Coronavirus Disease 2019 (COVID-19) - Whole Genome Sequencing for Surveillance
Consistent with O. Reg. 671/92 of the French Language Services Act, laboratory testing information on this page is only available in English because it is scientific or technical in nature and is for use only by qualified health care providers and not by members of the public.
Background
This page provides information on the whole genome sequencing (WGS) that is performed at Public Health Ontario (PHO) for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19 disease. SARS-CoV-2 WGS is performed for surveillance purposes only and cannot be used in place of standard molecular diagnostic testing. For molecular diagnostic testing, refer to this PHO webpage - PCR : Coronavirus Disease 2019 (COVID-19).
The information collected from SARS-CoV-2 WGS is summarized in a monthly report: SARS-CoV-2 Whole Genome Sequencing in Ontario.
Updates:
Public Health Ontario no longer performs PCR screening for COVID-19 Variants of Concern (VOCs).
This change reflects:
- A transition to whole genome sequencing for more effective variant surveillance.
- Widespread community immunity.
- Reduced clinical and public health impact of identifying specific variants.
As a result, VOC PCR test requests will NOT be accepted.
Testing Indications
SARS-CoV-2 WGS is not performed for diagnostic purposes. Traditional testing indications do not apply. If suspecting a SARS-CoV-2/COVID-19 infection, refer to PHO's Test Information Index for information on diagnostic test ordering.
Routine SARS-CoV-2 Surveillance by WGS
PHO’s laboratory randomly selects a proportion of all SARS-CoV-2 PCR positive respiratory specimens for WGS surveillance that meet the following criteria:
- Respiratory specimen in transport medium (nasopharyngeal swab collection preferred) AND
- SARS-CoV-2 positive by PCR AND
- SARS-CoV-2 PCR cycle threshold (Ct) value must be < 30 AND
- Remaining specimen volume at least 1 mL
Sampling proportions are adjusted periodically to account for overall positivity rates, the number of PCR positive specimens, and laboratory testing capacity. The proportion of positive specimens sequenced may reach 100% if capacity allows. This approach limits selection bias, better represents the SARS-CoV-2 variants in circulation in Ontario, and enhances our ability to detect new and emerging SARS-CoV-2 variants.
SARS-CoV-2 WGS Support in Other Scenarios
SARS-CoV-2 WGS support may be offered upon request in specific scenarios*:
- Suspected reinfection – sequencing can be requested to support the classification of a case as reinfection if the case does not meet the time-based reinfection criteria as outlined in the Coronavirus Case Definitions and Disease-Specific Information document.
- Outbreak investigations – a Lineage or In-Depth Genomic Analysis Report can be requested to support certain outbreak investigations.
* Requests for SARS-CoV-2 WGS in the above scenarios must be approved by a PHO Microbiologist.
Refer to the SARS-CoV-2 Whole Genome Sequencing and Analysis Request guidance document for explanations of available report formats and contact IORT@oahpp.ca for support. Refer to Submission and Collection Notes #3 below.
Specimens that do not meet the above criteria will not be tested. Positive specimens indicating suspected vaccine breakthrough will not be tested.
Specimen Requirements
| Test Requested | Required Requisition(s) | Specimen Type | Minimum Volume | Collection Kit |
SARS-COV-2 Whole Genome Sequencing |
SARS-CoV-2 PCR-positive respiratory specimen in transport medium (Ct ≤ 30) |
1.5 ml preferred (min 1.0 ml) |
NA |
Submission and Collection Notes
Complete all fields of the requisition form, including:
- Test(s) requests and indications for testing
- Patient setting/population/source
- Specimen location at the time of request
- Clinician information including name, CPSO and address
- One SARS-CoV-2 gene target Ct value (if multiple targets were tested choose the lowest Ct value ≤ 30)
For clinical specimens, label the specimen container(s) with the patient’s first and last name, date of collection, and one other unique identifier such as the patient’s date of birth or Health Card Number. For additional information see: Criteria for Acceptance of Patient Specimens. Failure to provide this information may result in rejection or testing delay.
Requests for SARS-CoV-2 WGS in Other Scenarios must be approved by a PHO Microbiologist. Email IORT@oahpp.ca to complete the request form and obtain approval/WGS-request ID before shipping any specimens. If approved, submitters should confirm the specimen’s eligibility and availability with the original testing lab (if not tested at PHO) and arrange for specimens to be sent to PHO. Submitters will receive information on signing up for a secure Incident and Outbreak Response Collaboration SharePoint Site account that will be used to provide the In-Depth Genomic Analysis report.
Limitations
This laboratory-developed approach is used for surveillance purposes and public health management only.
Some specimens with a SARS-CoV-2 PCR Ct value of ≤30 may not be successfully sequenced due to sensitivity differences across different SARS-CoV-2 PCR assays, nucleic acid degradation, or sequencing technical issues.
Storage and Transport
Specimens should be stored at refrigerated 2 to 8°C following initial SARS-CoV-2 PCR testing and shipped to PHO’s laboratory at the same temperature. All clinical specimens must be shipped in accordance to the Transportation of Dangerous Good Act.
If the anticipated transport time to the laboratory is >72 hours, the specimen should be kept frozen at -70 to -80°C.
Test Frequency and Turnaround Time (TAT)
SARS-CoV-2 WGS is performed daily at PHO’s laboratory-Toronto. Results are available within 14 calendar days from receipt of the specimen at PHO.
The SARS-CoV-2 WGS process is laboratory-developed. SARS-CoV-2 nucleic acids are extracted from a specimen and the complete or near-complete viral genome is amplified by multiplex PCR. The resulting PCR reaction is then sequenced on a commercial sequencing platform. The corresponding SARS-CoV-2 sequences are then analyzed using computational tools to determine the SARS-CoV-2 variant classification 1 .
Interpretation
The following table provides possible SARS-CoV-2 WGS results and interpretations:
| SARS-CoV-2 WGS Result* |
Possible Interpretation |
|---|---|
|
Negative |
A SARS-CoV-2 Variant of Concern was not detected. The lineage being defined is based on Pangolin schema and is not currently considered as variant of concern (VOC). |
|
Unable |
Unable to generate a SARS-CoV-2 genome sequence. SARS-CoV-2 was not successfully sequenced for a lineage assignment. This could be due to low viral load in the specimen, PCR inhibition, or other technical issues. |
|
SARS-CoV-2 B.1.1.529 |
SARS-CoV-2 B.1.1.529 VOC (Omicron) detected. This indicates that a SARS-CoV-2 variant of concern was detected. No additional action is required. |
*Note: The Whole Genome Sequencing assay is being used for surveillance purposes.
Reporting
Routine SARS-CoV-2 Surveillance by WGS
Results are reported to the physician, authorized health care provider (General O. Reg 45/22, s.18) or submitter as indicated on the requisition.
The information collected from SARS-CoV-2 WGS is summarized in a monthly report: SARS-CoV-2 Whole Genome Sequencing in Ontario.
SARS-CoV-2 WGS Support in Other Scenarios (Suspected reinfections and Outbreak investigations)
Genomic Analysis Reports are provided electronically via a secure portal accessible only to the requesting healthcare professional.
References
- World Health Organization (WHO). 2023. Statement on the update of WHO’s working definitions and tracking system for SARS-CoV-2 variants of concern and variants of interest. Available from: https://www.who.int/news/item/16-03-2023-statement-on-the-update-of-who-s-working-definitions-and-tracking-system-for-sars-cov-2-variants-of-concern-and-variants-of-interest
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