HIV – Diagnostic Serology

Testing Indications

HIV testing should be considered as part of routine care. Specifically, testing is recommended for individuals:

• with symptoms of acute HIV infection
• with symptoms associated with chronic HIV infection
• who have had a high-risk exposure
• who belong to a population with higher rates of HIV
• who are initiating immune suppressing therapy
• who are pregnant; if ordering as part of a prenatal screening, see Prenatal – Serology

For more information on HIV testing recommendations, refer to the Ontario Guidelines for Providers Offering HIV Testing.

Acceptance/Rejection Criteria

• Cadaveric blood testing services are not available through PHO. Requests can be submitted to Mt. Sinai Hospital Microbiology Laboratory at 416-586-4800 extension 4432.
• Donor testing is not available at PHO. Specimens from patients being screened as potential donors (e.g. organ, tissue, cells, fertility, etc.) should be referred to a laboratory that performs donor screening assays. Specimens submitted for donor screening to PHO will be rejected.
• Haemolysed, icteric, lipemic or microbially contaminated sera are unacceptable for testing.
• Mismatched information on the specimen tube label and requisition will be rejected.

Timing of Specimen Collection

PHO uses a 4^th generation HIV Ag/Ab screening test. In general, individuals will test reactive 2-4 weeks after exposure (median 18 days) and >99% of individuals will be reactive by 6 weeks.¹

Storage and Transport

Place the specimen tube in a biohazard bag and seal it. Place completed HIV Serology and PCR Test Requisition in the pouch at the front of the biohazard bag.

To avoid delays in testing, ship specimens for testing to PHO’s laboratory immediately after collection or processing. Spun SST or separated serum stored at 2-8°C must be shipped with ice packs to PHO’s laboratory.  If delayed shipping is anticipated, remove serum from clot and store frozen at -20°C or colder, and ship on dry ice.

All clinical specimens must be shipped in accordance with the Transportation of Dangerous Good Act.

Test Frequency and Turnaround Time (TAT)

HIV diagnostic serology testing is performed daily Monday to Friday.

Turnaround time is up to 3 business days for non-reactive specimens and up to 6 business days for reactive specimens from receipt by PHO’s laboratory.

Stat and Critical Samples Testing

STAT testing should only be requested when it directly affects patient care in an emergency medical circumstance. If warranted, the following HIV testing is expedited:

• Testing of the ‘source patient’ for needlestick/occupational exposure.
• A pregnant individual presenting in labour who has not received prenatal care or for whom results of previous testing are not readily available and are at high risk for HIV.

Prior to sending, PHO’s laboratory Customer Service Centre must be notified at 416-235-6556 / 1-877-604-4567. STAT samples must be shipped separately from routine specimens and delivered to PHO’s laboratory in a timely manner. The outer package must be clearly marked indicating ‘STAT’. It must be handled in accordance with the Canadian Biosafety Standards and shipped in accordance with the Transportation of Dangerous Goods Regulations. Failure to ship separately will delay testing.

After-hours STAT testing requires approval . Contact the after-hours Duty Officer at 416-605-3113 if outside of PHO’s laboratory operating hours.

PHO uses the following methods for HIV diagnostic testing:

1. HIV-1/-2 antigen (Ag)/antibody (Ab) combination screen: Chemiluminescent microparticle immunoassay (CMIA) that simultaneously detects HIV-1 p24 Ag and HIV-1 and HIV-2 Ab. This assay does not differentiate between p24 and HIV-1/-2 Ab.
2. HIV-1/-2 antibody confirmatory test: Geenius™ immunochromatographic test for confirmation and differentiation of antibodies to HIV-1 and HIV-2.
3. HIV-1/-2 nucleic acid test (NAT): Qualitative NAT for the detection and differentiation of HIV-1 and HIV-2 RNA.

Algorithm

Serum specimens submitted for HIV serology are tested following the algorithm below:

Note: To help determine HIV status and guide treatment decisions, HIV PCR will be prioritized over HIV serology for individuals ≤18 months of age. If there is a high suspicion of acute HIV in the window period (i.e. prior to seroconversion), consider requesting both HIV serology and HIV PCR testing.

Interpretation

Results should be interpreted in the context of the patient’s clinical presentation, risk factors, and exposure history, and treatment. The table below provides a general overview of the algorithm and the most common interpretations for HIV diagnostic testing for adults. It does not include all possible result scenarios. These interpretations may not apply to infants as maternal antibodies can circulate for 18-24 months after birth. Consider referral to a paediatric HIV specialist for management of infants suspected of HIV infection.

HIV-1/HIV-2 Ag/Ab Combo Screen	HIV-1/HIV-2 Ab Confirmatory	HIV-1/HIV-2 PCR	Interpretation
Non-reactive	Not tested	Not tested	Non-reactive for HIV-1 p24 antigen and HIV-1/-2 antibodies. For high-risk exposures, repeat HIV testing at 3 weeks and, if negative, at 6 weeks after possible exposure or on presentation of symptoms compatible with acute HIV infection. If there is a high suspicion of acute HIV in the window period (i.e. prior to seroconversion), consider requesting HIV PCR testing.
Reactive/ Indeterminate	Negative/ Indeterminate	HIV-1 and HIV-2 RNA not detected	Inconclusive for HIV infection.   Repeat testing in at least 4 weeks from date of collection. If two or more inconclusive results have been obtained on separate specimens, no further testing for HIV is recommended unless clinically warranted.
Reactive/ Indeterminate	HIV-1 positive	Not tested	Evidence of HIV-1 infection.
Reactive/ Indeterminate	HIV-2 positive	Not tested	Evidence of HIV-2 infection.
Reactive/ Indeterminate	HIV-1 and HIV-2 positive	Not tested	Evidence of HIV-1 and HIV-2 co-infection.
Reactive/ Indeterminate	Negative/ Indeterminate	HIV-1 RNA detected	Evidence of acute HIV-1 infection.
Reactive/ Indeterminate	Negative/ Indeterminate	HIV-2 RNA detected	Evidence of acute HIV-2 infection.
Reactive/ Indeterminate	Negative/ Indeterminate	HIV-1 and HIV-2 RNA detected	Evidence of acute HIV-1 and HIV-2 co-infection.

A PHO Microbiologist is available for questions about HIV serology results and interpretations by calling 416-235-6556 / 1-877-604-4567 or by email at CustomerServiceCentre@oahpp.ca.

For individuals who test positive for HIV-1, consider submitting a follow-up EDTA whole blood or plasma specimen for HIV-1 viral load testing. Refer to the HIV-1 Viral Load Test Information Sheet for detailed information.

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.

The PHO Laboratory Enhancement Program (LEP) questionnaire and a CATIE letter will accompany all first-time HIV-positive results .

• LEP Questionnaire: A voluntary questionnaire used to gather patient demographics (e.g., race/ethnicity, country of birth), HIV risk factors, and testing/treatment history. This data is essential for provincial HIV surveillance in Ontario and informs policy and programming decisions.
• CATIE Information Sheet: An information sheet that includes resources to help newly diagnosed individuals access treatment, understand the role of public health in follow-up and contact notification, and offers strategies to prevent transmission, along with resources for support services.

Specimens that are positive for HIV diagnostic testing are reported to the Medical Officer of Health as per the Ontario Health Protection and Promotion Act.

Due to changes in PHO’s and Ontario’s laboratory information systems, cumulative reporting is no longer available.

References

1. Ontario Guidelines for Providers Offering HIV Testing