Skip to content

Basket

You currently have no items in your basket.

Could mutations of SARS-CoV-2 suppress diagnostic detection?

Rockland Immunochemicals scientists have been working hard to support COVID-19 research efforts with expertise and the development of SARS-CoV antibodies.

The recent emergence of SARS-CoV-2 strains H69/V701,2, D796H3 and D614G4 in the United Kingdom and the N501Y strain in South Africa has prompted concerns as to their susceptibility to vaccine neutralization. I argue here another concern deserves equal attention: whether such strains can evade detection by diagnostics and compromise our ability to accurately track disease.

SARS-CoV-2 is arguably one of the most intensely studied viruses since the advent of HIV. Genotyping of the virus is occurring on a global scale and enabling nearly ‘real-time’ acquisition of viral genetic composition5. The mutation rate of the virus is ~2 nucleotides (nt) per month, which is considerably less than that of influenza (4 nt/month) or HIV (8 nt/month)6. Mutations put at risk detection strategies that do not accommodate changes in the viral genome.

The observed mutations in SARS-CoV-2 are not predicted to affect the utility of currently deployed vaccines7; however, changes in the viral nucleic acid and protein sequences put at risk the utility of certain in vitro diagnostic assays if the mutation occurs in an area critical for primer or antibody binding in RT-PCR and immunoassays. In addition, a particular concern is antibody-based COVID-19 diagnostic tests that assess the presence and concentration of SARS-CoV-2 viral proteins in biofluids (mainly lysates from nasopharyngeal, oropharyngeal or saliva extracts). The most commonly deployed immunoassays for detection of SARS-CoV-2 viral proteins include enzyme-linked immunosorbent assays (ELISAs) and lateral flow assays (LFAs). The targeted analytes in these assays are predominantly spike (S) or nucleocapsid (N) proteins, the two most abundant and immunogenic viral proteins present in the SARS-CoV-2 genome.

S protein is a seductive viral antigen. It is highly immunogenic and contains sequences unique to SARS-CoV-28, thereby potentially minimizing cross reactivity to sequences present in other known human coronaviruses, such as SARS-CoV, Middle Eastern respiratory syndrome (MERS) virus and human coronaviruses 229E, OC43, HKU-1 and NL639. However, it comes with risks. S protein is the most likely viral protein to undergo mutation, especially mutations that may affect viral function, including infection rate10, transmissibility11,12 and the ability to infect individuals younger in age13 (for example, a mutation near the receptor binding domain may affect entry into the host cell). As mutations occur, immunoassays that detect S protein are more susceptible to an increasing rate of false-negative results, and it is essential to obtain sufficiently accurate testing results to detect the virus during the pandemic.

Conversely, point mutations in the N protein are less likely to occur and less likely to affect viral function. Thus, N protein is considered the best target for in vitro diagnostic detection and vaccine development for COVID-19 because of the conservation of the N protein sequence, the expanding knowledge of its genetics and biochemistry, and its strong immunogenicity14. The N protein, however, is also not invulnerable to mutation, and in vitro diagnostic and vaccine design must account for potential and inevitable mutations.

Regarding in vitro diagnostic immunoassays, an assay design that includes polyclonal antibodies has distinct advantages over assays that rely on the detection of a single epitope using a monoclonal antibody. A polyclonal antibody recognizing multiple epitopes present on or within the N protein is most likely to continue to detect the protein, despite the presence of multiple mutations in the target analyte. Where a mutation occurs within an epitope, a monoclonal antibody reactive to only that single epitope may become ineffective in detecting the viral protein. ‘Escape variant’ detection is among the several well documented benefits of polyclonal antibodies in applications where multiepitope binding properties represent clear advantages15.

In terms of the emerging SARS-CoV-2 strains — N501Y in South Africa9, H69/V701,2, D796H3 and D614G4 — none represent mutations that would hinder the ability of a diagnostic polyclonal antibodies to N protein to detect SARS-CoV-2. Even the strain B.1.1.7 (Fig. 1), which was identified to have 17 mutations, would be detected using such antibodies.

Figure 1: Multiple sequence alignments of SARS-CoV-2 S protein. Relative portions of the sequence alignments of S are shown. The mutated positions in variants are highlighted in yellow. Click HERE for full size image

With this in mind, and as new variants of SARS-CoV-2 are identified, it is critical that diagnostic tests for the virus in wide use are regularly reconfigured. In particular, diagnostic tests configured to use a single monoclonal antibody, especially those targeting S protein, must revalidate the performance of the test against emerging strains of SARS-CoV-2 or consider adapting the assay to the detection of N protein using high-affinity polyclonal antibodies as critical detection reagents.

References

  1. Kemp, S. A. et al. Recurrent emergence and transmission of a SARS-CoV-2 Spike deletion H69/V70. Preprint at bioRxiv https://doi.org/10.1101/2020.12.14.422555 (2020).
  2. Gallagher, J. New coronavirus variant: what do we know? BBC News https://www.bbc.com/news/health-55388846 (20 December 2020).
  3. Kemp, S. A. et. al. Neutralising antibodies in Spike mediated SARS-CoV-2 adaptation. Preprint at medRxiv https://doi.org/10.1101/2020.12.05.20241927 (2020).
  4. Plante, J. A. et al. Nature https://doi.org/10.1038/s41586-020-2895-3 (2020).
  5. Yin, C. Genomics 112, 3588–3596 (2020).
  6. Callaway, E. Nature 585, 174–177 (2020).
  7. Dearlove, B. et al. Proc. Natl Acad. Sci. USA 117, 23652–23662 (2020).
  8. Jaimes, J. A., André, N. M., Chappie, J. S., Millet, J. K. & Whittaker, G. R. J. Mol. Biol. 432, 3309–3325 (2020).
  9. McIntosh, K. & Perlman, S. Coronaviruses, including severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS). In Mandell, Douglas, and Bennett’s Principles and Practice of Infectious Diseases 9th edn (Bennett, J. E., Dolin, R. & Blaser, M. J., eds) 2072–2080 (Elsevier, 2020).
  10. Chen, J., Wang, R., Wang, M. & Wei, G. W. J. Mol. Biol. 432, 5212–5226 (2020).
  11. Public Health England. Investigation of novel SARS-CoV-2 variant, Variant of Concern 202012/01. Technical briefing 2, https://assets.publishing.service.gov.uk/government/uploads/system/uploads/attachment_data/file/949639/Technical_Briefing_VOC202012-2_Briefing_2_FINAL.pdf (28 December 2020).
  12. World Health Organization. SARS-CoV-2 variants. Disease Outbreak News https://www.who.int/csr/don/31-december-2020-sars-cov2-variants/en/ (31 December 2020).
  13. Volz, E. et al. Cell 184, 64–75.e11 (2021).
  14. Dutta, N. K., Mazumdar, K. & Gordy, J. T. J. Virol. 94, e00647–e20 (2020).
  15. Ascoli, C. A. & Aggeler, B. Biotechniques 65, 127–136 (2018).

 

We use cookies to give you the best experience of using this website. By continuing to use this site, you accept our use of cookies. Please read our Cookie Policy for more information.