HIV-1 Drug Resistance and Genotype Testing

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Background
This page provides information for HIV-1 Drug Resistance Genotyping and other related testing at Public Health Ontario (PHO). Testing is referred to the British Columbia Centre for Excellence (BCCfE) in HIV/AIDS.

For information regarding other testing options for HIV, refer to the following PHO webpages:

Updates

Discontinuation of gp-41 resistance testing.
The specimen requirements table has been updated with revised test names and added testing indications.
Updated test names in the Specimen Requirements table and added testing indications.
Updated specimen collection and handling instructions.
Special instructions for submitting Untimed Drug Level / Therapeutic Drug Monitoring (TDM) requests have been added.
Updated HIV-1 Drug Resistance and Genotyping Testing requisition form.

* Copie d'impression non contrôlée. Valide uniquement le jour de l'impression : 28 août 2025.

Testing Indications

HIV-1 Resistance Genotyping detects resistance mutations in HIV-1 to guide antiretroviral (ARV) therapy decisions. The test requires a viral load of ≥250 copies/mL and measures susceptibility to the following ARV drug classes:
1. Nucleoside reverse transcriptase inhibitors (NRTIs)
2. Non-nucleoside reverse transcriptase inhibitors (NNRTIs)
3. Protease inhibitors (PIs)
4. Integrase strand transfer inhibitors (INSTIs)
Consider HIV-1 Resistance Genotyping for the following situations:

At the diagnosis of HIV-1 infection or before initiating ARV
Virologic failure or suboptimal virologic response to ARV
HIV-1-positive pregnant individual for prevention of mother-to-child transmission
Known or suspected transmission of drug-resistant HIV (e.g., sexual partner with resistant virus)
Switching regimens after a treatment failure or unknown resistance history

HIV-1 V3 Tropism testing (investigational) measures susceptibility to CCR5 antagonists (e.g. maraviroc) and should be done before starting a CCR5 antagonist. Two tests are available based on the patient’s viral load:

Plasma CCR5 Tropism – can be requested if the patient's HIV-1 viral load is ≥500 copies/mL.
Proviral DNA Tropism – can be requested if the patient's HIV-1 viral load is <500 copies/mL.

HLA-B*57:01 Abacavir Hypersensitivity test is used to determine whether the patient has the HLA-B*57:01 allele and therefore is at higher risk of abacavir hypersensitivity. Testing should be done before initiating an abacavir-containing regimen. The test only needs to be done once as the results do not change over time. No minimum viral load is required.

BCCfE offers therapeutic drug monitoring (TDM), both time and untimed, as an investigational test in cases of treatment failure. TDM is not offered routinely. Contact PHO Customer Service before sending TDM requests by calling 1-877-604-4567 or emailing customerservicecentre@oahpp.ca.

Acceptance/Rejection Criteria

The following specimens will be rejected:

HIV-1 Resistance Genotyping requests on HIV-1 viral load specimens that are <250 copies/mL.
Plasma CCR5 Tropism request on HIV-1 viral load specimens that are <500 copies/mL.
Specimens submitted with an incomplete HIV-1 Drug Resistance and Genotype Testing Requisition.
Specimens with incomplete labels.
Specimens for Proviral DNA Tropism or HLA-B*57:01 Abacavir Hypersensitivity Testing collected on plasma preparation tubes (PPT).

Specimen Collection and Handling

Specimen Requirements

Test Requested	Required Requisition(s)	Specimen Type	Minimum Volume	Collection Kit
HIV-1 Drug Resistance Genotyping	HIV-1 Drug Resistance and Genotype Testing Requisition	Plasma or previously submitted HIV-1 viral load specimen ^2,5,6	2.5 mL	- PPT or - EDTA Tube
HIV-1 Drug Resistance Genotyping Reinterpretation	HIV-1 Drug Resistance and Genotype Testing Requisition	Previously submitted HIV-1 Resistance Genotyping specimen^3,6	n/a	n/a
Plasma CCR5 Tropism	HIV-1 Drug Resistance and Genotype Testing Requisition	Plasma or previously submitted HIV-1 viral load specimen^4,5,6	2.5 mL	- PPT or - EDTA Tube
Proviral DNA CCR5 Tropism	HIV-1 Drug Resistance and Genotype Testing Requisition	Unspun EDTA Whole blood⁷	3 mL	- EDTA Tube
HLA-B*57:01 Abacavir Hypersensitivity	HIV-1 Drug Resistance and Genotype Testing Requisition	Unspun EDTA Whole blood⁷	3 mL	- EDTA Tube
Therapeutic Drug Monitoring		See Special Instructions.

Submission and Collection Notes

Use the dedicated HIV-1 Drug Resistance and Genotype Testing Requisition. Complete all sections of the requisition.

HIV-1 Resistance Genotyping requires a minimum HIV-1 viral load (VL) of ≥250 copies/mL and can be requested on a new or previously submitted HIV-1 VL specimen.

HIV-1 Resistance Genotyping reinterpretation can only be ordered on a previously tested specimen; drug class(es) must be selected.

Plasma CCR5 Tropism requires a minimum HIV-1 VL of ≥500 copies/mL and can be requested on a new or previously submitted HIV-1 VL specimen.

HIV-1 VL will be tested to determine eligibility for tests that require a minimum HIV-1 VL, if not already done.

If requesting a test on a previously submitted specimen, a completed HIV-1 Drug Resistance and Genotype Testing Requisition must be faxed to PHO Customer Service at 416-235-6552. A PHO’s lab specimen number and result date must be provided on the requisition.

Proviral DNA CCR5 Tropism and HLA B*57:01 Abacavir Hypersensitivity testing requires one (1) unspun 3 mL EDTA whole blood tube. Do NOT centrifuge. One tube is sufficient for both tests.

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. Patient information on the specimen must match the requisition, or the specimen will be rejected.

Storage and Transport

Place the specimen in a biohazard bag and seal it. To avoid delays in testing, ship specimens to PHO’s laboratory immediately after collection or processing. EDTA whole blood must be shipped within 24 hours of collection on ice packs to the nearest PHO location. If a delay is anticipated, transfer the EDTA whole blood into aliquot tube(s), store at -20°C and ship frozen to the nearest PHO location. Plasma specimens can be stored and/or transported at 2-8°C for up to 6 days or at ≤ -18°C for up to 12 weeks. If extended storage is required, plasma specimens should be frozen at ≤-60°C.

Shipping of specimens should be done by Transportation of Dangerous Goods (TDG) certified individuals in accordance with TDG regulations.

Special Instructions

Before submitting samples for Therapeutic Drug Monitoring (timed and untimed), contact PHO’s Customer Service Centre by calling 1-877-604-4567 or emailing customerservicecentre@oahpp.ca for collection and submission instructions.

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

HIV-1 Resistance Genotyping and other related tests are referred to the British Columbia Centre for Excellence (BCCfE) in HIV/AIDS and shipped weekly on Mondays from PHO’s laboratory, Toronto location.

Turnaround time is up to 21 business days from receipt of specimens at the PHO.

Test Methods

BCCfE uses reverse transcription-polymerase chain reaction (RT-PCR) and sequencing of specific HIV regions to detect mutations associated with HIV-1 drug resistance. Mutations are interpreted using several databases, including the Stanford HIV Drug Resistance database.

Algorithm

HIV-1 Resistance Genotyping is automatically reflexed for all baseline and first-time HIV-1 viral load testing in patients with a HIV-1 viral load of ≥250 copies/mL.

Reporting

Results are reported to the physician, authorized health care provider (General O. Reg 45/22, s.18) or submitter as indicated on the requisition.