A webinar sponsored by Thermo Fisher Scientific
Assay design and variants of concern
In a recent webinar on public health and emerging SARS-CoV-2 variants, Alan McNally, Ph.D., a professor in microbial genomics at the University of Birmingham, Emmanuel André, M.D., Ph.D., co-director of the Infectious Disease Laboratory at the University Hospital of Leuven, and Manoj Gandhi, M.D., Ph.D., Thermo Fisher’s senior director of medical affairs, Genetic Sciences, discuss assay design for PCR-based SARS-CoV-2 testing with an emphasis on the role these assays have played in epidemiological surveillance and the detection of new variants of concern. The conversation centered around molecular multi-target assay design, the importance of early recognition of new variants of concern, and the convergent evolution of SARS-CoV-2 following selective pressures applied to the virus.
Given the potential for variants of concern to enhance transmission, influence disease severity, and possibly evade natural or vaccine-induced immune responses, identifying cases of coronavirus disease (COVID-19) caused by variants of concern and monitoring their prevalence is critical.
Many laboratories are now investigating how to quickly test for variants of concern and include as many SARS-CoV-2 positive cases as possible in the variant testing. The Applied Biosystems™ TaqMan® SARS-CoV-2 Mutation Panel is a customizable menu of verified real-time PCR assays designed to detect mutations of concern that characterize the SARS-CoV-2 variants posing a high risk to the public health. The Applied Biosystems™ TaqMan® SARS-CoV-2 Mutation Panel allows you to build your own custom (targeted) panel to determine the prevalence of variants of concern or interest in the community you serve. This scalable follow-up testing solution lets you run a few or hundreds of samples to monitor for key SARS-CoV-2 mutations—all on your current real-time PCR instrumentation. The panel is regularly updated with assays targeting novel emerging SARS-CoV-2 variants and offers flexibility as assays can easily be added or removed from the custom panels depending on the current epidemiological situation.
Convergent evolution of SARS-CoV-2
All replicating organisms acquire mutations due to errors during the replication process, and these mutations accumulate over time enabling the organisms to develop novel properties and evolve in a divergent manner from their ancestors. The course of the evolution is set by the selective pressures of the changing environment that favor the survival of those organisms that carry mutations enabling them to thrive under the existing conditions. Convergent evolution has also been observed in nature and represents the phenomenon in which organisms develop similar functional properties, even though they have evolved independently and from a different lineage. Some of the most common examples of convergent evolution are the development of echolocation by both bats and whales, even though the two animal groups are clearly evolutionary distant, or the wing structures in insects, birds and mammals.
The uncontrolled spread of SARS-CoV-2 virus at the beginning of the COVID-19 pandemic has enabled the virus to evolve in a divergent manner. Several lineages were defined, carrying different mutations in the SARS-CoV-2 genome. However, over the course of the pandemic, the introduction of the disease control measures such as mask wearing or social distancing, has acted as an environmental pressure which led to the selection of SARS-CoV-2 lineages with mutations that increase the infectivity and transmissibility of the virus. When the genomes of SARS-CoV-2 variants were analyzed a striking observation was that the same mutations were found in independently emerging lineages. One example is the N501Y mutation in the S gene of SARS-CoV-2 encoding the spike protein which increases the affinity of the virus for the ACE2 receptor on human cells, thus rendering the virus more transmissible. This mutation was found in the three variants of concern: the Alpha (B.1.1.7, UK), Beta (B.1.351, South African) and Gamma (P.1, Brazilian) which evolved independently in different parts of the world at a similar timepoint in 2020. As the pandemic continued and many people developed immunity against SARS-CoV-2 due to an infection or vaccination, the variants with mutations conferring immune escape, such as E484K/Q or K417N/T started to be detectable in emerging variants. This particular feature of the SARS-CoV-2 virus, to acquire same mutations in different variants of concern, was utilized to design the RT-PCR based approach for surveillance testing. In each variant of concern these key mutations are present in different combinations with other mutations and by genotyping these particular positions one is able to discriminate between the different circulating variants. The fact that single assays are used offers a particular flexibility as one can easily adjust the selected panel of mutations by adding or removing assays, making this surveillance approach effective even as the virus continues to mutate.
TaqMan Mutation Panel
Introducing the Applied Biosystems TaqMan SARS-CoV-2 Mutation Panel, a customizable solution of 30+ verified real-time PCR assays, enabling detection of all currently known variants of concern or interest. Build your own panel for identification of one or many SARS-CoV-2 mutations—all on your current real-time PCR instrumentation.
In Dr. André’s laboratory, a selection of TaqMan genotyping assays enables correct detection and identification of SARS-CoV-2 variants of concern, including Alpha (501Y.V1), Beta (501Y.V2) and Gamma (501Y.V3). Designing tests for a pathogen that is still evolving is a challenge, but by focusing on mutations that have emerged convergently in multiple strains and are likely to emerge again (N501Y, E484K, K417N/T), the TaqMan SARS-CoV-2 Mutation Panel increases our chances to detect upcoming emerging variants of concern. Compared to WGS, PCR-based surveillance testing has several advantages including cost reduction, quick turnaround time, and an increase in scalability as higher proportion of SARS-CoV-2 positive cases can be analyzed. These are features that can make a significant difference in the response of public health institutions aimed at limiting the viral spread and ending the ongoing COVID-19 pandemic.
- TaqMan SARS-CoV-2 Mutation Panel
- Emerging SARS-CoV-2 Mutations and Variants
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