The overall goal of this proposal is to improve methods for the early diagnosis of HIV infection in infants born to HIV seropositive mothers and to better define intrauterine vs. intrapartum acquisition of infection in a prospective longitudinal study of mothers and infants. We will assess virological and immunological parameters of risk of either intrauterine or intrapartum infection in mother/infant pairs by correlating the following maternal parameters (specific HIV-AB including AB to GP120, GP41 and HIV regulatory proteins, plasma, viremia, presence of virus in amniotic or cervical/vaginal fluid at delivery and detection of HIV1 virus in the placenta) with the earliest diagnosis of infection in the infant. We will assess whether HIV viral antigens (P24,TAT,VIF, and NEF) cross the placenta an correlate with immune response or infection in the infant. We will also assess placenta or aborted fetal tissue for evidence of HIV infection by PCR and in situ hybridization assays. The proposed innovative diagnostic studies are aimed at establishing unequivocally as quickly as possible an infant as HIV infected. We have adapted to a micro scale for babies a series of assays designed directly to detect HIV1 virus or its products including quantitative HIV PCR-DNA and PCR-RNA in situ hybridization DNA and RNA in PBL compared to quantitative co-culture, plasma culture, and P24 antigen. We proposed a unique set of assays for three structural proteins HIV-nef-vif-tat which include antigen and antibody assays to be compared in mother and baby pairs for discrepant results indicating newborn infection. Because of transplacental IgG antibody, immunological responses unique to the infant (in vitro production HIV IgG-IgM and IgM IgD IgE and IgA antibody response to specific HIV proteins) will be assessed serially. All children will be followed for 2 years to ultimately define infection and statistical analysis of individual and combinations of tests which will accurately predict a baby infected or not infected will be done. We propose a unique pilot study using the SCID-hu mouse model to assess the ability of this model to amplify either HIV virus replication and/or specific HIV1 antibody production from injected human neonatal cells. Using these micro assays we have the unique ability to sequentially follow mother and infant to not only identify infection early but also better define the timing of virus acquisition in the fetus/infant.
