Early Infant Diagnosis of HIV

Dr Jayanthi . P.ElwitigalaConsultant Microbiologist

NSACP

• Worldwide, in 2008, an estimated 430 000 new infections due to HIV occurred in children

• 90% were acquired through mother to- child transmission of HIV.

• Of the 430 000 new infections, between 280 000 and 360 000 were acquired during labourand in the pre-partum period.

• Of the remaining new infections, the majority were acquired during breastfeeding.

HIV transmission from mother to child

• During pregnancy, labor and delivery, or breast feeding is known as perinatal transmission and is the most common route of HIV infection in children.

• The risk of transmission is 25-30%

• Studies suggest that 35% of the infected child die in their 1st year, 50% die by 2nd year and 60% die by 3rd year, in resource limited settings.

• However, women with HIV who take antiretroviral medication during pregnancy as recommended can reduce the risk of transmitting HIV to their babies to less than 1% (PMTCT programmes)

• Early Infant Diagnosis (EID) is crucial to reduce morbidity and mortality in HIV-infected children

• The Children with HIV Early Antiretroviral Therapy (CHER) study showed that mortality among HIV-infected infants randomized to ART within the first 12 weeks of life was significantly reduced compared to those receiving later ART based on clinical criteria as per prior WHO guidelines.

Violari A, Cotton MF, Gibb DM, et al. Early antiretroviral therapy and mortality among HIV-infected infants. N Engl J Med. 2008;359:2233–44.

Recently published data has confirmed dramatic survival benefits for infants started with ART.

• Hence, it is strongly recommended that laboratory testing has to be carried out to diagnose HIV-1 infection in infants below 18 months.

• However, very few infants are gaining access to early diagnosis and for the timely ART .

• Addressing HIV/AIDS in Infants & Children below 18 months is significant global challenge

Who needs testing?

• Two types of infants will need HIV diagnostic testing

- Infants - HIV-exposed (mother known HIV positive)- Infants - sick with signs and symptoms of HIV, even if

unknown HIV exposure status (referred by MO/Pediatrician)

• Identification of infected infants can take place through one of the following:

(If available )– infant diagnosis algorithm used in the country– National pediatric guidelines

Diagnostic tests

• HIV antibody detection assays, used to diagnose HIV infection in adults cannot be used early in life because of the persistence of maternal HIV antibodies acquired by passive transfer.

Clinical Guidelines Portalhttp://aidsinfo.nih.gov/

Diagnosis of HIV Infection in Infants and Children(Last updated: February 12, 2014; last reviewed: February 12, 2014)

Recommendations• Virologic assays that directly detect HIV must be

used to diagnose HIV infection in infants younger than 18 months (AII).

• HIV DNA polymerase chain reaction and HIV RNA assays are recommended as preferred virologicassays (AII).

• Virologic diagnostic testing in infants with known perinatal HIV exposure is recommended at ages 14 to 21 days, 1 to 2 months, and 4 to 6 months (AII).

• Virologic diagnostic testing at birth should be considered for infants at high risk of HIV infection

• Virologic diagnostic testing should be considered 2 to 4 weeks after cessation of antiretroviral (ARV) prophylaxis for infants receiving combination ARV infant prophylaxis, if the results of prior virologic testing were negative while the infant was receiving prophylaxis

• A positive virologic test should be confirmed as soon as possible by a repeat virologic test on a second specimen

WHO

Recommendation 3: • It is strongly recommended that HIV

virological testing be used to diagnose HIV infection in infants and children below 18 months of age.

(Strong recommendation – High quality of evidence)

Recommendation 4: In infants and children undergoing virological

testing, the following assays (and respective specimen types) are strongly recommended for use:

• HIV DNA on whole blood specimen or DBS; • HIV RNA on plasma or DBS; • Us p24 Ag on plasma or DBS.(Strong recommendation – High quality of

evidence)

Recommendation 5:It is strongly recommended that all HIV-exposed

infants have HIV virological testing

at 4–6 weeks of age or at the earliest opportunity thereafter.

• The current test of choice in HIV care and treatment program is the

HIV-1 DNA PCR test which detects HIV-1 Proviral DNA integrated in Human Genome. Sensitivity (99.3%) and Specificity (99.6%)(Shermann et al., 2005 )

• HIV RNA PCR and other nucleic acid detection techniques have also proved accurate and reliable & they provide additional information about virological status that can assist in clinical management

(However, false positive results can occur although rarely as a result of laboratory and operator error.)

HIV Life Cycle : Pre-integration

Fusion

ssRNA

Penetration and Uncoating

dsDNA cDNA Nuclear Transport

Host Cell

Reverse Transcription

Host chromosome

Integration

HIV Life Cycle: Post-integration

Maturation

Budding

Host Cell

Transcription

ssRNA

mRNA

ProteinsAssembly

Translation

DBS

• With the development of dried blood spot (DBS) method to simplify sample management and transport, polymerase chain reaction (PCR) testing of infants has become feasible in rural settings using centralized laboratories. The use of DBS-PCR methods in infants has been demonstrated to be both highly sensitive and specific, enabling EID in even resource-limited settings

DBS• The child is pricked and blood is collected on

a filter paper card - Dried Blood Spot card (DBS).

6 wks age to 4 months - heel prick4 months to 10 months – big toe prick

10 months to 18 months - finger prick (depending on health of the child)

• This card is made with a special Filter paper called Whatmann Filter Paper 903.

HIV DNA PCR

• HIV DNA PCR is a sensitive technique used to detect specific HIV viral DNA in peripheral blood. Good accuracy in whole blood and DBS in almost all circumstances.

• mononuclear cells. The specificity of the HIV DNA PCR is 99.8% at birth and 100% at 1, 3, and 6 months.

• The sensitivity of the test performed at birth is 55% but increases to more than 90% by 2 to 4 weeks of age,

• and 100% at ages 3 months and 6 months.6-9

RNA assays

• Good accuracy; however, there are concerns if the infant is on ART because of the reduced amount of detectable RNA.

• Branched-DNA (b-DNA) assays: lower specificity (around 97%) and are therefore associated with more FP results.

• Us p24 Ag: good accuracy; however, there are concerns if the infant is on ART because of the reduced amount of detectable p24 Ag.

POC

• There are no Point of Care (POC) tests currently available for EID testing, although several are under development. The most effective placement of the POC instruments to optimize HIV diagnosis has not yet been determined. It is anticipated that technologies in the pipeline will have lower instrument and per-test costs

Summary of recommended testing approaches

Category Test required Purpose Action

Well, HIV-exposedinfant

Virological testing at 4–6 weeks of age

To diagnose HIV Start ART if HIV-infected

Infant – unknownHIV exposure

Maternal HIV serological test orinfant HIV serological test

To identify or confirm HIV exposure

Need virological test if HIV-exposed

Well, HIV-exposedinfant at 9 months

HIV serological test (at last immunization, usually 9 months)

To identify infants who have persisting HIV antibody or have seroreverted

HIV seropositive need virological test and continued follow up; HIV negative, assume uninfected, repeat testing required if still breastfeeding

Category Test required Purpose Action

Infant or child withsigns andsymptomssuggestive of HIV

HIV serological test To confirm exposure

Perform virologicaltest if <18 monthsof age

Well or sick childseropositive>9 months and<18 months

Virological testing To diagnose HIV Reactive – start HIVcare and based on nationalstart criteria

Infant or child whohas completelydiscontinuedbreastfeeding

Repeat testing sixweeks or more afterbreastfeedingcessation – usuallyinitial HIV serologicaltesting followed byvirological testing forHIV-positive childand <18 monthsof age

To exclude HIVinfection afterexposure ceases

Infected infantsand children<24 months of age,need to start HIVcare, including ARTb

• While early treatment is known to dramatically decrease morbidity and mortality from HIV infection,

• In 2011 out of the estimated 3.4 million children living with HIV globally, 230,000 died from AIDS-related illnesses

• Clearly there is more work to be done to identify HIV infected infants earlier and initiate lifesaving treatment.

• An effective EID program is the first step to ensuring the survival of this vulnerable population.

EID Challenges

• An effective EID service should be able to:Identify all HIV-exposed infants and their mothers, provide appropriate HIV testing services

• Retain HIV-exposed infants and their mothers in care to ensure the mother-infant pair successfully navigates the EID

• cascade to the point of a definitive diagnosis, without being lost to follow-up

• Identify all HIV-infected infants and link them to treatment services to ensure timely initiation of ART.

The way forwardLaboratory networking• Laboratories are designated as national, regional based on

the types of samples they can collect. • Forming an EID- laboratory network in each country is

essential. • An EID-laboratory network should be built in two

dimensions.• In one dimension, the EID- laboratory network should be

embedded within the national laboratory tiered system in which the national reference laboratory supports regional EID

• Regional laboratories should then support the hospital EID laboratories by providing technical assistance.

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