While many technologies can be used to determine presence or absence of the SARS-CoV-2 virus in a sample, there is also an immediate need for epidemiological research to track viral transmission patterns, including identification of mutations within the viral genome. One key mutation is found in the SARS-CoV-2 spike protein gene, which influences cell entry and is a key target for vaccine development research. Identifying all variants in a SARS-CoV-2 genome in a given sample from a collection of samples can be accomplished by targeted next-generation sequencing (NGS), which enables sequencing of the complete SARS-CoV-2 viral genome and provides variant-level information that other molecular methods cannot.
Dr. Dirk Dittmer, Professor and Director of Virology and Global Oncology Programs at the UNC Lineberger Comprehensive Cancer Center and UNC Viral Genomics Core facility at UNC-Chapel Hill, is leveraging NGS for his research that focuses on the discovery of new viruses. Initially, his lab worked to discover viruses associated with cancers and most recently in respiratory infections, which enabled him to quickly pivot to SARS-CoV-2 research as the virus moved from urban travel centers into less densely populated areas in North Carolina.
In SARS-CoV-2 and infectious disease applications, molecular methodologies like qPCR and NGS are complementary tools and equally vital as each serves a unique purpose. Because qPCR focuses on detecting specific key targets, such as the spike protein for SARS-CoV-2, this technology is regularly used in healthcare settings to determine the presence of a viral pathogen.
On the other hand, Dr. Dittmer says NGS can provide more information of critical scientific value, as it enabling sequencing of the entire viral genome to identify critical viral genome variants that inform viral evolution, disease risk and severity, and host response. He explains that the idea of sequencing viruses is a well-established and proven method to track infections and person-to-person transmission patterns for other viral pathogens like influenza and HIV.
Dr. Dittmer applies the same principles for his studies in immunodeficiency and HIV/AIDS to his research of the SARS-CoV-2 virus, leveraging targeted NGS to identify super-spreaders and super-spreading events.
“We need to know if the virus spreads and evolves differently in different communities that differ by geography, race, ethnicity, preferred mode of transport and congregation. Sequencing an entire viral genome does not make any assumptions about which genes are important in any particular situation, allowing us to discover regions of the SARS-CoV-2 virus that no one has ever considered [and] gives us a footprint of evolutionary forces,” he says.
He hopes the insights gleaned from sequencing the entire viral genome will generate new insights about the biology of SARS-CoV-2 and novel drug targets that could inform vaccine development research.
Dr. Dittmer and his team have been studying a specific strain of the SARS-CoV-2 virus most commonly found in North Carolina since the beginning of the global crisis. This particular strain has a mutation in its spike protein called D614G that appears to be more contagious than the original strain from Asia. Since NGS enables analysis of specific viral strains and can be used to identify potential mutations that may occur in the spike protein, the data collected can be crucial for vaccine research. Any potential mutation in the spike protein will need to be taken into consideration during vaccine development research, or else it may render a vaccine candidate less effective as the virus continues to evolve.
For his research, Dr. Dittmer and his team use the highly automated Ion Torrent Genexus System, which makes NGS accessible to almost any lab. He describes it as “a closed system that takes care of even the bioinformatics for pre-defined assays” like the Ion AmpliSeq SARS-CoV-2 Research Panel. When used together, the Ion Torrent Genexus Integrated Sequencer and Ion AmpliSeq SARS-CoV-2 Research Panel enables automated sample-to-variant report NGS in under a single day.
“For SARS-CoV-2 and any other scenario that has the purpose of informing decision-making, any turnaround time greater than twenty-four hours is a non-starter,” he says.
In addition to rapid turnaround time, an added benefit of the Genexus System is sample flexibility, to help keep running costs low in his other research efforts.
“When we perform NGS in cancer clinical trials, the samples are collected over months and run together in batches. In this scenario, cost, precision and accuracy is more important than speed.”
The automation capabilities of Ion Torrent NGS allow Dr. Dittmer’s team to eliminate all hand pipetting steps for the SARS-CoV-2 assay, allowing them to “track every error which makes all the difference in reporting actionable information.”
Finally, Dr. Dittmer highlights the use of the Ion GeneStudio S5 series system as a complement to the Genexus System, primarily as a discovery and training platform as they develop workflows to study other infectious diseases. This scalable NGS system enables higher throughput and is flexible to run a broad range of infectious disease research applications, including host and immune response, microbiome analysis and transcriptome studies.
Contributions from researchers like Dr. Dittmer increase our understanding of the SARS-CoV-2 biology and transmission patterns, enabling more confidence in approaches to potential vaccine development and therapeutics to help minimize the impact of the virus.
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 McNamara, R.P., Caro-Vegas, C., Landis, J.T., Moorad, R., Pluta, L.J., Eason, A.B., Thompson, C., Bailey, A., Villamor, F.C.S., Lange, P.T., Wong, J.P., Seltzer, T., Seltzer, J., Zhou, Y., Vahrson, W., Juarez, A., Meyo, J.O., Calabre, T., Broussard, G., Rivera-Soto, R., Chappell, D.L., Baric, R.S., Damania, B., Miller, M.B., Dittmer, D.P., High-density amplicon sequencing identifies community spread and ongoing evolution of SARS-CoV-2 in the Southern United States, Cell Reports (2020), doi: https://doi.org/10.1016/j.celrep.2020.108352.