Characterization of Entry Pathways, Species-Specific Angiotensin-Converting Enzyme 2 Residues Determining Entry, and Antibody Neutralization Evasion of Omicron BA.1, BA.1.1, BA.2, and BA.3 Variants.

The SARS-CoV-2 Omicron variants were first detected in November 2021, and several Omicron lineages (BA.1, BA.2, BA.3, BA.4, and BA.5) have since rapidly emerged. Studies characterizing the mechanisms of Omicron variant infection and sensitivity to neutralizing antibodies induced upon vaccination are ongoing by several groups. In the present study, we used pseudoviruses to show that the transmembrane serine protease 2 (TMPRSS2) enhances infection of BA.1, BA.1.1, BA.2, and BA.3 Omicron variants to a lesser extent than ancestral D614G. We further show that Omicron variants have higher sensitivity to inhibition by soluble angiotensin-converting enzyme 2 (ACE2) and the endosomal inhibitor chloroquine compared to D614G. The Omicron variants also more efficiently used ACE2 receptors from 9 out of 10 animal species tested, and unlike the D614G variant, used mouse ACE2 due to the Q493R and Q498R spike substitutions. Finally, neutralization of the Omicron variants by antibodies induced by three doses of Pfizer/BNT162b2 mRNA vaccine was 7- to 8-fold less potent than the D614G. These results provide insights into the transmissibility and immune evasion capacity of the emerging Omicron variants to curb their ongoing spread. IMPORTANCE The ongoing emergence of SARS-CoV-2 Omicron variants with an extensive number of spike mutations poses a significant public health and zoonotic concern due to enhanced transmission fitness and escape from neutralizing antibodies. We studied three Omicron lineage variants (BA.1, BA.2, and BA.3) and found that transmembrane serine protease 2 has less influence on Omicron entry into cells than on D614G, and Omicron exhibits greater sensitivity to endosomal entry inhibition compared to D614G. In addition, Omicron displays more efficient usage of diverse animal species ACE2 receptors than D614G. Furthermore, due to Q493R/Q498R substitutions in spike, Omicron, but not D614G, can use the mouse ACE2 receptor. Finally, three doses of Pfizer/BNT162b2 mRNA vaccination elicit high neutralization titers against Omicron variants, although the neutralization titers are still 7- to 8-fold lower those that against D614G. These results may give insights into the transmissibility and immune evasion capacity of the emerging Omicron variants to curb their ongoing spread.

Cross-species tropism and antigenic landscapes of circulating SARS-CoV-2 variants.

Mutations in the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike receptor-binding domain (RBD) may alter viral host tropism and affect the activities of neutralizing antibodies. Here, we investigated 153 RBD mutants and 11 globally circulating variants of concern (VOCs) and variants of interest (VOIs) (including Omicron) for their antigenic changes and cross-species tropism in cells expressing 18 ACE2 orthologs. Several RBD mutations strengthened viral infectivity in cells expressing ACE2 orthologs of non-human animals, particularly those less susceptible to the ancestral strain. The mutations surrounding amino acids (aas) 439-448 and aa 484 are more likely to cause neutralization resistance. Strikingly, enhanced cross-species infection potential in the mouse and ferret, instead of the neutralization-escape scores of the mutations, account for the positive correlation with the cumulative prevalence of mutations in humans. These findings present insights for potential drivers of circulating SARS-CoV-2 variants and provide informative parameters for tracking and forecasting spreading mutations.

"Is Omicron mild"? Testing this narrative with the mutational landscape of its three lineages and response to existing vaccines and therapeutic antibodies.

SARS-CoV-2 Omicron with its lineages BA.1, BA.2, and BA.3 has triggered a fresh wave of Covid-19 infections. Though, Omicron has, so far, produced mild symptoms, its genome contains 60 mutations including 37 in the spike protein and 15 in the receptor-binding domain. Thirteen sites conserved in previous SARS-CoV-2 variants carry mutations in Omicron. Many mutations have shown evolution under positive selection. Omicron's giant mutational leap has raised concerns as there are signs of higher virus infectivity rate, pathogenesis, reinfection, and immune evasion. Preliminary studies have reported waning of immunity after two-dose primary vaccine regime, need for the boosters, folds reduction in vaccine effectiveness and neutralizing antibodies even after boosting and significant neutralization resistance with the therapeutic monoclonal, polyclonal, and convalescent antibodies against Omicron. The narrative that "Omicron is mild," therefore, needs time to be tested with a deeper, scientific dwelling into the facts.

SARS-CoV-2 Delta Variant Displays Moderate Resistance to Neutralizing Antibodies and Spike Protein Properties of Higher Soluble ACE2 Sensitivity, Enhanced Cleavage and Fusogenic Activity.

The SARS-CoV-2 B.1.617 lineage variants, Kappa (B.1.617.1) and Delta (B.1.617.2, AY) emerged during the second wave of infections in India, but the Delta variants have become dominant worldwide and continue to evolve. Here, we compared B.1.617 variants for neutralization resistance by convalescent sera, mRNA vaccine-elicited sera, and therapeutic neutralizing antibodies using a pseudovirus neutralization assay. B.1.617.1, B.1.617.2, and AY.1 pseudoviruses showed a modest 1.5- to 4.4-fold reduction in neutralization by convalescent sera and vaccine-elicited sera. In comparison, similar modest reductions were also observed for C.37, P.1, R.1, and B.1.526 pseudoviruses, but 7- and 16-fold reductions for vaccine-elicited and convalescent sera, respectively, were seen for B.1.351 pseudoviruses. Among twenty-three therapeutic antibodies tested, four antibodies showed either complete or partial loss of neutralization against B.1.617.2 pseudoviruses and six antibodies showed either complete or partial loss of neutralization against B.1.617.1 and AY.1 pseudoviruses. Our results indicate that the current mRNA-based vaccines will likely remain effective in protecting against B.1.617 variants. Finally, the P681R substitution confers efficient cleavage of B.1.617 variants' spike proteins and the spike of Delta variants exhibited greater sensitivity to soluble ACE2 neutralization, as well as fusogenic activity, which may contribute to enhanced spread of Delta variants.