of adaptive immune escape threatening vaccine effectiveness and increasing hospitalisations. Here, we demonstrate that the most recent Omicron variants have enhanced capacity to antagonise or evade human innate immune defenses. We find Omicron BA.4 and BA.5 replication is associated with reduced activation of epithelial innate immune responses versus earlier BA.1 and BA.2 subvariants. We also find enhanced expression of innate immune antagonist proteins Orf6 and N, similar to Alpha, suggesting common pathways of human adaptation and linking VOC dominance to improved innate immune evasion. We conclude that Omicron BA.4 and BA.5 have combined evolution of antibody escape with enhanced antagonism of human innate immunity to improve transmission and possibly reduce immune protection from severe disease.

The P681H mutation in the Spike glycoprotein confers Type I interferon resistance in the SARS-CoV-2 alpha (B.1.1.7) variant (preprint)

Variants of concern (VOCs) of severe acute respiratory syndrome coronavirus type-2 (SARS-CoV-2) threaten the global response to the COVID-19 pandemic. The alpha (B.1.1.7) variant appeared in the UK became dominant in Europe and North America in early 2021. The Spike glycoprotein of alpha has acquired a number mutations including the P681H mutation in the polybasic cleavage site that has been suggested to enhance Spike cleavage. Here, we show that the alpha Spike protein confers a level of resistance to the effects of interferon-{beta} (IFN{beta}) in lung epithelial cells. This correlates with resistance to restriction mediated by interferon-induced transmembrane protein-2 (IFITM2) and a pronounced infection enhancement by IFITM3. Furthermore, the P681H mutation is necessary for comparative resistance to IFN{beta} in a molecularly cloned SARS-CoV-2 encoding alpha Spike. Overall, we suggest that in addition to adaptive immune escape, mutations associated with VOCs also confer replication advantage through adaptation to resist innate immunity.

Mutations that adapt SARS-CoV-2 to mustelid hosts do not increase fitness in the human airway. (preprint)

SARS-CoV-2 has a broad mammalian species tropism infecting humans, cats, dogs and farmed mink. Since the start of the 2019 pandemic several reverse zoonotic outbreaks of SARS-CoV-2 have occurred in mink, one of which reinfected humans and caused a cluster of infections in Denmark. Here we investigate the molecular basis of mink and ferret adaptation and demonstrate the spike mutations Y453F, F486L, and N501T all specifically adapt SARS-CoV-2 to use mustelid ACE2. Furthermore, we risk assess these mutations and conclude mink-adapted viruses are unlikely to pose an increased threat to humans, as Y453F attenuates the virus replication in human cells and all 3 mink-adaptations have minimal antigenic impact. Finally, we show that certain SARS-CoV-2 variants emerging from circulation in humans may naturally have a greater propensity to infect mustelid hosts and therefore these species should continue to be surveyed for reverse zoonotic infections.

Reduced neutralisation of the Delta (B.1.617.2) SARS-CoV-2 variant of concern following vaccination (preprint)

Vaccines are proving to be highly effective in controlling hospitalisation and deaths associated with SARS-CoV-2 infection but the emergence of viral variants with novel antigenic profiles threatens to diminish their efficacy. Assessment of the ability of sera from vaccine recipients to neutralise SARS-CoV-2 variants will inform the success of strategies for minimising COVID19 cases and the design of effective antigenic formulations. Here, we examine the sensitivity of variants of concern (VOCs) representative of the B.1.617.1 and B.1.617.2 (first associated with infections in India) and B.1.351 (first associated with infection in South Africa) lineages of SARS-CoV-2 to neutralisation by sera from individuals vaccinated with the BNT162b2 (Pfizer/BioNTech) and ChAdOx1 (Oxford/AstraZeneca) vaccines. Across all vaccinated individuals, the spike glycoproteins from B.1.617.1 and B.1.617.2 conferred reductions in neutralisation of 4.31 and 5.11-fold respectively. The reduction seen with the B.1.617.2 lineage approached that conferred by the glycoprotein from B.1.351 (South African) variant (6.29-fold reduction) that is known to be associated with reduced vaccine efficacy. Neutralising antibody titres elicited by vaccination with two doses of BNT162b2 were significantly higher than those elicited by vaccination with two doses of ChAdOx1. Fold decreases in the magnitude of neutralisation titre following two doses of BNT162b2, conferred reductions in titre of 7.77, 11.30 and 9.56-fold respectively to B.1.617.1, B.1.617.2 and B.1.351 pseudoviruses, the reduction in neutralisation of the delta variant B.1.617.2 surpassing that of B.1.1351. Fold changes in those vaccinated with two doses of ChAdOx1 were 0.69, 4.01 and 1.48 respectively. The accumulation of mutations in these VOCs, and others, demonstrate the quantifiable risk of antigenic drift and subsequent reduction in vaccine efficacy. Accordingly, booster vaccines based on updated variants are likely to be required over time to prevent productive infection. This study also suggests that two dose regimes of vaccine are required for maximal BNT162b2 and ChAdOx1-induced immunity.

In vitro evolution of Remdesivir resistance reveals genome plasticity of SARS-CoV-2 (preprint)

Remdesivir (RDV) is used widely for COVID-19 patients despite varying results in recent clinical trials. Here, we show how serially passaging SARS-CoV-2 in vitro in the presence of RDV selected for drug-resistant viral populations. We determined that the E802D mutation in the RNA-dependent RNA polymerase was sufficient to confer decreased RDV sensitivity without affecting viral fitness. Analysis of more than 200,000 sequences of globally circulating SARS-CoV-2 variants show no evidence of widespread transmission of RDV-resistant mutants. Surprisingly, we also observed changes in the Spike (i.e., H69 E484, N501, H655) corresponding to mutations identified in emerging SARS-CoV-2 variants indicating that they can arise in vitro in the absence of immune selection. This study illustrates SARS-CoV-2 genome plasticity and offers new perspectives on surveillance of viral variants.