Coronavirus Sequencing

At a basic level, diagnostic testing helps clinicians manage patients, and infectious disease surveillance is required to manage populations.

Diagnostic testing provides important yes/no answers for individual patients so that appropriate management can be provided.

Surveillance helps public health officials track the path of the epidemic, understand transmission routes, perform contact tracing, determine the rate of viral evolution, and understand if the virus is changing in ways that could impact diagnostic or therapeutic effectiveness.

NGS can provide unbiased detection of a novel pathogen in patient samples without prior knowledge of the organism.

A key challenge in infectious disease detection is that many of the microbes, including viruses, that cause respiratory, digestive, and other diseases in humans, have not been researched and characterized and thus are not known or detected by targeted approaches such as PCR. Development of PCR assays requires knowledge of the pathogen genome. NGS plays a critical role in discovering these unknown, novel pathogens; the resulting genome sequence can then be used to develop routine tests such as PCR to help clinicians manage patients.

NGS can be used to track the evolution of the pathogen genome to help public health officials monitor the spread of infection and determine the best isolation plan at a population level. Sequencing the virus from different patients over time can determine the rate of viral evolution, and address whether the virus is changing in ways that could impact pathogenicity as well as diagnostic or therapeutic effectiveness. PCR is designed to detect the presence of specific regions of the pathogen genome and will not identify new mutations across rapidly evolving pathogen genomes. Furthermore, PCR performance can suffer if mutations occur in the primer or probe binding regions.

Epidemiologists can utilize NGS to study viral genome mutations from patient samples across the globe. They can use this information to build a genetic tree (or map) that can indicate the path of transmission between patients. Clusters due to genetic similarities in the pathogen belong to patients within the same transmission chains. These transmission chains allow public health officials to quickly determine the pathogen origin, track the path of the epidemic, understand transmission routes, and inform appropriate containment measures.

A shotgun metagenomics workflow enables sequencing of both novel and known species. During an outbreak involving an unknown pathogen, multiple molecular diagnostic tests are often utilized; this may lead to unnecessary costs and delays in identifying the pathogen. Shotgun metagenomics can be used as a single comprehensive screening assay for identifying and characterizing pathogens. This research workflow can help accelerate outbreak investigations and support development of new lab tests for large-scale screening efforts.

Once a pathogen such as SARS-CoV-2 is identified, a target enrichment workflow can provide the high sensitivity needed to detect the virus, and provide information about its epidemiology and evolution. This information can help researchers optimize infection control strategies, including monitoring when it's acceptable to de-escalate isolation mechanisms and resume normal activities, and aid in the development of vaccines.

These complementary workflows using Illumina sequencing can be performed alongside traditional testing methods and integrated into a comprehensive outbreak response model.

The Respiratory Virus Oligo Panel includes 7,800 probes to sequence common respiratory viruses, recent flu strains, SARS-CoV-2, and other coronaviruses, as well as human probes to act as positive controls. These probes are 80-mer oligos, spaced very close together, providing full genome coverage of all viruses in the panel. Table of viruses in the panel:

Target enrichment is a resequencing method that captures genomic regions of interest by hybridization to target-specific biotinylated probes. Target enrichment through hybrid–capture methods allows for highly sensitive detection and therefore does not require high read depth. Additionally, the target enrichment NGS workflow allows for near-complete sequence data of targets and opens up applications such as variant analysis for viral evolution or viral surveillance.

Alternatively, amplicon sequencing is designed to detect the presence of the target pathogen in a sample by identifying specific regions of the pathogen genome. This method does not enable identification of new mutations across rapidly evolving pathogen genomes (as is required for viral evolution or viral surveillance studies).

The target enrichment NGS workflow allows for near-complete sequence data of targets and opens up applications such as variant analysis for coronavirus evolution or viral surveillance studies. Compared to other targeted resequencing methods, such as amplicon sequencing, enrichment through hybrid capture allows for dramatically larger probe panels with more comprehensive profiling of the target regions. Additionally, the oligo probes used for hybrid–capture protocols remain effective even within highly mutagenic regions (which can be difficult for amplicon-based assays such as qPCR), allowing targeting of rapidly evolving viruses such as RNA viruses.

Once a pathogen like the SARS-CoV-2 coronavirus has been identified, amplicon sequencing can provide cost-effective, rapid, and scalable detection of the pathogen. When used as a general whole-genome sequencing diagnostic approach, it allows for broader target coverage, making it less susceptible to mutational effects. For research, viral whole-genome sequencing can be used to monitor viral mutations and allows phylogenetic analysis.

Once a pathogen like the SARS-CoV-2 coronavirus has been identified, a viral enrichment sequencing panel provides high sensitivity detection coupled with epidemiology information by detecting the full genome and the genomic mutations found across different samples. This information helps define the epidemiology of transmission and can assist public health officials in optimizing infection control strategies.

The Illumina Respiratory Virus Oligo Panel expands detection to ~30 families of respiratory viruses and allows researchers to study co-infections with other viruses in the panel.

The Illumina COVIDSeq Test (3076 samples) and COVIDSeq Assay (96 samples) are amplicon-based NGS products designed to detect RNA from the SARS-CoV-2 virus.

We recommend visiting our NGS for Beginners web section.

Visit our Sequencing Platforms page to explore our portfolio. The choice of sequencer depends on which method(s) you use most frequently. See the workflows above for recommendations on which sequencer is optimal for which method.