Animal reservoirs of Covid-19 may trigger new rounds of human diseases

Watch out for the dog (and the cat too)

New variants of SARS-CoV-2 lurk under the streets of New York. A recent paper by Smyth et al. extracts SARS-CoV-2 samples from fourteen city wastewater treatment plants. The team developed methods to detect mutations in a critical region of the genome, the receptor binding domain of the Spike protein.

As expected, they found a number of different variants of Covid-19 in their samples, including Alpha, Beta, Delta, and Gamma. They also found four distinct “cryptic” lineages, WNY1, 2, 3 and 4, from three of the sewage sites. These four cryptic variants give surprising and disturbing results. Receptor-binding domain RNA extracted from samples contained up to 29 mutations in all four variants detected, some previously seen in variants of concern or of interest and others unique to these samples. Figure 1 illustrates the overlap of mutations found in many other variations. Most of the variants unique to the sewer variants are extraordinarily rare among the 3.5 million sequences found in the GISAID SARS-CoV-2 database.

The mutations shown in Figure 1 all occur in the small region of the viral genome sampled by sewer detectives. This region specifies the receptor binding domain, the site of the virus binding to the ACE2 receptor on the surface of the host cell. The receptor binding domain is also the target of the vast majority of protective antibodies. Mutations within this region can profoundly affect transmission and immunity.

Most cryptic variant mutations occur between amino acid positions 437 and 508. This is the part of the Spike protein that is in direct contact with the ACE2 receptor.

Extensive mutations in the receptor binding domain are functional. The authors report that pseudotyped viruses carrying a receptor binding domain substituted by that of the cryptic variants are infectious. This is not entirely unexpected as the polymorphisms shared with variants of interest and concern enhance the function of the Spike protein. Amino acid substitutions shared at positions N440, L452 and N501 enhance receptor binding affinity. Mutations at positions K417, N439, K444, N460, E484, Q493 and S494 reduce viral neutralization by convalescent sera and selected monoclonal antibodies. Obviously, the amino acid substitutions at the twelve new sites are viable. This is particularly notable for the deletion at position 484, a site known to interact directly with ACE2 and neutralizing antibodies. It is important to determine the potential role of each of these cryptic mutations in immune evasion and improved function. They can anticipate changes yet to be seen in worrying new variants.

Black-smith et al. speculate that these variants do not come from human sources, but from non-human hosts. The speculation is based on several observations. First, cryptic variants are seen only in some samples, but not in all. The authors argue that if variants were prevalent in the human population, they would be found throughout the city and would not be confined to specific sewage sites. The second is that their observation of pseudotyped viruses carrying the “cryptic” receptor binding domain are capable of infecting non-human ACE2 receptors, particularly those of rats and mice.

Finally, we know that SARS-CoV-2 can infect many species other than pangolins and humans. Up to 40% of all dogs tested in the United States have antibodies to SARS-CoV-2. Several variants infect house mice and wild field mice. Up to 30% of all white-tailed deer in the northeast are positive for Covid-19 antibodies. There were reports of infections from domestic cats and large cats kept in zoos at the start of the pandemic.

SARS-CoV-2, like the influenza virus, can engage in zoonotic volleyball. Infections of animal origin can find their way into humans. Strains of SARS-CoV-2 infected mink, and mink returned the favor by infecting humans. As early as January 2021, mink farmed in Denmark showed signs of infection with SARS-CoV-2. These viruses have been transmitted to human populations and then sequenced. A to study by Bayarri Olmos et al. indicates that a mutation directly derived from zoonotic transmission from mink, Y453F in the receptor binding domain, results in up to four times higher affinity for the ACE2 receptor, suggesting a much more transmissible virus. In the future, we need to be aware of the new variants from the fauna that inhabit our ecosystem.

Black-smith et al. examined the resistance of pseudotyped viruses carrying the mutant receptor binding domain to monoclonal antibodies currently approved for treatment: etesevimab, bamlanivimab and imdevimab. They report that WNY1 and 2 are partially neutralized by etesevimab and imdevimab. The WNY3 and 4 variants are completely resistant to all the monoclonal antibodies tested. All four variants are also completely resistant to bamlanivimab. The researchers also tested these mutations against naturally occurring antibodies in convalescent antisera. Convalescent sera neutralized both WNY1 and 2 but were much less effective against WNY3 and 4. Neutralizing titers of sera from fully vaccinated individuals were also significantly reduced.

Black-smith et al. note that the pseudotyped virus used to test for neutralization of antibodies only includes mutations in the receptor binding domain, while the full-length variant is much more likely to decrease neutralization of antibodies to a greater extent. They conclude that “the characteristics of these variant lines give them the capacity to be an increased threat to human health”.

Examining this small region of the virus has been instructive. Cryptic variants display the full range of variations that can occur in SARS-CoV-2 RNA. The receptor binding domain here shows a greater degree of mutability. This embodies the potential for viral variation, not only in the receptor binding domain, but also throughout the rest of the Spike protein and viral genome, potentially improving virulence, vaccine and monoclonal antibody resistance, immune suppression, and the transmission.

As a final thought, it would be remiss not to add the possibility that the extreme variation observed in the highly localized cryptic variants of New York City is not due to zoonotic variants, but rather to the effluent of clusters of highly regional variants reproducing in human populations, possibly neighborhoods. treat immunocompromised patients. Extreme variation in S protein is documented in such patients. Either way, the final takeaway message is that the variants documented so far are just a subset of what we can expect from future SARS-CoV-2 infections.