In vitro Characterization of Fitness and Convalescent Antibody Neutralization of SARS-CoV-2 Cluster 5 Variant Emerging in Mink at Danish Farms.

Ria Lassaunière,Jens Bukh,Jannik Fonager,Alonzo Alfaro-Núñez,Charlotta Polacek,Anders Frische,Louise Lohse,Anders Fomsgaard,Bitten Aagaard,Graham J Belsham,Signe Bollerup,Anette Bøtner,Anni Assing Winckelmann,Thomas Bruun Rasmussen,Susanne Gjørup Sækmose,Nina Weis,Morten Rasmussen,Kåre Mølbak,Alexander Underwood

doi:10.3389/fmicb.2021.698944

Abstract

In addition to humans, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can transmit to animals that include hamsters, cats, dogs, mink, ferrets, tigers, lions, cynomolgus macaques, rhesus macaques, and treeshrew. Among these, mink are particularly susceptible. Indeed, 10 countries in Europe and North America reported SARS-CoV-2 infection among mink on fur farms. In Denmark, SARS-CoV-2 spread rapidly among mink farms and spilled-over back into humans, acquiring mutations/deletions with unknown consequences for virulence and antigenicity. Here we describe a mink-associated SARS-CoV-2 variant (Cluster 5) characterized by 11 amino acid substitutions and four amino acid deletions relative to Wuhan-Hu-1. Temporal virus titration, together with genomic and subgenomic viral RNA quantitation, demonstrated a modest in vitro fitness attenuation of the Cluster 5 virus in the Vero-E6 cell line. Potential alterations in antigenicity conferred by amino acid changes in the spike protein that include three substitutions (Y453F, I692V, and M1229I) and a loss of two amino acid residues 69 and 70 (ΔH69/V70), were evaluated in a virus microneutralization assay. Compared to a reference strain, the Cluster 5 variant showed reduced neutralization in a proportion of convalescent human COVID-19 samples. The findings underscore the need for active surveillance SARS-CoV-2 infection and virus evolution in susceptible animal hosts.

Highlights

RESULTSSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in December 2019 in Wuhan, China
Accumulation of Amino Acid Changes in the Spike Protein of Mink-Associated SARS-CoV-2 In Denmark, SARS-CoV-2 variants co-circulating in mink and humans collectively acquired at least 35 different amino acid changes in the spike protein (Larsen et al, 2021)
Potential causes may include: (i) reduced efficiency of viral entry due to the spike mutations; (ii) modulated viral replication resulting from amino acid changes and/or deletions in ORF1a ( M85 and N2082) and ORF1b (P314L, T730I, and T2163I) that encode the viral replication and transcription complex (V’Kovski et al, 2021); or (iii) altered virus induced apoptosis mediated by ORF3a in the presence of the H182Y change (Ren et al, 2020)

Summary

RESULTS

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in December 2019 in Wuhan, China. In Denmark, SARS-CoV-2 variants co-circulating in mink and humans collectively acquired at least 35 different amino acid changes in the spike protein (Larsen et al, 2021). These SARSCoV-2 variants belong to lineage B.1.1.298. SARS-CoV-2 E gene genomic and subgenomic RNA measurements for the Cluster 5 virus were notably lower at 24 h post-inoculation compared to FIGURE 1 | The mink-associated mutations in the SARS-CoV-2 spike protein. Neutralization titres were initially evaluated against Cluster 5 variant relative to an early pandemic SARS-CoV-2 isolate (H1) (Figures 3A,B). One plasma sample had a greater than fourfold reduction, a threshold set for neutralization resistance (Li et al, 2020)

DISCUSSION

MATERIALS AND METHODS

Findings

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