The University of Zimbabwe Birth Cohort Study (UZBCS) stands as a crucial initiative in understanding the intricate relationship between maternal health and infant development, far removed from the complexities of automotive key programming codes like 01494. This detailed investigation delves into how maternal health conditions, including persistent viral infections such as HIV, CMV, and hepatitis B, alongside other infectious and non-communicable diseases and nutritional status, impact pregnancy outcomes, infant mortality, development, immunity, and overall health. Notably, approximately half of the pregnant women participating in the UZBCS are HIV-infected, making this study a vital resource for understanding and addressing maternal and infant health in regions with high HIV prevalence. Upon enrollment, each woman engaged in a comprehensive structured questionnaire, providing extensive clinical, socio-demographic, environmental, and household details. Furthermore, the study meticulously recorded socio-economic factors such as employment status, family income, food budgets, and household food security. For HIV-infected participants, data on the initiation and adherence to combination antiretroviral therapy (cART) were also documented. Aligning with Zimbabwean national guidelines advocating lifelong cART for all HIV-infected pregnant women (Option B+), the study acknowledges the practical challenges of timely HIV diagnosis during pregnancy. The established standard of care in Zimbabwe for preventing mother-to-child transmission of HIV involves non-nucleoside reverse transcriptase inhibitors, specifically TENOLAM-E (Tenofovir, Lamivudine, and Efavirenz). Mothers are encouraged to breastfeed for as long as they choose, with exclusive breastfeeding promoted for the first six months of the infant’s life.
Infants exposed to HIV are initiated on Cotrimoxazole prophylaxis until breastfeeding ceases or they test positive for HIV-PCR, whichever comes first. Upon confirmed HIV-1 infection, infants are promptly started on cART, typically including protease inhibitors like Lopinavir/ritonavir and nucleoside reverse transcriptase inhibitors, Zidovudine and Lamivudine.
The UZBCS enrolled 1200 pregnant women – 600 HIV-infected and 600 HIV-uninfected – commencing at or after 20 weeks of gestation, from February 2016 to June 2019. The mother-infant pairs are being monitored for two years, with the study anticipated to conclude in June 2021.
Participant Selection for the UZBCS Cohort
Potential participants were identified during routine antenatal care appointments at four of the twelve City of Harare Polyclinics. These clinics—Kuwadzana, Rujeko, Budiriro, and Glenview—are located in the South-western high-density suburbs of Harare, areas characterized by lower socio-economic conditions. Follow-up assessments for mothers and infants were conducted at delivery, within 10 days postpartum, and subsequently at 6, 10, 14, and 24 weeks postpartum. Data collected at these follow-up points included the date and mode of delivery, birth weight and length, feeding practices, and comprehensive information on the infants’ overall health and developmental milestones.
Focus Group for CMV Analysis
From the initial 527 mothers recruited between February 2016 and August 2017, 280 of whom were HIV-infected, data and biosamples were obtained. Due to funding constraints, Cytomegalovirus (CMV)-DNAemia measurements were limited to a subset of pregnant women. The primary selection criterion included all mothers who transmitted HIV to their infants by six months of age (n=11), categorized as MTCT cases. For comparison, 120 HIV-infected mothers who did not transmit HIV were randomly chosen as controls using a computer-generated pseudo-random number algorithm. Notably, three of these control mothers had twin births. Depending on the specific analytical objectives, this control group is referred to as either 120 HIV-infected non-transmitting mothers or 123 infant-mother dyads. An additional control group of 46 HIV-uninfected mothers and their infants were also randomly selected using the same methodology.
Procedures for Blood Collection and Analysis
At the time of enrollment, ten milliliters of venous blood were collected from each mother. Maternal HIV diagnosis was performed using rapid qualitative immunochromatographic assays, specifically the SD Bioline HIV-1/2 3.0 (Standard Diagnostics Inc., Kyonggi-do, South Korea) and Abbott’s Determine® HIV–1/2*. Western Blot testing was employed to resolve any indeterminate results.
Full blood counts were determined from whole ethylenediaminetetraacetic acid (EDTA) blood samples using a Mindray© Haematology 3-part differential, 16 parameters BC3600 Analyzer (Shenzhen, China). The World Health Organization (WHO) definition was utilized for diagnosing anemia in pregnancy (hemoglobin < 11 g/dL [21]).
Absolute CD4+ T-lymphocyte counts in EDTA blood samples from HIV-infected mothers were enumerated within 6 hours of sample collection using a Partec Cyflow counter (Cyflow, Partec, Munster, Germany).
Detection and Quantification of Maternal CMV/HIV
For analyzing HIV-RNA and CMV-DNA loads, blood samples were centrifuged at 3000 rpm for 5 minutes, and the plasma was aliquoted into labeled cryo-vials and stored at -80°C until testing.
Total CMV nucleic acids were extracted from 200 µL of plasma using the QIAamp MinElute Virus Spin Kit (Qiagen, Hilden, Germany). Subsequently, 60 µL of total viral nucleic acids, including CMV-DNA, were eluted and immediately stored at -80°C for further analysis. CMV-DNAemia detection and quantification were conducted using quantitative PCR (qPCR) with the RealStar CMV PCR kit v1.0 (Altona Diagnostics, Hamburg, Germany). The QuantStudio 3 Real-Time PCR System (Applied Biosystems, CA) was used for CMV-DNA quantification. The detection limit was established at 1 copy per mL.
For HIV viral load quantification, nucleic acids were extracted from 1 mL of maternal baseline plasma, and HIV-1-RNA was quantified using an automated TaqMan Roche Amplicor 1.5 Monitor Test (Cobas AmpliPrep/Cobas TaqMan, Roche Diagnostics, Branchburg NJ), following the manufacturer’s instructions. The detection limit for HIV-1-RNA was 20 copies per mL.
Assessment of Adverse Birth Outcomes
Adverse birth outcomes considered in the study included preterm birth (delivery before 37 weeks of gestation), low birth weight (less than 2500g), and small for gestational age infants (birth weight below the 10th percentile for gestational age and sex based on INTERGROWTH-21st standards).
Early Infant HIV Diagnosis
Venous blood (EDTA) or heel prick blood spots were collected and preserved on 903 protein saver cards (number 105311018) at delivery, 10 days, and 6, 10, 14, and 24 weeks postpartum. Every infant blood sample from each study visit was processed and stored as dried blood spots and plasma at -20°C and -80°C, respectively, for future assessments, including HIV and CMV evaluations. Infant HIV infection was detected using a qualitative 1.5 Roche Amplicor HIV-1 proviral DNA PCR kit (Roche Diagnostics Incorporation, Branchburg, New Jersey).
Data Management and Statistical Analysis
Data were entered and managed using Research Electronic Data Capture (REDCap v 8.0, © 2020); accessible at http://www.redcap.uzchs.ac.zw/redcap/. Data entry accuracy was ensured through independent double entries and verification to resolve any discrepancies.
Statistical comparisons between groups (HIV-infected/uninfected, CMV-DNA positive/negative) were performed using the Kruskal Wallis test, Mann–Whitney U test, Chi-squared test, and Fisher exact test, as appropriate. For serology positive results, undetectable levels of HIV-1-RNA load or CMV-DNA were assigned a value of zero. Multivariate linear and logistic regression analyses were conducted to test the following predictors at enrollment: CMV-DNAemia > 50 copies per mL, HIV-1-RNA load, years since HIV diagnosis, duration of ongoing cART use, and maternal age.
For infant outcomes, all infants, including twins, were included, with maternal parameters applied to each twin. For maternal outcomes, such as predicting maternal HIV-1-RNA load, mothers with twin deliveries were counted once.
Automated parameter elimination for Akaike information criterion optimization was performed using the “glmulti” R package. Due to statistical power limitations, interactions between predictors were not considered. All calculations were performed in R Studio 1.2.5001.
