Investigation of Severe Acute Respiratory Syndrome Coronavirus 2 Infection in Rats

Abstract

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with its ability to infect multiple animal species, has become a significant concern, especially with the emergence of variants carrying the mutation N501Y in the spike protein, which increases the virus's affinity for the ACE2 receptor in both mice and humans, thereby enhancing viral infectivity and facilitating adaptation to these hosts. As commensal rodents, rats often live close to humans in high-density populations. The striking similarity of 90% between the amino acids in Mus musculus ACE2 (mACE2) and those in Rattus norvegicus suggests a potential for SARS-CoV-2 to infect and spread among rat populations. The work described in this thesis aims to investigate the ability of SARS-CoV-2 and its variants of concern (VOCs) to infect rats and to be transmitted between them using molecular and histopathological approaches. This thesis initially investigated whether wild-type Wistar rats, as animal models, could be infected with multiple SARS-CoV-2 strains, including the original SARS-CoV-2 Liverpool strain (hCoV-2/human/Liverpool/REMRQ0001/2020), the B.1.1.7 and B.1.351 variants. The results demonstrated the ability of these three strains to infect rats and spread to various tissues, including the lungs, nasal passages, intestines, hearts, and brains. Additionally, data from Oxford Nanopore sequencing revealed distinct mutations in the viral RNA genome during the infection experiment, providing insight into viral evolution in rats, a potential host species. After confirming the susceptibility of rats to SARS-CoV-2 infection, the ability of rats to transmit B.1.1.7 and B.1.351 variants to naïve (non-infected) rats, even in the absence of direct contact, was confirmed by using a partitioned cage that only allowed the airflow to pass through two parts of the cage: the infected and non-infected parts. Lung tissues obtained from non-infected rats exhibited the presence of SARS-CoV-2 nucleocapsid protein (NP) associated with bronchus-associated lymphoid tissue (BALT) formation. As the expression of hACE2 enhances the susceptibility of animals to SARS-CoV-2, transgenic hACE2 Sprague Dawley (SD) rats were utilised and infected with the Omicron BA.5 sub-lineage. Results obtained from this experiment showed a significant decrease in viral load and viral transcription levels from day 1 to day 5 post-infection. Moreover, the reduction in NP expression and BALT formation in day 7 lung tissues compared with day 3 tissues supports the viral load results and indicates the immune response activity of hACE2 rats against viral infection. Nasal tissues exhibited a higher viral level than lung tissues, indicating that the upper respiratory system is the main target for the BA.5 sub-lineage. The results discussed in this thesis provide insight into SARS-CoV-2 infection and transmission between rats as animal models, enhancing the understanding of virus-host interactions and zoonotic spillover concerns. Exploring and monitoring SARS-CoV-2 infections in rat populations should be continued to assess the potential for emerging new mutations that could increase zoonotic spillover risks.