HIV Testing and Monitoring

HIV testing detects infection with human immunodeficiency virus type 1 or 2 by antibody, antigen or nucleic acid. Fourth generation laboratory assays detect both HIV antibody and the p24 antigen, shortening the window period to about 14 to 21 days after exposure. Rapid point-of-care tests (finger prick or oral fluid) detect antibody in 15 to 20 minutes.

Nucleic acid tests (HIV RNA) detect infection within 10 to 14 days and are used for acute infection and infant diagnosis. Once infection is confirmed, CD4 count and HIV viral load stage the disease and monitor treatment. Resistance genotyping directs the choice of antiretroviral drugs.

Early diagnosis allows treatment that restores life expectancy to near normal, prevents transmission (undetectable equals untransmittable) and prevents mother-to-child transmission to under 1%. Delayed diagnosis remains common: around 40% of people present with CD4 counts under 350 cells/microL.

About 39 million people live with HIV worldwide, with 1.3 million new infections and 630 000 deaths in 2022. About two thirds of people with HIV live in sub-Saharan Africa. About 86% of those with HIV know their status, and 76% are on treatment.

Indications.

• Everyone aged 15 to 65 years once, and repeated by risk level.
• All pregnant women at the first antenatal visit, and again in the third trimester in high prevalence settings.
• Patients with tuberculosis, sexually transmitted infections, hepatitis B or C, lymphoma, unexplained weight loss, fever of unknown origin, candidiasis, shingles, unexplained cytopenias or dementia.
• People who inject drugs, sexual partners and children of infected people, and exposed healthcare workers.
• Infants born to infected mothers, and before pre-exposure prophylaxis.

Specimen. Serum, plasma or whole blood for laboratory assays. Capillary blood or oral fluid for rapid tests. EDTA plasma for viral load (separated within 6 hours, or dried blood spots for infant diagnosis). EDTA whole blood for CD4 count and genotyping. Consent and counselling are needed, with confidentiality and linkage to care.

Testing sequence.

• Initial test: fourth generation antigen and antibody assay or a rapid test. Non-reactive means no infection, unless exposure was in the last 2 to 4 weeks (retest at 4 to 6 weeks, or use an RNA test).
• Reactive initial test: confirmed by an assay that separates HIV-1 and HIV-2 antibodies (differentiation immunoassay), then HIV-1 RNA if antibody results are negative or indeterminate (acute infection).
• Rapid test algorithms in resource limited settings use two or three different rapid tests in series (first reactive, second and third reactive), with an unreactive result on a different test excluding infection.
• A negative RNA with reactive antibody: false positive of the screening assay.
• Positive RNA with negative antibody: acute HIV infection.

Window period. Time from infection to detection: RNA 10 to 14 days, fourth generation 14 to 21 days, third generation antibody 3 to 4 weeks, rapid antibody tests 3 to 12 weeks. Almost all infections are detected by 6 weeks with a fourth generation test, and 12 weeks with rapid tests.

Infant diagnosis. Maternal antibody persists to 18 months, so antibody tests cannot diagnose infants. Nucleic acid testing (DNA or RNA PCR) at birth in high risk infants, 4 to 6 weeks, and 4 to 6 weeks after stopping breastfeeding, with a final antibody test at 18 months in exposed infants.

CD4 count. Normal 500 to 1500 cells/microL (30 to 60% of lymphocytes).

• Over 500: low risk of opportunistic infection.
• 200 to 500: moderate immunosuppression, with risk of tuberculosis, shingles, and oral candidiasis.
• Under 200: AIDS-defining threshold. Risk of Pneumocystis pneumonia, cryptococcal meningitis, toxoplasmosis and mycobacterial disease. Prophylaxis with co-trimoxazole is started.
• Under 100: risk of cytomegalovirus, cryptococcus and disseminated Mycobacterium avium complex.
• Under 50: highest risk, with very high mortality.

HIV viral load. Expressed as copies/mL and log10.

• Baseline: typically 10^4 to 10^6 copies/mL. Over 100 000 indicates high risk of rapid progression.
• On treatment: viral suppression (under 50 copies/mL, or under 200 by some definitions) should be reached by 12 to 24 weeks. Failure is a confirmed value over 1000 copies/mL after at least 6 months of adherent treatment.
• Low level viraemia (50 to 1000 copies/mL) is monitored, and persistent values over 200 raise concern for adherence problems or resistance.
• Viral blips (single values 50 to 1000 with return to suppression) are not failure.
• Monitoring: at 3 months after starting treatment, then every 6 to 12 months once suppressed.

Resistance genotyping. Before starting therapy if resistance prevalence is over 10% in the community, in treatment failure (while the patient is still on the failing regimen, with viral load over 500 to 1000 copies/mL), in pregnancy, and in post-exposure prophylaxis failure.

HLA-B*5701. Predicts abacavir hypersensitivity, and is tested before abacavir is started. Tropism assay is tested before maraviroc.

Other baseline tests. Hepatitis B and C serology, syphilis serology, tuberculosis screening, creatinine, liver function, full blood count, lipids, glucose, G6PD where relevant, toxoplasma IgG, cryptococcal antigen if CD4 is under 200, and pregnancy test.

Critical values and findings needing immediate action.

• A reactive initial test: confirm the same day where possible, counsel, and link to care without delay.
• CD4 under 200: start co-trimoxazole prophylaxis, screen for tuberculosis and cryptococcal antigen, and start antiretroviral therapy promptly (tuberculosis and cryptococcal meningitis timing is individualised).
• Positive cryptococcal antigen in serum: lumbar puncture, pre-emptive antifungal therapy.
• Acute HIV infection (fever, rash, lymphadenopathy with negative antibody, positive RNA): start treatment early, and notify partners.
• Viral load over 1000 copies/mL on treatment: adherence support, resistance test, and regimen change chosen from results.
• Pregnant woman positive: start treatment at once, and arrange infant prophylaxis and testing.
• Occupational exposure: post-exposure prophylaxis within 72 hours (ideally within 2 hours).

Interferences and pitfalls.

• Testing within the window period gives false negatives, and retesting is needed when recent exposure is reported.
• False positive screening results occur with pregnancy, autoimmune disease, recent vaccination, and acute infections. Confirmation is always required before a diagnosis is given.
• Indeterminate Western blot or line immunoassay results need RNA testing.
• Patients on pre-exposure prophylaxis or early treatment may show delayed or weaker antibody responses.
• Rapid tests are less sensitive in early infection and with oral fluid.
• Elite controllers have undetectable viral load without treatment.
• HIV-2 is not reliably measured by HIV-1 viral load assays.
• CD4 count varies by 20 to 30% with the time of day, intercurrent infection, steroids, and splenectomy. Percentage is more stable.
• Viral load blips after vaccination or infection can be mistaken for failure.
• Stigma and consent: results must be given in person with counselling, and never through third parties.

Special groups.

• Pregnancy: universal testing, repeat in late pregnancy for high risk women, treatment to a suppressed viral load before delivery reduces transmission to under 1%.
• Infants: nucleic acid testing, and antiretroviral prophylaxis for 4 to 6 weeks (longer if the mother is not suppressed).
• Adolescents and young women in sub-Saharan Africa: disproportionately high incidence, with self-testing as an option.
• Older adults: late diagnosis is frequent, with lower CD4 counts at presentation.
• Regions: self-testing kits, point-of-care viral load and CD4 platforms, and community testing extend access.

Follow up tests.

• HIV viral load and CD4 count, and genotypic resistance test.
• Hepatitis B surface antigen, hepatitis C antibody, and syphilis serology.
• Tuberculosis screening (symptoms, chest radiograph, Xpert, urine lipoarabinomannan).
• Cryptococcal antigen, toxoplasma serology, and cytomegalovirus testing.
• Cervical screening, creatinine and urinalysis (tenofovir), lipid profile and glucose, and HLA-B*5701.
• HPV and anal cytology in selected groups.