SARS-CoV-2 Exposure in Norway Rats (Rattus norvegicus) from New York City

Abstract

Millions of Norway rats (Rattus norvegicus) inhabit New York City (NYC), presenting the potential for transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from humans to rats. We evaluated SARS-CoV-2 exposure among 79 rats captured from NYC during the fall of 2021. Our results showed that 13 of the 79 rats (16.5%) tested IgG- or IgM-positive, and partial SARS-CoV-2 genomes were recovered from all 4 rats that were qRT-PCR (reverse transcription-quantitative PCR)-positive. Genomic analyses suggest these viruses were associated with genetic lineage B, which was predominant in NYC in the spring of 2020 during the early pandemic period. To further investigate rat susceptibility to SARS-CoV-2 variants, we conducted a virus challenge study and showed that Alpha, Delta, and Omicron variants can cause infections in wild-type Sprague Dawley (SD) rats, including high replication levels in the upper and lower respiratory tracts and induction of both innate and adaptive immune responses. Additionally, the Delta variant resulted in the highest infectivity. In summary, our results indicate that rats are susceptible to infection with Alpha, Delta, and Omicron variants, and wild Norway rats in the NYC municipal sewer systems have been exposed to SARS-CoV-2. Our findings highlight the need for further monitoring of SARS-CoV-2 in urban rat populations and for evaluating the potential risk of secondary zoonotic transmission from these rat populations back to humans. IMPORTANCE The host tropism expansion of SARS-CoV-2 raises concern for the potential risk of reverse-zoonotic transmission of emerging variants into rodent species, including wild rat species. In this study, we present both genetic and serological evidence for SARS-CoV-2 exposure to the New York City wild rat population, and these viruses may be linked to the viruses that were circulating during the early stages of the pandemic. We also demonstrated that rats are susceptible to additional variants (i.e., Alpha, Delta, and Omicron) that have been predominant in humans and that susceptibility to infection varies by variant. Our findings highlight the reverse zoonosis of SARS-CoV-2 to urban rats and the need for further monitoring of SARS-CoV-2 in rat populations for potential secondary zoonotic transmission to humans.

Keywords: Delta; Norway rats; Omicron; Rattus norvegicus; SARS-CoV-2; brown rats; rat COVID-19; rat coronavirus; reverse zoonosis; surveillance; wildlife.

FIG 1

Genomic sequences of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wild rats. (A) SARS-CoV-2 genomes found in rats compared with reference wild-type virus and variants of concern. Analyses were performed and visualized using https://clades.nextstrain.org. Reference sequences were downloaded from GISAID. (B) Phylogenetic tree of rat SARS-CoV-2 sequences with reference sequences from wild-type viruses and variants of concern. Branches with a posterior probability of ≥0.7 are labeled. (C) Distance matrices of regions covered by each rat-derived SARS-CoV-2 genome. Lineage B is represented by hCoV-19/Wuhan/IME-WH05/2019|EPI_ISL_529217|2019-12-30, Delta by hCoV-19/USA/NY-Wadsworth-21052117-01/2021|EPI_ISL_2278740|2021-05-01, Omicron by hCoV-19/USA/NY-NYULH1825/2021|EPI_ISL_2427410|2021-05-11, Alpha by hCoV-19/USA/NY-Wadsworth-21070773-01/2021|EPI_ISL_2868594|2021-05-31, and Beta by hCoV-19/USA/NY-PRL-2021_1008_02C08/2021|EPI_ISL_5285364|2021-10-03.

FIG 2

Sprague Dawley (SD) rats are susceptible to infection with Alpha, Delta, and Omicron variants. (A) Prevalence of Alpha, Delta, and Omicron variants in NYC. Figure adapted from https://outbreak.info. (B) Scheme of the virus challenge experiment using 6-week-old SD rats. (C) Amino acid changes of Alpha, Delta, and Omicron variants across the receptor binding domain (RBD) compared to Wuhan-Hu-1 (NCBI access no. MN908947.3). (D) Body weight of rats mock-infected or infected with either Alpha, Delta, or Omicron variant. (E and F) Viral RNA copies (E) and infectious viral titers (F) in the turbinate and lungs from rats infected with either Alpha, Delta, or Omicron variant at 2 or 4 days postinfection (dpi). *, P < 0.05; **, P < 0.01. (G) Detection of SARS-CoV-2 nucleocapsid protein in bronchial epithelial cells by immunohistochemistry at 2 and 4 dpi. Scale bar = 100 μm.

FIG 3

Innate and adapted immune responses induced by SARS-CoV-2 infection in SD rats. (A) Pro-inflammatory cytokine/chemokine expression in lungs from rats infected with either Alpha, Delta, or Omicron variant. Asterisks indicate significant differences between the indicated variant and Delta. *, P < 0.05; **, P < 0.01. (B) Wuhan-Hu-1 spike protein- or RBD-specific IgG or IgM antibody titers. Absorbance at 450 nm (A₄₅₀)/cutoff was interpreted as negative if ≤1.0 and seropositive if >1.0. The cutoff value was 3-fold of the mean of negative serum samples. (C) Alpha-, Delta-, or Omicron-specific neutralizing antibody titers induced in mock-infected rats or rats infected with either Alpha, Delta, or Omicron variant.

FIG 4

Interactions between the RBD of SARS-CoV-2 variants Alpha, Delta, and Omicron and rat ACE2. (A) Rat ACE2 in complex with RBD. The three major contact sites in box-1, box-2, and box-3 are shown in panels B, C, and D, respectively. Interactions mediated by Alpha, Delta, and Omicron RBDs are compared side-by-side. Black thick outlines highlight favorable interactions. (E) List of ACE2 amino acid variations between rat, mouse, and human at the RBD interface. Many RBD mutations in Alpha, Delta, and Omicron variants are located near host-specfic ACE2 residues, indicated by black arrows.