Sarbecoviruses (a subgenus of Coronaviridae) are considered zoonotic and the subgenus of Sarbecoviruses specifically are gaining a notoriety of jumping from animals to humans. As viruses jump from host to host, evolutionary mutations in the viruses' nucleic acid are selected to improve viral infection and replication in the new host.
Since our awareness of Sarbecovirus in humans is a recent phenomenon, it is unclear if the virus is present in other species naturally. It is important to note that natural transmission from humans to most research animal species has not been reported and is considered unlikely.
In rapid response to the COVID-19 pandemic, our scientific staff has designed a SAR-CoV PCR assay and MFIA COVID-Plex serology (antibody) test. Charles River has developed both tests to identify if your research animals have been infected with SARS-CoV-2, the virus that causes COVID-19.
MFIA COVID-Plex Serology
CRL's serological COVID-Plex test is specific for detection of SARS-CoV2 (COVID-19) antibodies and qualified for surveillance (and research) purposes of laboratory animals. The COVID-Plex multiplex assay utilizes SARS-CoV2 (COVID-19) specific spike-full length (spike-FL) and nuclear proteins (NP). A sample is considered positive only if both COVID-19-specific spike-FL and NP assays are individually positive.
The animals with just COVID-19 spike-FL reaction (possibly due to cross reacting seasonal coronavirus strain antibodies) may not be used in studies involving SARS-CoV2 spike protein (i.e., vaccine studies).
Serology Sample Types
Label each individual HemaTIP™. To label, grasp the HemaTIP™ and wrap the label around the base, overlapping it on itself to secure in place. From LTM, you can use the "Labels-Order Samples Wrap-Around" report with Avery label, 5167/5267 (or equivalent office supply store brand, to print labels formatted for use with HemaTIPs.
Use your current blood sampling procedure.
Touch HemaTIP™ to blood until tip is completely saturated/red. Do not submerge in blood.
Place HemaTIP™ upright in clamshell case to dry for 1 hr at ambient temperature. For best results, leave the clamshell open while drying.
Serum Diluted (1:4)
Submit at least 200uL of serum diluted 1 part serum into 4 parts phosphate buffered saline (PBS) with antimicrobial agent (i.e., 40uL serum into 160uL of PBS with antimicrobial agent).
Sample should not be heat inactivated; please indicate if heat inactivation or another treatment has occurred.
Submit at least 40uL serum. Sample should not be heat inactivated; please indicate if heat inactivation or another treatment has occurred.
- Summary Report
Sarbecovirus PCR Assay
Based on a conserved genetic region with a low mutation rate, this assay will not only detect the circulating human and known animal strains, but also will likely detect novel strains in other species.
How to Submit PCR Samples
All sample collection materials (except nasal swabs) are provided free by request through our Laboratory Testing Management® program. For best sensitivity, animal-derived samples should be shipped for testing as soon as possible but can be held at 4◦ C for one week or -20◦ C for one month.
Samples collected directly from research animals considered to be a higher susceptibility risk for Sarbecoviruses (e.g., nonhuman primates, dogs, cats, swine, ferrets, bats) will be processed in our BSL-2 Laboratory.
Oral and Nasal Swabs
- When collecting oral and nasal swabs swab shafts should be removed before placing the swab tip into the tube to improve assay sensitivity.
- Swabs should be placed in a 5 mL tube and shipped at room temperature or on ice packs.
Bronchial Wash 200-500 uL total of bronchial should be submitted in a 2.0 mL microcentrifuge tube. Ship on ice packs or dry ice. Lung Tissue or Respiratory Tract Tissue 100-300 mg of tissue can submitted in a 2.0 mL microcentrifuge tube on ice packs or dry ice. Fecal Swabs or Pellets
- Small animal fecal pellets can be shipped at room temperature in a 2.0 mL microcentrifuge tube.
- Large animal samples must be a swab of fecal material shipped on ice packs and placed in a 5 mL conical tube.
Whole Blood 200-500 uL of whole blood can be submitted in a non-heparin anticoagulant in a non-glass vial. The window of detection is shorter than for a respiratory sample because the virus is first cleared from the blood by the humoral immune response. Research Biologics Cell cultures or tumor cell lines may be evaluated. Submit sample according to guidelines for CLEAR testing.
*Only samples for precautionary screening of laboratory animal may be submitted. Please do not submit: human samples, samples collected from research animals inoculated with or knowingly infected with Sarbecoviruses, or research reagents, cell lines, or other biologics that contain Sarbecoviruses.
Which research animals are at the highest risk of having a Sarbecovirus or acquiring from another host?
Some animal species may have virus receptors that are more susceptible to SARS viruses, but natural transmission from human to research animals has not been reported and is considered unlikely. These species include nonhuman primates, dogs, cats, swine, Peromyscus, ferrets, mink, otters, and bats. Research rodents are unlikely to be infected through natural transmission, although infection through high titer inoculation of mice has been demonstrated , but transmission to naïve cohorts have not. Transmission from experimentally inoculated hamsters, Peromyscus, ferrets, and cats to naïve like-species has been reported. Awareness of Sarbecoviruses is a recent phenomenon, it is unclear which other members of this subgenus may be present in other animal species.
Should routine testing of study animals be performed?
Routine testing of laboratory animals is not recommended. There have been no reports of natural infection in research animals. In addition, standard industry practices of good hand hygiene, use of site uniforms, and appropriate PPE (gloves, face masks, etc.) should be sufficient to minimize possible spread from asymptomatic staff to research animals.
When should clinical cases be tested for COVID-19?
In line with government and national organizations (CDC, USDA, AVMA), testing of animals is discouraged unless all other, more common causes of illness have been excluded. Further guidance on the decision to test animals is available from the CDC.
What are the recommended sample types for Sarbecovirus PCR testing?
Only samples for precautionary screening of laboratory animals may be submitted.
- Oral and nasal swabs
- Bronchial wash
- Lung tissue or respiratory tract tissue
- Fecal swab or pellets
- Whole blood
- Research biologics
Where is the highest concentration of Sarbecoviruses?
Sarbecoviruses are primarily respiratory viruses. The highest concentration of shedding the virus is expected to be from the naso/oral cavity.
What is the importance of designing a SARS-CoV assay beyond the ability to detect only human Sarbecoviruses?
Coronaviruses and the subgenus of Sarbecoviruses are gaining a notorious history of spreading viruses from animals to humans. Though the exact evolutionary transmission has not been determined, bats, pangolins, and civets are believed to play a role in transmission to humans. As viruses jump from host to host, evolutionary mutations in the viruses' nucleic acid are selected to improve receptor binding and replication within the new hosts intracellular machinery.
Single mutations in critical locations in a PCR primer or probe can greatly reduce the sensitivity of the assay. For this reason, Charles River targeted a highly conserved region among all known Sarbecovirus sequences where there has been limited evidence of mutation. The chosen target region serves an important function where single nucleotide base pair mutations could compromise virus replication. Should mutations occur in known circulating strains or unknown novel strains in other animal species, this assay will provide the best possibility for cross-reactivity.
Why are fecal swabs being used to detect Sarbecoviruses?
SARS-CoV has been detected in the defecate of humans and therefore may also be detected in the defecate of other animal species, although this is primarily a respiratory virus.
Should I be concerned about potential SARS-CoV-2 infection of study animals?
Our assessment is that the risk is very low. Coronaviruses are widespread in nature and most are limited in their host species range. As of April 6, 2020, the COVID-19 websites of the World Health Organization, the World Organisation for Animal Health and the Center for Disease Control and Prevention say there is no evidence of risk of infection from companion animals.
Will the Sarbecovirus PCR cross-react with rodent coronaviruses such as MHV?
In addition to computer target analysis of the PCR primer and probe targets, we have also demonstrated in the laboratory assay qualification that the Sarbecovirus PCR does not detect rodent coronaviruses.
What about common lab animals?
There are no reports of natural infection of SARS or MERS in laboratory species. Each of these models required experimental intranasal instillation of relatively large amounts of viral particles and showed limited clinical signs. While common research strains of mice could be infected with variants of SARS-CoV-2, the virus was eliminated rapidly and did not infect naïve cohorts. However, genetically engineered mice that were designed to have certain human cell characteristics, were useful models to study both SARS and MERS.
Work is just starting on animal models for SARS-CoV-2. Results to date indicate that SARS-CoV-2 does not infect common strains of mice. Several laboratories are just beginning to explore the potential to study the virus using the same SARS and MERS transgenic mouse models.
Several labs across the globe are reporting ongoing work with SARS-CoV-2 in nonhuman primates.
How does Charles River keep study animals safe?
Maintaining our animals' health starts with selection of quality vendors and the fact that we only purchase animals that are purpose-bred for biomedical research. Charles River does not use wild or feral animals. Nearly all rodents come from commercial barrier-maintained colonies with stringent health standards, the vast majority are from other CRL sites. The rare non-commercial rodents used must meet the same health standards. All large animals are sourced from vendors that are audited and approved by CRL veterinarians.
Rodents and rabbits are transported to our sites in biosecure crates in dedicated climate-controlled vehicles. Crates are disinfected prior to housing the animals in their study room. All animals are checked by trained staff for any problems and if found, are reported to veterinary staff.
Can study animals get the virus from a sick employee?
The risk is very low. Our site policies are that people who are ill must remain home, particularly during cold and flu season. Our PPE standards are designed to prevent disease transmission between animals and humans.
In addition, experts have not expressed concern about transmission to or from animals. Multiple international and domestic health organizations have indicated that pets and other domestic animals are not considered at risk for contracting or spreading SARS-CoV-2.
What resources are available to learn more information?
Veterinary Medical Association From ferrets to mice and marmosets, labs scramble to find right animals for coronavirus studies – STAT, March 6, 2020 Animal models for SARS and MERS coronaviruses, Current Opinion in Virology, August, 2015