Human Immunodeficiency Virus - HIV | Choose the Right Test

Medical Experts

Contributor

Hillyard

David R. Hillyard, MD

Professor of Pathology, University of Utah

Medical Director, Molecular Infectious Diseases, ARUP Laboratories

Contributor

McMillin

Gwendolyn A. McMillin, PhD, DABCC (CC,TC), FADLM

Professor of Pathology (Clinical), University of Utah

Scientific Director, Mass Spectrometry Platform; Medical Director, Clinical Toxicology, ARUP Laboratories

Contributor

Slev

Patricia R. Slev, PhD, D(ABCC)

Professor of Pathology (Clinical), University of Utah

Section Chief, Immunology; Medical Director, Immunology Core Laboratory, ARUP Laboratories

HIV type 1 (HIV-1) and type 2 (HIV-2) are the two etiologic agents of AIDS. Although pre- and postexposure treatments have become available to avoid contracting HIV, infection rates remain high among at-risk populations, and HIV continues to be a major public health problem.

Both population screening and more frequent screening in those at risk are recommended in asymptomatic individuals because early detection of the disease is critical for improving treatment outcomes. Diagnostic testing includes “fourth generation testing” that detects both HIV antigens and antibodies, HIV-1/HIV-2 antibody differentiation testing, and nucleic acid amplification testing (NAAT). For more information, refer to the Screening and Diagnostic Testing section.

Antiretroviral drug resistance testing is recommended before the start of antiretroviral therapy (ART) for HIV to guide treatment selection and whenever treatment failures occur. For more information, refer to the Drug Resistance Tests section.

CD4 lymphocyte count and viral load testing can be used to monitor treatment response. For more information, refer to the Monitoring section.

Quick Answers for Clinicians

How quickly can HIV be detected after infection?

The length of time after exposure before HIV infection can be detected (window period) depends on the type of test used. Fourth generation antigen/antibody tests can detect HIV 18-45 days after infection; antibody-only tests require additional time to yield reliable results (23-90 days after exposure). HIV RNA testing can identify HIV 10-33 days after infection, but this testing is not generally recommended for initial screening in asymptomatic patients. However, HIV RNA testing is recommended in certain cases, such as in patients who present with symptoms of HIV infection with known recent exposure, in infants born to individuals with HIV, and for monitoring of disease (refer to the Laboratory Testing section).^,

Is repeat testing recommended after a negative HIV test result?

If a test result is negative in a patient with a possible exposure, testing should be repeated after the “window period” (the length of time after exposure before HIV infection can be detected) to confirm the negative result. In general, repeat testing should be considered in high-risk patients if clinically indicated.

Can preexposure prophylaxis affect HIV test results?

The use of preexposure prophylaxis (PrEP) can reduce the sensitivity of diagnostic tests, including antibody testing, antigen testing, and HIV RNA testing.

What is transmitted drug resistance, and how does it relate to antiretroviral testing?

Individuals with HIV who have resistant viral strains may pass those strains on to others. Transmitted drug resistance is linked to impaired virologic response to initial antiretroviral therapy (ART) regimens. At least 10% of individuals with HIV in high-income countries who have not yet been treated with ART have resistance to at least one ART drug,which underscores the importance of resistance testing in treatment-naive patients.

Should antiretroviral therapy be delayed until drug resistance test results are obtained?

Antiretroviral therapy (ART) should generally begin within 7 days of an HIV diagnosis, and as early as the day of diagnosis if immediate treatment can be coordinated. The start of treatment should not be postponed until resistance test results are received; the regimen can be adjusted if necessary after results are in.

What is the optimal timing for resistance testing in cases of treatment failure?

In patients who experience virologic failure during antiretroviral therapy (ART), resistance testing is most informative if performed while the patient is still receiving the regimen or within 4 weeks of therapy cessation to ensure that the resistant virus will be detected if present. Acquired drug resistance in HIV occurs during therapy as a result of drug pressure (selection). When an ART regimen is discontinued or interrupted, the HIV viral population within a patient may revert to primarily wild type within 4-6 weeks.The drug-resistant virus may still be present, but at concentrations below the limit of detection of genotypic or phenotypic tests. However, even low concentrations of drug-resistant virus can lead to treatment failure when ART is reinitiated.

What is the role of medical laboratory testing for infants exposed to HIV?

In infants and children age <18 months with perinatal or postnatal HIV exposure, including through breastfeeding, virologic assays (HIV RNA or HIV DNA nucleic acid amplification testing [NAAT]) that detect HIV directly must be used to diagnose HIV. At least two sequential negative virologic tests are required for definitive exclusion of HIV infection in nonbreastfed infants. A positive virologic test should be confirmed by repeat testing as soon as possible. An expedited antibody test should be used to identity perinatal HIV exposure if the birthing parent’s HIV test results are unavailable. Follow-up HIV NAAT should be performed in the infant if the antibody test is positive, and the birthing parent should be offered standard HIV testing as soon as possible. For specific testing intervals, refer to the Guidelines for the Use of Antiretroviral Agents in Pediatric HIV Infection.

Indications for Testing

Recommendations for laboratory testing to screen for HIV vary based on the patient and recommending organization. Screening at least once is recommended in all individuals; more frequent screening is recommended in individuals at high risk and those who are pregnant. Groups at greatest risk for HIV include men who have sex with men (MSM); individuals with HIV-positive sexual partners; individuals who use intravenous drugs; individuals with other sexually transmitted infections; and individuals who have had multiple sex partners since their last HIV test or have had partners with HIV risk factors or an unknown sexual history. Additionally, screening is also appropriate in individuals considering pre- or postexposure prophylaxis.

Guideline Recommendations for HIV Screening
CDC^a (endorsed by AAFP, ACOG, ACP, IDSA)	All individuals 13-64 yrs of age (testing recommended at least once) Patients at high risk for HIV (annual screening recommended) Pregnant individuals (routine prenatal screening recommended)
USPSTF^b	All individuals 15-65 yrs of age Individuals <15 yrs or >65 yrs at increased risk of infection Pregnant individual, including untested individuals who present in labor with unknown HIV status
^aRecommendations issued initially in 2006 and current in 2018. ^bRecommendations issued in 2019. AAFP, American Academy of Family Physicians; ACOG, American College of Obstetricians and Gynecologists; ACP, American College of Physicians; IDSA, Infectious Diseases Society of America; USPSTF, U.S. Preventive Services Task Force Sources: CDC, 2014; CDC, 2018; USPSTF, 2019; ACOG, 2014; ACOG, 2018

Laboratory testing to diagnose HIV infection is appropriate in individuals with a positive screening test result, symptoms of HIV infection, or recent exposure to a person with HIV, and in infants born to mothers with HIV.^,^,

Testing to monitor treatment is appropriate for patients who are currently receiving ART for HIV.

Laboratory Testing

Screening and Diagnostic Testing

Many types of tests are used to screen for and diagnose HIV. Rapid screening tests can be performed at the point of care (POC) and provide results within minutes; however, these tests are not as reliable as combined antigen/antibody tests (fourth generation tests), which can be used for screening or diagnosis. Confirmatory tests are supplemental tests used to confirm the diagnosis in patients with repeatedly reactive fourth generation antigen/antibody tests. Finally, follow-up tests are used if discrepancies exist between the screening and confirmatory tests.

The types of tests used in screening and diagnosis are detailed below. Refer to ARUP's Human Immunodeficiency Virus in Adults and Adolescents Testing Algorithm for more information.

Treatment-Related Testing

Initial Pretreatment Tests

After an established diagnosis of HIV and before beginning treatment with ART, initial recommended laboratory tests include the HIV RNA level test, CD4 T-lymphocyte cell count, tests for other infections (eg, hepatitis B virus [HBV] and hepatitis C virus [HCV]), and tests to assess general health, such as a CBC, kidney and liver function, lipid level, and glucose level tests.^, A pregnancy test should be given to individuals who have a uterus and are of childbearing age before beginning treatment. ART initiation should not be delayed by these test results unless kidney or liver damage is discovered or transmitted drug resistance is strongly suspected.

Drug Resistance Tests

Drug resistance testing involves genotypic or phenotypic testing to evaluate viral strains and guide the selection of therapy regimens. These tests are designed to detect resistance to the various classes of ART drugs. Resistance testing should be performed in early HIV infection even if the patient has opted to postpone ART so that the results will be available when the individual begins therapy. However, repeat testing at the start of therapy should be considered because of the possibility that a drug-resistant virus (superinfection) has been acquired between the initial diagnosis and ART initiation. Resistance testing should also occur at treatment failure and when a regimen switch is planned.

Genotypic Tests

Genotypic testing is preferred over phenotypic testing in most contexts (eg, in patients with known drug resistant-variant patterns and in those anticipated to have uncomplicated variant patterns) because of its faster turnaround time, lower cost, and greater sensitivity for combinations of wild-type and resistant virus. Additionally, test interpretation is more straightforward for genotypic than phenotypic tests.

Genotypic assays generally use either Sanger sequencing or next generation sequencing (NGS) to identify variants in the reverse transcriptase (RT), protease resistance (PR), and integrase genes of circulating HIV RNA, which may assist in identifying resistance to nucleoside reverse transcriptase inhibitors (NRTIs), non-NRTIs (NNRTIs), integrase strand transfer inhibitors (INSTIs), and protease inhibitors (PIs). Testing is also available to detect variants in the gp41 gene, which are associated with resistance to fusion inhibitors (ie, enfuvirtide). The International Antiviral Society–USA has a list of resistance-associated variants, and the Stanford University HIV Drug Resistance Database is a useful resource to aid in genotypic test interpretation.

Therapy-naive patients: Genotypic testing for resistance-associated variants is indicated in patients who have not previously been treated with ART. If transmitted resistance to INSTIs is suspected, testing for variants that cause INSTI resistance (ie, variants in integrase genes) should be considered.

Therapy-experienced patients: In patients currently being treated with ART, resistance testing is recommended in those with an unsatisfactory decrease in viral load and in those with treatment failure, as indicated by virologic failure and HIV RNA measurements of more than 1,000 copies/mL. Drug resistance testing may be less sensitive but can still be considered in patients with HIV RNA values less than 1,000 copies/mL, such as patients with an HIV RNA value between 500 and 1,000 copies/mL. In cases of virologic failure, resistance testing should be performed while the patient is still undergoing the treatment regimen or within 4 weeks of treatment discontinuation to increase the likelihood that resistance variants will be detected. If treatment failure occurs during therapy with INSTIs, testing for INSTI resistance should be performed. INSTI resistance testing should include NRTI, NNRTI, and PI testing or should be performed alongside testing for those resistances.

Phenotypic Tests

Phenotypic assays are used to evaluate whether the virus is able to replicate in the presence of various ART drug concentrations. Phenotypic testing should be performed in addition to genotypic testing in patients who are known or anticipated to have complex drug-resistant variant patterns. Phenotypic coreceptor tropism assays are also preferred over genotypic assays when a CCR5 antagonist (or receptor inhibitor [RI]) is under consideration or when a patient experiences virologic failure while receiving a CCR5 antagonist (eg, maraviroc). Coreceptor tropism testing is also useful to guide treatment in ART-experienced patients with significant drug resistance or consistently high concentrations of HIV RNA.

Drug Hypersensitivity Test

The HLA-B*5701 genotyping test is required by the FDA before treatment with abacavir to identify patients at risk for abacavir hypersensitivity.^, For more information, refer to the ARUP Consult Germline Pharmacogenetics and HLA Testing topics.

Monitoring

Quantitative viral load testing can help to determine a therapeutic plan and is used for monitoring during treatment. After the initiation of ART, patients should be evaluated within 6 weeks to assess their viral load, as well as their ability to tolerate the therapy and adherence to treatment. Appropriate monitoring tests include quantitative NAAT and quantitative CD4 lymphocyte tests. Although the absolute CD4 lymphocyte count can aid in the monitoring of HIV, CD4 testing is not required for patients once viral suppression has been established (ie, is stable). The HIV Medicine Association recommends the CD4 count be monitored every 3-6 months for the first 2 years after treatment begins; after 2 years, in patients with an undetectable viral load and CD4 counts of 300-500 cells/mm³, the CD4 count should be measured annually. CD4 monitoring can be considered optional once the CD4 count is consistently greater than 500 cells/mm³.