HIV This Week

Zhang Q, Wang L, Jiang Y, Fang L, Pan P, Gong S, Yao J, Tang YW, Vermund SH, Jia Y. Early infant HIV-1 diagnosis suitable for resource-limited settings with multiple circulating subtypes: Nested, 3-monoplex DNA PCR on dried blood spots. J Clin Microbiol. 2007; 46(2):721-6.

Early infant diagnosis of HIV-1 infection is complicated by the persistence of maternal antibodies and by diverse HIV-1 subtypes. Zhang and colleagues developed a nested, 3-monoplex HIV-1 DNA polymerase chain reaction (N3M-PCR) assay to detect diverse HIV-1 subtypes in infants born to infected mothers. They optimized the test for use with dried blood spot samples for ease of storage and transport from rural China to central laboratories. Six pairs of primers were designed targeting env, gag, and pol genomes run in three reactions with an analytical sensitivity of 10 copies DNA per reaction to cover nine HIV-1 subtypes A, B, C, D, F, G, CRF01_AE, CRF08_BC, and CRF07_BC. Assay performance was evaluated on 347 dried blood spot specimens from 151 exposed infants in four diverse provinces of China with multiple circulating subtypes. Results were compared with HIV antibody enzyme immunoassay and Western blot confirmation in the infants at >/=18 months of age, or convincing clinical and epidemiologic data for deceased infants. Sensitivity of the N3M-PCR assay was 30.0% (3/10) for infants at 48 hours after birth, 91.7% (11/12) at 1-2 months, and 93.7% (15/16) at 3-6 months of age. Specificity was 100% (94/94) at all three time points. The polymerase chain reaction reproducibility in the three DNA regions was 100% for samples at 48 hours after birth, 96.7% at 1-2 months, and 100% at 3-6 months of age. The HIV-1 DNA N3M-PCR assay on dried blood spots offers a simple and affordable approach for early infant HIV-1 diagnosis in regions with diverse HIV-1 circulating subtypes.

Editors’ note: The numbers of samples tested in this study in China are small but the results are very encouraging. This polymerase chain reaction (PCR) test is detecting the virus, not antibody, and its performance is judged against the presence of antibodies after 18 months of age. Test sensitivity is good by one month when the test is missing up to 10% of infected infants and test specificity is excellent (no false positive results) from 48 hours of life on. Dried blood spots (DBS) require minimal storage facilities because they are stable at room temperature for prolonged periods and can be safely and easily shipped for centralised testing with economies of scale. As for the DBS-ELISA of Patton et al (below), further testing of the DBS-PCR for infant diagnosis is now needed on a larger scale.

The diagnostic accuracy of the modified p24 Ag assay on paediatric dried blood spots was evaluated. Samples analyzed within 6 weeks of collection yielded no false positive results (specificity 100%) and few false negative results (sensitivity 96.5%-98.3%). Laboratory services with limited resources should assess this option for routine infant diagnosis.

Editors’ note: In this South African study, dried blood spot specimens from 147 six-week old babies born to HIV-seropositive mothers and 99 children known to be infected (median age 20 months) were tested with good sensitivity and excellent specificity. Dried blood spots were obtained from capillary blood obtained by heel stick. The test was an ultra sensitive 24 antigen ELISA and specimens were collected on two types of filter paper. Storage with a desiccant conserved test sensitivity. As with the DBS-PCR of Zhang et al (above), testing should proceed in other settings with larger numbers to validate these findings.