Transmission of SARS-CoV-2 in free-ranging white-tailed deer in the United States

Abstract

SARS-CoV-2 is a zoonotic virus with documented bi-directional transmission between people and animals. Transmission of SARS-CoV-2 from humans to free-ranging white-tailed deer (Odocoileus virginianus) poses a unique public health risk due to the potential for reservoir establishment where variants may persist and evolve. We collected 8,830 respiratory samples from free-ranging white-tailed deer across Washington, D.C. and 26 states in the United States between November 2021 and April 2022. We obtained 391 sequences and identified 34 Pango lineages including the Alpha, Gamma, Delta, and Omicron variants. Evolutionary analyses showed these white-tailed deer viruses originated from at least 109 independent spillovers from humans, which resulted in 39 cases of subsequent local deer-to-deer transmission and three cases of potential spillover from white-tailed deer back to humans. Viruses repeatedly adapted to white-tailed deer with recurring amino acid substitutions across spike and other proteins. Overall, our findings suggest that multiple SARS-CoV-2 lineages were introduced, became enzootic, and co-circulated in white-tailed deer.

Fig. 1. Surveillance of SARS-CoV-2 viruses in free-ranging white-tailed deer in the United States (November 2021-April 2022).

a temporal distribution of SARS-CoV-2 quantitative reverse transcription polymerase chain reaction (qRT-PCR) positive nasal swab samples collected in white-tailed deer and those selected for whole genome sequencing (WGS); b geographic distribution of SARS-CoV-2 positive white-tailed deer samples with WGS; c SARS-COV-2 genetic variants in white-tailed deer nasal swab samples; d circulating SARS-CoV-2 Variants of Concern (VOCs) in humans in the United States (Jan 2021-October 2022) and those sampled in white-tailed deer from November 4, 2021 to April 4, 2022. Out of 383 SARS-CoV-2 WGS with complete metadata (Supplementary Data 1), 355 had identified Pango lineages, including 1 non-VOC variants (B.1) and 354 VOCs (70 Alpha, 9 Gamma, 273 Delta, 2 Omicron); 282 high-quality WGS were used further in evolutionary and transmission analyses. Arrow is used to highlight the small number in a particular week. Note that as d) focuses on the VOCs comparison between human and white-tailed deer, a single white-tailed deer sample, which belongs to a non-VOC variant (B.1), was collected on December 1, 2021 in Pennsylvania and was not shown in panel d. The source data for each subpanel is available in the Source Data file.

Fig. 2. Genetic association between SARS-CoV-2 viruses from humans and those from free-ranging white-tailed deer in the United States (November 2021-April 2022).

a The maximum clade credibility tree for white-tailed deer SARS-CoV-2 sequences (n = 282) and their potential precursor viruses in humans inferred 109 independent spillovers events of SARS-CoV-2 from humans (directly or indirectly) to white-tailed deer (Supplementary Data 2 and Supplementary Data 11). Three types of spillover events were identified: Human-Deer (green), where each event consists of at least one human precursor sequence and one white-tailed deer SARS-CoV-2 sequence; Human-Deer-Deer (blue), where each event consists of at least one human precursor sequence and at least two white-tailed deer SARS-CoV-2 sequences; Human-Deer-Human (red) where each event consists of at least one human precursor sequence, at least two white-tailed deer sequences, and an additional human SARS-CoV-2 sequence. The timescale of the phylogenetic tree was represented in units of years, and the scale bar indicates the divergence time in years. b The number of spillover events by variant of concern. c Geographic distribution of the Human-Deer, d Human-Deer-Deer, and e Human-Deer-Human events. The source data for subpanel b–e is available in the Source Data file.

Fig. 3. Repeated amino acid substitutions in the SARS-CoV-2 variants detected in wild white-tailed deer populations.

a # of repeated amino acid substitutions across SARS-CoV-2 proteins; b repeated amino acids substitutions under positive selection and their association with the independent transmission events shown in Fig. 2. Each row represents a transmission event, and a red box in each column represents an adaptive substitution observed in a specific event (row). Supplementary Data 2 and Supplementary Data 11 contain a list of the spillover events. For additional information on amino acid substitutions, please refer to Supplementary Data 3. The source data for each subpanel is available in the Source Data file.

Fig. 4. Visualization of repeated amino acid substitutions in the SARS-CoV-2 variants detected in wild white tailed deer populations.

a Papain-like proteinase (non-structural protein 3 [NSP3]; template with Protein Data Bank [PDB] accession #6wuu), b RNA-dependent RNA polymerase (NSP7, NSP8, and NSP12; template with PDB #6m71), and c spike protein (template with PDB #6vxx). The structures were visualized by PyMOL. Additional substitutions are listed in Supplementary Data 3. The source data for the bar figure in subpanel a is available in the Source Data file.

Fig. 5. Transmission of SARS-CoV-2 in free-ranging white-tailed deer in the state of New York (NY).

a County specific positive rate by quantitative reverse transcription polymerase chain reaction (qRT-PCR) or surrogate virus neutralization test (sVNT) in NY, and only those counties with at least four samples were included; b the maximum clade credibility tree of white-tailed deer SARS-CoV-2 viruses in NY with six transmission events with at least four deer sequences detected; c transmission events of SARS-CoV-2 viruses in NY analyzed by Bayesian Stochastic Search Variable Selection method; d the maximum clade credibility trees of white-tailed deer SARS-CoV-2 viruses associated with cross-county transmission events (case #1-4, in orange) and within-county (case # 5-6 in pink) detected by using phylogeographic analyses. The Bayes factors for these across-county transmission events are listed in Supplementary Data 5. The nodes in red were SARS-CoV-2 sequences from white-tailed deer, and those in cyan from human. The source data for panel a is available in Supplementary Data 4 and the Source Data file.

Fig. 6. A potential transmission event of white-tailed deer-adapted SARS-CoV-2 from white-tailed deer to humans.

a The maximum clade credibility tree illustrating the genetic relation among the white-tailed deer SARS-CoV-2 sequences and those in humans, and the human virus potentially originated from white-tailed deer is boxed; the presence of white-tailed deer-specific repeatedly adaptive amino acid substitutions were labeled in red for each node in the tree. The nodes in red were SARS-CoV-2 sequences from white-tailed deer, those in green from humans, and those in blue from zoo lions. The timescale of the phylogenetic tree was represented in units of years, and the scale bar indicates the divergence time in years. b frequency of the two white-tailed deer-specific repeatedly adaptive substitutions in the white-tailed deer or human SARS-CoV-2 sequences in public databases. ORF open reading frame, S spike protein, and NSP non-structural protein. The source data for subpanel b is available in the Source Data file.

Fig. 7. A potential transmission event of white-tailed deer-adapted AY.103 SARS-CoV-2 from white-tailed deer to humans in North Carolina (NC).

a The maximum clade credibility tree illustrating the genetic relation among the SARS-CoV-2 sequences from white-tailed deer and those from humans, and the human SARS-CoV-2 sequence potentially originated from white-tailed deer is boxed; the genomic diversity (comparing to Wuhan-Hu-1/2019) and the presence of two white-tailed deer-specific repeated amino acid substitutions as well as one event specific amino acid substitution (divide by blue line) was labeled in red for each node in the tree. The timescale of the phylogenetic tree was represented in units of years, and the scale bar indicates the divergence time in years. b frequency of the three white-tailed deer-specific substitutions in the white-tailed deer or human SARS-CoV-2 sequences. The nodes in red were SARS-CoV-2 sequences from white-tailed deer, those in green from humans, and those in blue from zoo lions. We analyzed for inter-host single nucleotide variations (iSNVs) in the 12 white-tailed deer sequences and defined a minor iSNV as having at least 5% but less than 50% prevalence among the reads. Among these 12 samples, we identified a single iSNV in 22-002350-031_NC_Randolph_2021-11-20_AY.103, which was ORF1b:V12271L with 461 reads coding for amino acid L (39.6%) and 703 reads coding for V (60.4%) at this position. The source data for subpanel b is available in the Source Data file.

Fig. 8. A potential transmission event of white-tailed deer-adapted AY.44 SARS-CoV-2 from white-tailed deer to humans in North Carolina (NC).

The maximum clade credibility tree illustrating the genetic relation among the white-tailed deer SARS-CoV-2 sequences and those in humans, and the human virus potentially originated from white-tailed deer is boxed; the genomic diversity (comparing to Wuhan-Hu-1/2019) and the presence of 3 white-tailed deer-specific repeated amino acid substitutions (star marks the one under positive selection) was labeled in red for each node in the tree. No human SARS-CoV-2 viruses (except for the one listed in the box) from public databases contain the combination of these three white-tailed deer specific repeated amino acid substitutions. The nodes in red were SARS-CoV-2 sequences from white-tailed deer, and those in cyan from human. The timescale of the phylogenetic tree was represented in units of years, and the scale bar indicates the divergence time in years.

Fig. 9. A potential transmission event of white-tailed deer-adapted AY.119 SARS-CoV-2 from white-tailed deer to humans in Massachusetts (MA).

Three white-tailed deer specific polymorphisms was identified in the human SARS-CoV-2 viruses in box. No human SARS-CoV-2 viruses (except for the one listed in the box) from public databases contain the combination of these three polymorphisms. The frequency of each of three individual polymorphisms in other human SARS-CoV-2 viruses are listed in Supplementary Data 7. The nodes in red were SARS-CoV-2 sequences from white-tailed deer, and those in cyan from human. The timescale of the phylogenetic tree was represented in units of years, and the scale bar indicates the divergence time in years.

Fig. 10. The maximum clade credibility tree illustrating the evolutionary relation between the white-tailed deer SARS-CoV-2 sequences collected from this study and those from public database.

A total of 332 white-tailed deer SARS-CoV-2 genomes were downloaded from GISAID (Global Initiative on Sharing All Influenza Data) on December 5, 2022. Among these genomes, 118 had complete and high coverage sequences (>99% coverage) and were included in the evolutionary analyses along with the white-tailed deer SARS-CoV-2 samples collected in this study. The timescale of the phylogenetic tree was represented in units of years, and the scale bar indicates the divergence time in years.