Modeling SARS-CoV-2 propagation using rat coronavirus-associated shedding and transmission

Abstract

At present, global immunity to SARS-CoV-2 resides within a heterogeneous combination of susceptible, naturally infected and vaccinated individuals. The extent to which viral shedding and transmission occurs on re-exposure to SARS-CoV-2 is an important determinant of the rate at which COVID-19 achieves endemic stability. We used Sialodacryoadenitis Virus (SDAV) in rats to model the extent to which immune protection afforded by prior natural infection via high risk (inoculation; direct contact) or low risk (fomite) exposure, or by vaccination, influenced viral shedding and transmission on re-exposure. On initial infection, we confirmed that amount, duration and consistency of viral shedding, and seroconversion rates were correlated with exposure risk. Animals were reinfected after 3.7-5.5 months using the same exposure paradigm. 59% of seropositive animals shed virus, although at lower amounts. Previously exposed seropositive reinfected animals were able to transmit virus to 25% of naive recipient rats after 24-hour exposure by direct contact. Rats vaccinated intranasally with a related virus (Parker's Rat Coronavirus) were able to transmit SDAV to only 4.7% of naive animals after a 7-day direct contact exposure, despite comparable viral shedding. Cycle threshold values associated with transmission in both groups ranged from 29-36 cycles. Observed shedding was not a prerequisite for transmission. Results indicate that low-level shedding in both naturally infected and vaccinated seropositive animals can propagate infection in susceptible individuals. Extrapolated to COVID-19, our results suggest that continued propagation of SARS-CoV-2 by seropositive previously infected or vaccinated individuals is possible.

Fig 1. SDAV infection and re-infection paradigm.

A. Initial infection with SDAV. Naïve animals were inoculated intranasally with 2X10^e4 pfu SDAV. After 48 hours, inoculated animals or their dirty cages were used to expose naïve animals to the virus. For direct contact exposure, one inoculated animal was placed in a clean cage with naïve animals for 24 hours. For fomite exposure, naïve animals were placed in a dirty cage that had been inhabited by an inoculated rat for 24 hours. After 24 hours, all rats were relocated to new clean cages to constitute four groups: inoculated rats, direct exposure rats, and two fomite exposure groups–a fomite cohabitation group constituting 2–3 animals, and a fomite single group with only one animal. Oral swabs were taken on Day 2, 3, 4, 7 and 10 days post-exposure (dpe) on all animals, and serology performed 5–6 weeks later. A control group (not shown) was inoculated with DMEM alone, and similarly swabbed and bled. After initial exposure, animals assumed either seronegative or seropositive status. All groups were evenly split by sex. B. Reinfection with SDAV Naïve seronegative rats were inoculated intranasally with 2X10^e4 pfu SDAV to provide a source of infection. Rats that had originally received intranasal infection with SDAV were re-infected intranasally again with the same viral dose. Seropositive and seronegative animals from the initial infection experiment were randomly assigned direct contact, fomite contact-cohabitation, and fomite contact-singly housed contact groups for their second exposure. Time between initial and second exposure ranged from 113–165 days. Oral swabs were taken on Day 2, 3, 4, 7 and 10 dpe on all animals. Animals were sacrificed at 10 dpe and assessed for seroconversion. All groups were evenly split by sex.

Fig 2. Transmission of SDAV to naïve rats by SDAV-exposed or RCV-vaccinated rats.

A. Transmission of SDAV to naïve rats by previously SDAV infected rats. Inoculated rats that had received an initial dose of 2X10^e4 pfu SDAV, and had subsequently shed virus and seroconverted, received a second similar intranasal inoculation 112–140 days later (n = 13). After 48 hours, these animals were placed in a clean cage with susceptible recipient rats (n = 13; direct contact paradigm). After 24 hours, recipient rats were separated from the inoculated rat and placed in a new clean cage. Body weights and oral swabs were taken on both groups of animals at 2, 3, 4, 7 and 10 dpi. Animals were sacrificed at 10 dpe and assessed for seroconversion. All groups were evenly split by sex. B. Transmission of SDAV to naïve rats by RCV vaccinated rats. Eight-week-old naïve rats (n = 24) were inoculated intranasally with 1X10^e3 pfu RCV and assessed for seroconversion 4 weeks later. All animals seroconverted, and were divided into a single vaccine group (n = 12) and a double vaccine group (n = 12). This latter group received an additional RCV inoculation at the same dose. Six weeks after their last RCV exposure, seropositive animals were inoculated with 2X10^e4 pfu SDAV. After 48 hours, these animals were co-housed in a clean cage with a susceptible rat (n = 12 for single vaccine group; n = 9 for double vaccine group) for 7 days. Body weights and oral swabs were taken on both groups of animals at 2, 3, 4, 7 and 10 dpe. Animals were sacrificed at 10 dpe and assessed for seroconversion. All groups were evenly split by sex.