A CNN model for predicting binding affinity changes between SARS-CoV-2 spike RBD variants and ACE2 homologues

The cellular entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) involves the association of its receptor binding domain (RBD) with human angiotensin converting enzyme 2 (hACE2) as the first crucial step. Efficient and reliable prediction of RBD-hACE2 binding affinity changes upon amino acid substitutions can be valuable for public health surveillance and monitoring potential spillover and adaptation into non-human species. Here, we introduce a convolutional neural network (CNN) model trained on protein sequence and structural features to predict experimental RBD-hACE2 binding affinities of 8,440 variants upon single and multiple amino acid substitutions in the RBD or ACE2. The model achieves a classification accuracy of 83.28% and a Pearson correlation coefficient of 0.85 between predicted and experimentally calculated binding affinities in five-fold cross-validation tests and predicts improved binding affinity for most circulating variants. We pro-actively used the CNN model to exhaustively screen for novel RBD variants with combinations of up to four single amino acid substitutions and suggested candidates with the highest improvements in RBD-ACE2 binding affinity for human and animal ACE2 receptors. We found that the binding affinity of RBD variants against animal ACE2s follows similar trends as those against human ACE2. White-tailed deer ACE2 binds to RBD almost as tightly as human ACE2 while cattle, pig, and chicken ACE2s bind weakly. The model allows testing whether adaptation of the virus for increased binding with other animals would cause concomitant increases in binding with hACE2 or decreased fitness due to adaptation to other hosts.

Detection of SARS-CoV-2 Omicron variant (B.1.1.529) infection of white-tailed deer

White-tailed deer (Odocoileus virginianus) are highly susceptible to infection by SARS-CoV-2, with multiple reports of widespread spillover of virus from humans to free-living deer. While the recently emerged SARS-CoV-2 B.1.1.529 Omicron variant of concern (VoC) has been shown to be notably more transmissible amongst humans, its ability to cause infection and spillover to non-human animals remains a challenge of concern. We found that 19 of the 131 (14.5%; 95% CI: 0.10-0.22) white-tailed deer opportunistically sampled on Staten Island, New York, between December 12, 2021, and January 31, 2022, were positive for SARS-CoV-2 specific serum antibodies using a surrogate virus neutralization assay, indicating prior exposure. The results also revealed strong evidence of age-dependence in antibody prevalence. A significantly ({chi}2, p < 0.001) greater proportion of yearling deer possessed neutralizing antibodies as compared with fawns (OR=12.7; 95% CI 4-37.5). Importantly, SARS-CoV-2 nucleic acid was detected in nasal swabs from seven of 68 (10.29%; 95% CI: 0.0-0.20) of the sampled deer, and whole-genome sequencing identified the SARS-CoV-2 Omicron VoC (B.1.1.529) is circulating amongst the white-tailed deer on Staten Island. Phylogenetic analyses revealed the deer Omicron sequences clustered closely with other, recently reported Omicron sequences recovered from infected humans in New York City and elsewhere, consistent with human to deer spillover. Interestingly, one individual deer was positive for viral RNA and had a high level of neutralizing antibodies, suggesting either rapid serological conversion during an ongoing infection or a "breakthrough" infection in a previously exposed animal. Together, our findings show that the SARS-CoV-2 B.1.1.529 Omicron VoC can infect white-tailed deer and highlights an urgent need for comprehensive surveillance of susceptible animal species to identify ecological transmission networks and better assess the potential risks of spillback to humans. Key FindingsThese studies provide strong evidence of infection of free-living white-tailed deer with the SARS-CoV-2 B.1.1.529 Omicron variant of concern on Staten Island, New York, and highlight an urgent need for investigations on human-to-animal-to-human spillovers/spillbacks as well as on better defining the expanding host-range of SARS-CoV-2 in non-human animals and the environment.

Multiple spillovers and onward transmission of SARS-Cov-2 in free-living and captive White-tailed deer (Odocoileus virginianus)

Many animal species are susceptible to SARS-CoV-2 and could potentially act as reservoirs, yet transmission of the virus in non-human free-living animals has not been documented. White-tailed deer (Odocoileus virginianus), the predominant cervid in North America, are susceptible to SARS-CoV-2 infection, and experimentally infected fawns can transmit the virus. To test the hypothesis that SARS-CoV-2 may be circulating in deer, we tested 283 retropharyngeal lymph node (RPLN) samples collected from 151 free-living and 132 captive deer in Iowa from April 2020 through December of 2020 for the presence of SARS-CoV-2 RNA. Ninety-four of the 283 deer (33.2%; 95% CI: 28, 38.9) samples were positive for SARS-CoV-2 RNA as assessed by RT-PCR. Notably, between November 23, 2020 and January 10, 2021, 80 of 97 (82.5%; 95% CI 73.7, 88.8) RPLN samples had detectable SARS-CoV-2 RNA by RT-PCR. Whole genome sequencing of the 94 positive RPLN samples identified 12 SARS-CoV-2 lineages, with B.1.2 (n = 51; 54.5%), and B.1.311 (n = 19; 20%) accounting for ~75% of all samples. The geographic distribution and nesting of clusters of deer and human lineages strongly suggest multiple zooanthroponotic spillover events and deer-to-deer transmission. The discovery of sylvatic and enzootic SARS-CoV-2 transmission in deer has important implications for the ecology and long-term persistence, as well as the potential for spillover to other animals and spillback into humans. These findings highlight an urgent need for a robust and proactive "One Health" approach to obtaining a better understanding of the ecology and evolution of SARS-CoV-2. One-Sentence SummarySARS-CoV-2 was detected in one-third of sampled white-tailed deer in Iowa between September 2020 and January of 2021 that likely resulted from multiple human-to-deer spillover and deer-to-deer transmission events.

Computational prediction of the effect of amino acid changes on the binding affinity between SARS-CoV-2 spike protein and the human ACE2 receptor

The association of the receptor binding domain (RBD) of SARS-CoV-2 viral spike with human angiotensin converting enzyme (hACE2) represents the first required step for viral entry. Amino acid changes in the RBD have been implicated with increased infectivity and potential for immune evasion. Reliably predicting the effect of amino acid changes in the ability of the RBD to interact more strongly with the hACE2 receptor can help assess the public health implications and the potential for spillover and adaptation into other animals. Here, we introduce a two-step framework that first relies on 48 independent 4-ns molecular dynamics (MD) trajectories of RBD-hACE2 variants to collect binding energy terms decomposed into Coulombic, covalent, van der Waals, lipophilic, generalized Born electrostatic solvation, hydrogen-bonding, {pi}-{pi} packing and self-contact correction terms. The second step implements a neural network to classify and quantitatively predict binding affinity using the decomposed energy terms as descriptors. The computational base achieves an accuracy of 82.2% in terms of correctly classifying single amino-acid substitution variants of the RBD as worsening or improving binding affinity for hACE2 and a correlation coefficient r of 0.69 between predicted and experimentally calculated binding affinities. Both metrics are calculated using a 5-fold cross validation test. Our method thus sets up a framework for effectively screening binding affinity change with unknown single and multiple amino-acid changes. This can be a very valuable tool to predict host adaptation and zoonotic spillover of current and future SARS-CoV-2 variants.

Recurrent mutations in SARS-CoV-2 genomes isolated from mink point to rapid host-adaptation

SARS-CoV-2, the agent of the COVID-19 pandemic, can infect a wide range of mammals. Since its spread in humans, secondary host jumps of SARS-CoV-2 from humans to a variety of domestic and wild populations of mammals have been documented. The evolution of SARS-CoV-2 in different host species is of fundamental interest while also providing indication of how SARS-CoV-2 may have adapted to human hosts soon after the initial host jump, a time window for which there are no genome sequences available. Moreover, the study of SARS-CoV-2 circulating in animals is critical to assess the risk that the transmission of animal-adapted viral lineages back into humans (i.e., spillback) may pose. Here, we compared the genomic landscapes of SARS-CoV-2 isolated from animal species relative to that in humans, profiling the mutational biases indicative of potentially different selective pressures in animals. We focused on viral genomes collected in infected mink (Neovison vison) and white-tailed deer (Odocoileus virginianus) for which reports of multiple independent spillover events and subsequent animal-to-animal transmission are available. We identified six candidate mutations for animal-specific adaptation in mink (NSP9_G37E, Spike_F486L, Spike_N501T, Spike_Y453F, ORF3a_T229I, ORF3a_L219V), and one in deer (NSP3a_L1035F), though these mutations appear to confer minimal advantage for circulation in humans. Additionally, circulation of SARS-CoV-2 in mink and deer has not caused considerable changes to the evolutionary trajectory of SARS-CoV-2 thus far. Finally, our results suggest that minimal adaptation was required for human-to-animal spillover and subsequent onward transmission in mink and deer, highlighting the generalist nature of SARS-CoV-2 as a pathogen of mammalian hosts.

Limited window for donation of convalescent plasma with high live-virus neutralizing antibodies for COVID-19 immunotherapy

The optimal timeframe for donating convalescent plasma to be used for COVID-19 immunotherapy is unknown. To address this important knowledge deficit, we determined in vitro live-virus neutralizing capacity and persistence of IgM and IgG antibody responses against the receptor-binding domain and S1 ectodomain of the SARS-CoV-2 spike glycoprotein in 540 convalescent plasma samples obtained from 175 COVID-19 plasma donors for up to 142 days post-symptom onset. Robust IgM, IgG, and viral neutralization responses to SARS-CoV-2 persist, in the aggregate, for at least 100 days post-symptom onset. However, a notable acceleration in decline in virus neutralization titers [≥]160, a value suitable for convalescent plasma therapy, was observed starting 60 days after first symptom onset. Together, these findings better define the optimal window for donating convalescent plasma useful for immunotherapy of COVID-19 patients and reveal important predictors of an ideal plasma donor, including age and COVID-19 disease severity score. One Sentence SummaryEvaluation of SARS-CoV-2 anti-spike protein IgM, IgG, and live-virus neutralizing titer profiles reveals that the optimal window for donating convalescent plasma for use in immunotherapy is within the first 60 days of symptom onset.