Diagnostic testing for HIV infection in infants and children younger than 18 months

INTRODUCTION — Promptly establishing the diagnosis of human immunodeficiency virus (HIV) infection in infants and young children is important because infants who are infected with HIV have a high morbidity and mortality if treatment with antiretroviral therapy (ART) is delayed [1]. In infants and young children, HIV testing must be performed using nucleic acid tests (NATs). Antibody-only and antigen/antibody combination assays that are commonly used for HIV testing in adults and older children are not reliable in infants and young children, because of the persistence of transplacentally acquired maternal antibody.

This topic reviews the strategy for diagnosing HIV infection in infants and young children of mothers with known HIV infection or of uncertain HIV serostatus.

We recognize that not all pregnant, postpartum, and lactating individuals identify as women or mothers. Using gender-inclusive language, however, is not possible in all languages and all countries and for all our readers. The topics discussed here are based on risks driven by biological sex and not gender identity. Therefore, throughout this topic, we use the term “woman” to signify a person of the female sex (regardless of gender identity) and the term “mother” to signify the female biological parent of a child (regardless of gender identity).

The diagnosis of HIV infection in adults and older children (older than age 18 months) and strategies for screening during pregnancy to prevent mother-to-child transmission of HIV are discussed separately:

●(See "Screening and diagnostic testing for HIV infection".)

●(See "Prevention of HIV transmission during breastfeeding in resource-limited settings".)

●(See "Antiretroviral selection and management in pregnant individuals with HIV in resource-rich settings".)

●(See "Prenatal evaluation of women with HIV in resource-rich settings".)

●(See "Acute and early HIV infection: Clinical manifestations and diagnosis", section on 'Diagnostic test performance in early HIV infection'.)

CHALLENGES OF HIV TESTING IN INFANCY — Antibody assays are commonly used for detection of HIV infection in adults and older children. These tests are highly sensitive and specific, though they may be falsely negative in the first few weeks after acute infection. (See "Screening and diagnostic testing for HIV infection", section on 'Antibody-only tests'.)

Newer antigen/antibody combination assays can detect HIV-1/2 antibodies and HIV-1 p24 antigen. Including the latter shortens the window period and increases the detection of early infection. (See "Screening and diagnostic testing for HIV infection", section on 'Combination HIV antigen and antibody tests'.)

The diagnosis of HIV infection in infants and young children differs from that in adults and older children because antibody-only and antigen/antibody combination assays will detect transplacentally transferred maternal HIV antibodies that persist for many months [2,3]. In a study of 234 uninfected children born to HIV-infected mothers who were followed for up to two years, the mean age of loss of maternal antibody was 10.3 months; one child did not lose maternal antibody until >18 months of age [2].

To identify HIV infection in infants and young children (ie, <18 months of age), HIV virologic testing must be performed using assays that detect HIV deoxyribonucleic acid (DNA) or ribonucleic acid (RNA), collectively referred to as "nucleic acid tests" (NATs) [4]. Cord blood should not be used for testing, because of the possibility of contamination of the sample with maternal blood. Scheduled repeated testing is necessary because test sensitivity increases with time from HIV virus exposure and very recent infection may not be detected.

DIAGNOSTIC STRATEGIES — The approach to the diagnosis of HIV in infants and young children (younger than 18 months of age) depends on whether the HIV status of the mother is known [5]. The following recommendations for establishing the diagnosis of HIV infection in infants and young children are based on the guidelines of the United States Department of Health and Human Services, the National Institutes of Health, and the American Academy of Pediatrics [5-7]. These guidelines also outline standard procedures for obtaining consent and for counseling.

Mother with HIV — In an infant or young child born to a mother with HIV, serial testing is performed with an HIV nucleic acid test (NAT) [7]. Timing of testing depends upon whether the infant’s risk for HIV acquisition is categorized as low or higher risk, based on factors including maternal prenatal care, antiretroviral therapy (ART), and viral suppression (table 1).

Formula-fed infants

●Low risk – Infants born to mothers with HIV who received standard ART during pregnancy with sustained viral suppression (usually defined as <50 copies/mL) and who were adherent to their ART regimen are at low risk of acquiring HIV.

For low-risk infants, testing is performed at 14 to 21 days after birth, one to two months of age, and four to six months of age [7]. A positive result at any point should be confirmed as soon as possible with a repeat NAT.

●High risk – Infants born to mothers with HIV who did not receive standard care for HIV during pregnancy are at higher risk of perinatal transmission. This includes infants born to mothers living with HIV who [7]:

•Did not receive prenatal care

•Did not receive antepartum ART or received only intrapartum antiretrovirals (ARVs)

•Initiated ART late in pregnancy (during the late second or third trimester)

•Received a diagnosis of acute or primary HIV infection during pregnancy or in labor

•Had detectable viral loads (≥50 copies/mL) close to the time of delivery, including those who received ART but did not have sustained viral suppression

For higher-risk infants, testing should be performed at birth, 14 to 21 days after birth, one to two months of age, two to six weeks after the completion of presumptive HIV therapy, and four to six months of age. A positive result at any point should be confirmed as soon as possible with a repeat NAT.

The rationale for additional testing in high-risk infants:

•Testing at birth – Testing at birth allows early HIV diagnosis and initiation of ART. It also permits determination of timing of the infection if the infant is infected. A positive result at ≤48 hours after birth reflects HIV infection in utero, while infants who have a negative test result during the first week of life and subsequently have positive test results are considered to have intrapartum infection.

•Testing two to six weeks after cessation of ARV prophylaxis – ARV prophylaxis, particularly combination ART, may reduce the sensitivity of testing during prophylaxis. Rather than deferring the earlier testing (at four to eight weeks of age) until after prophylaxis is discontinued (ie, 8 to 12 weeks of age), a strategy of testing at both time points allows prompt recognition of infected infants and may capture additional cases.

Breastfed infants — Infants who are breastfed by mothers with HIV are at higher risk of HIV acquisition due to ongoing potential exposure to HIV through breastmilk. The risks, benefits, and details on how to minimize risk through breastfeeding are discussed separately. (See "Intrapartum and postpartum management of pregnant women with HIV and infant prophylaxis in resource-rich settings", section on 'Breastfeeding'.)

Infants born to people with HIV who opt to breastfeed after comprehensive counseling should be closely monitored. Testing should proceed according to the high-risk testing schedule (table 1) and then at least every three months throughout breastfeeding, followed by monitoring at four to six weeks, three months, and six months after cessation of breastfeeding [7]. A positive result at any point should be confirmed as soon as possible with a repeat NAT.

Strategies to prevent HIV transmission during breastfeeding in resource-limited settings are discussed separately. (See "Prevention of HIV transmission during breastfeeding in resource-limited settings".)

Maternal HIV status unknown — Ideally, all women should be tested to identify HIV infection either prior to pregnancy or early in the prenatal course. Recommendations for HIV screening during pregnancy and in the general adult population are discussed in separate topics. (See "Prenatal care: Initial assessment", section on 'HIV' and "Screening and diagnostic testing for HIV infection", section on 'Routine screening'.)

If a woman presents in labor or at delivery without documentation of her HIV status, HIV testing should be performed on either the mother or infant. The results of serologic testing (mother or infant) reflect the HIV status of the mother, not the infant, since it is maternal antibodies to HIV that will be detected. If serologic testing for HIV in the mother or infant is positive, testing of the infant should proceed as described above. (See 'Mother with HIV' above.)

It is important that testing in the peripartum period be expedited so that ARV prophylaxis or presumptive HIV therapy can be given to the newborn as soon as possible, ideally within six hours of birth.

DIAGNOSTIC TESTS

HIV DNA polymerase chain reaction testing — A qualitative HIV-1 DNA polymerase chain reaction (PCR) assay that detects HIV proviral DNA within peripheral blood mononuclear cells has been the primary test used for the diagnosis of HIV-1 infection in infants born to mothers with HIV [5,6]. Whole-blood specimens or dried blood spot samples may be used for testing. The test should be positive in patients infected with HIV even while they are being treated with antiretroviral therapy (ART).

Approximately two-thirds of mother-to-child transmissions of HIV occurs perinatally, and DNA PCR testing will not detect very recent infection. The sensitivity of HIV-1 DNA PCR testing when performed at birth is 55 percent; it increases to >90 percent by two to four weeks of age and 100 percent at three and six months [8-10]. The specificity of HIV-1 DNA PCR is 99.8 percent at birth and 100 percent at one, three, and six months.

HIV clade B is the predominant viral strain in the United States. Non-clade-B strains are more prevalent in other parts of the world. For example, clade C predominates in Africa and India and clade E in Southeast Asia. Although some early HIV DNA PCR assays may have been less sensitive in detecting non-B clades [11-13], assays with greater sensitivity for non-B clades have been developed [14].

HIV RNA assays — HIV infection may be identified using either qualitative or quantitative RNA assays. There are a variety of methodologies used to detect HIV RNA including reverse-transcription PCR, b-DNA, transcription-mediated amplification, and nucleic acid sequence-based amplification. For diagnosis, HIV-1 qualitative RNA assays are used in the United States and other resource-rich countries in place of the HIV DNA PCR. Quantitative RNA assays are the standard viral load test performed for monitoring the response to treatment of HIV infection but may be used as a supplemental diagnostic assay to confirm HIV infection.

Compared with DNA PCRs, RNA assays may better detect HIV infection at birth, enabling the early initiation of ART [15], which has been shown to reduce infant morbidity and mortality. Studies have suggested that the sensitivity and specificity of qualitative and quantitative HIV RNA assays for identifying neonatal HIV infection are at least as good as those of HIV DNA testing [8-10,14,16,17]. However, these studies were not performed in children born to mothers who were receiving potent combination ART or in children receiving multidrug prophylaxis. Treatment of either mother or infant with combination ART could be expected to lead to an undetectable viral load measurement and thus a negative HIV-1 RNA test [18]. (See "Techniques and interpretation of HIV-1 RNA quantitation".)

Recommendations for serial testing between birth and four to six months of age allow for diagnostic testing to be done after cessation of antiretroviral (ARV) prophylaxis for infants receiving combination ARV infant prophylaxis if the results of testing were negative while the infant was receiving prophylaxis.

The available HIV-1 RNA assays are capable of sensitive detection of all major HIV-1 genetic variants. (See "Techniques and interpretation of HIV-1 RNA quantitation", section on 'Ability to detect different HIV-1 subtypes'.)

HIV-2 testing — Infants born to individuals with HIV-2 infection should be tested with HIV-2-specific nucleic acid tests (NATs) performed at time points similar to those used for HIV-1 testing. HIV-2 DNA PCR testing can be arranged by the HIV surveillance program of the state or local health department through their public health laboratory or the Centers for Disease Control and Prevention because this assay is not commercially available [7]. (See "Clinical manifestations and diagnosis of HIV-2 infection", section on 'Testing for HIV-2 infection'.)

Other tests — Other tests used to detect HIV viral infection are not recommended in infants and young children:

●HIV-1 peripheral blood cell culture is not recommended, as it is expensive, it is not widely available, and results take a long time (up to four weeks) to become available [5-7,19].

●HIV-1 p24 antigen assays are not recommended in the United States, as they have lower sensitivity compared with other available assays for detecting HIV in infants <6 months of age [6,7].

INTERPRETATION

Confirmed HIV infection — HIV infection in the infant or young child is diagnosed if there is a positive nucleic acid test (NAT) result at any point confirmed with at least one repeat NAT. Infants with confirmed HIV infection should be managed by a pediatric infectious disease specialist. The details of antiretroviral therapy (ART) for pediatric HIV infection are beyond the scope of this topic review. Guidelines on the treatment of pediatric HIV infection are available through the HIV.gov Clinical Info website. (See 'Society guideline links' below.)

Presumptive exclusion of HIV infection — Presumptive exclusion of HIV infection in non-breastfed infants is based on:

●Two or more negative NATs (one obtained at age ≥14 days and one at age ≥4 weeks), or

●One negative NAT at age ≥8 weeks (and at least two weeks after discontinuing multi-drug ARV prophylaxis or presumptive therapy), or

●One negative HIV antibody test at age ≥6 months

To be classified as presumptively uninfected, the infant must have no other laboratory evidence (ie, no positive virologic test results or low CD4 T cell count/percentage) or clinical evidence of HIV and must not be breastfeeding.

Presumptively uninfected infants and breastfed infants with initial negative testing (one obtained at age ≥14 days and one at age ≥4 weeks) do not need Pneumocystis pneumonia (PCP) prophylaxis, assuming the infant does not have other immunosuppression or clinical evidence of HIV infection and the breastfed infant is being monitored as per infant feeding guidelines. (See 'Breastfed infants' above.)

Definitive exclusion of HIV infection — Definitive exclusion of HIV infection in non-breastfed infants is based on [7]:

●Two or more negative NATs (one obtained at ≥1 month of age (and at least two weeks after discontinuing multidrug ARV prophylaxis or presumptive therapy) and one at ≥4 months of age), or

●Two negative antibody tests from separate specimens obtained at ≥6 months of age

As with presumptive exclusion, the infant must have no other laboratory evidence (ie, no positive virologic test results or low CD4 T cell count/percentage) or clinical evidence of HIV and must not be breastfeeding to be classified as definitively uninfected. For breastfed infants, the testing schedule is outlined elsewhere. (See 'Breastfed infants' above.)

Additional HIV testing is not routinely required for non-breastfed infants who meet the criteria for definitive exclusion of HIV and who have had no known or suspected HIV exposure after birth. However, some experts recommend a follow-up antibody test at 18 to 24 months of age to document seroreversion or clearance of maternal antibody and confirm the child's HIV-negative status [7,20].

Indeterminate HIV infection status — HIV-exposed infants who meet neither criteria for having HIV infection nor criteria for exclusion of HIV infection are characterized as having indeterminate HIV status [7]. This includes infants who do not meet the minimum requirement for presumptively uninfected (eg, having one negative test result at four weeks of age).

In addition, some infants may have NAT results that fall in an indeterminate range (also called "weakly positive"). These infants may be characterized as having indeterminate HIV status pending repeat testing. Follow-up testing should be performed as soon as possible. In a systemic review and meta-analysis of 32 studies, indeterminate results accounted for 17 percent of initial non-negative test results, of which three-quarters were negative on repeat testing [21]. These findings highlight the importance of serial testing to establish or exclude the diagnosis of HIV infection in infants born to mothers with HIV.

Infants with indeterminate HIV infection should receive PCP prophylaxis starting at four to six weeks until they are determined to be definitively or presumptively HIV uninfected. PCP prophylaxis is discussed in detail separately. (See "Treatment and prevention of Pneumocystis infection in patients with HIV", section on 'Antimicrobial prophylaxis'.)

NONPERINATAL EXPOSURES — Nonperinatal routes of HIV transmission in infants and children include sexual abuse, transfusion with contaminated blood products, and needlestick with contaminated needles. In these circumstances, testing is recommended regardless of the presence of signs and/or symptoms of HIV infection. When HIV testing is performed in infants <18 months old with HIV exposure from one of these routes, initial testing should be performed with a NAT, as described above (see 'Diagnostic tests' above). Initial testing in infants aged ≥18 months should be performed with an HIV antigen/antibody assay. (See "Screening and diagnostic testing for HIV infection", section on 'Tests'.)

Additional monitoring, including serial HIV testing, may be necessary. In addition, post-exposure prophylaxis may be warranted, depending on the nature of the exposure. Additional information on post-exposure prophylaxis can be found on the Centers for Disease Control and Prevention website.

HIV testing is also appropriate for infants and children with signs and/or symptoms of HIV, even in the absence of documented or suspected HIV exposure.

DIAGNOSIS IN CHILDREN OLDER THAN 18 MONTHS — The diagnosis of HIV infection in children ≥18 months is discussed separately. (See "Screening and diagnostic testing for HIV infection" and "Acute and early HIV infection: Clinical manifestations and diagnosis", section on 'Diagnostic test performance in early HIV infection'.)

SOCIETY GUIDELINE LINKS — Links to society and government-sponsored guidelines from selected countries and regions around the world are provided separately. (See "Society guideline links: HIV infection in infants and children".)

SUMMARY AND RECOMMENDATIONS

●Challenges of HIV testing in infancy – Antibody-only and antibody/antigen combination assays that are commonly used for HIV testing in adults and older children are not reliable in infants and young children, because of the persistence of transplacentally acquired maternal antibody. For this reason, HIV nucleic acid testing (NAT; ie, assays that detect HIV DNA or RNA) must be used to diagnose HIV infection in this population. (See 'Challenges of HIV testing in infancy' above.)

●Approach to testing

•Infants born to mothers with HIV – Infants born to mothers with HIV require serial HIV testing with NATs. The recommended schedule for virologic testing is summarized in the table (table 1). A positive result at any point should be confirmed as soon as possible with a repeat NAT. (See 'Mother with HIV' above.)

Qualitative RNA assays are the most commonly available test in the United States, but HIV DNA polymerase chain reactions (PCRs) remain the preferred test in infants in resource-limited settings. (See 'Diagnostic tests' above.)

•Unknown maternal HIV status – If the maternal HIV status is unknown, serologic testing for HIV should be performed on the mother or infant. If the serologic test is positive, proceed with testing the infant with NAT in the same manner as for the infant of a mother with HIV. (See 'Maternal HIV status unknown' above.)

●Interpretation

•Confirmed HIV infection – HIV infection in the infant or young child is diagnosed if there is a positive NAT result at any point confirmed with at least one repeat NAT. (See 'Confirmed HIV infection' above.)

•Presumptive exclusion of HIV infection – Presumptive exclusion of HIV infection in non-breastfed infants is based on (see 'Presumptive exclusion of HIV infection' above):

-Two negative NATs (one obtained at age ≥14 days and one at age ≥4 weeks), or

-One negative NAT at age ≥8 weeks, or

-One negative HIV antibody test at age ≥6 months

•Definitive exclusion of HIV infection – Definitive exclusion of HIV infection in non-breastfed infants is based on (see 'Definitive exclusion of HIV infection' above):

-Two or more negative NATs (one obtained at ≥1 month of age and one at ≥4 months of age), or

-Two negative antibody tests from separate specimens obtained at ≥6 months of age

-Although it is not required, some experts recommend a follow-up antibody test at 18 to 24 months of age to document seroreversion or clearance of maternal antibody and confirm the child's HIV-negative status

•Intermediate HIV status – HIV-exposed infants who meet neither criteria for having HIV infection nor criteria for exclusion of HIV infection are characterized as having indeterminate HIV status. Infants with NAT results that fall in an indeterminate range (also called "weakly positive") are also included in this category pending repeat testing. Follow-up testing should be performed as soon as possible. (See 'Indeterminate HIV infection status' above.)

Infants with indeterminate HIV status should receive prophylaxis against Pneumocystis pneumonia (PCP) starting at four to six weeks until they are determined to be definitively or presumptively without HIV. PCP prophylaxis is discussed in detail separately. (See "Treatment and prevention of Pneumocystis infection in patients with HIV", section on 'Antimicrobial prophylaxis'.)

ACKNOWLEDGMENT — The UpToDate editorial staff acknowledges Heidi Schwarzwald, MD, MPH, who contributed to an earlier version of this topic review.