P20-12. Heterogeneity of Gag mutational pathways in primary HIV-1 subtype C infection

Poster presentatio

Birth Weight for Gestational Age Norms for a Large Cohort of Infants Born to HIV-Negative Women in Botswana Compared with Norms for U.S.-Born Black Infants

Background: Standard values for birth weight by gestational age are not available for sub-Saharan Africa, but are needed to evaluate incidence and risk factors for intrauterine growth retardation in settings where HIV, antiretrovirals, and other in utero exposures may impact birth outcomes. Methods: Birth weight data were collected from six hospitals in Botswana. Infants born to HIV-negative women between 26-44 weeks gestation were analyzed to construct birth weight for gestational age charts. These data were compared with published norms for black infants in the United States. Results: During a 29 month period from 2007-2010, birth records were reviewed in real-time from 6 hospitals and clinics in Botswana. Of these, 11,753 live infants born to HIV-negative women were included in the analysis. The median gestational age at birth was 39 weeks (1st quartile 38, 3rd quartile 40 weeks), and the median birth weight was 3100 grams (1st quartile 2800, 3rd quartile 3400 grams). We constructed estimated percentile curves for birth weight by gestational age which demonstrate increasing slope during the third trimester and leveling off beyond 40 weeks. Compared with black infants in the United States, Botswana-born infants had lower median birth weight for gestational age from weeks 37 through 42 (p < .02). Conclusions: We present birth weight for gestational age norms for Botswana, which are lower at term than norms for black infants in the United States. These findings suggest the importance of regional birth weight norms to identify and define risk factors for higher risk births. These data serve as a reference for Botswana, may apply to southern Africa, and may help to identify infants at risk for perinatal complications and inform comparisons among infants exposed to HIV and antiretrovirals in utero

High Prevalence of Hypertension and Placental Insufficiency, but No In Utero HIV Transmission, among Women on HAART with Stillbirths in Botswana

Background: Increased stillbirth rates occur among HIV-infected women, but no studies have evaluated the pathological basis for this increase, or whether highly active antiretroviral therapy (HAART) influences the etiology of stillbirths. It is also unknown whether HIV infection of the fetus is associated with stillbirth. Methods: HIV-infected women and a comparator group of HIV-uninfected women who delivered stillbirths were enrolled at the largest referral hospital in Botswana between January and November 2010. Obstetrical records, including antiretroviral use in pregnancy, were extracted at enrollment. Verbal autopsies; maternal HIV, CD4 and HIV RNA testing; stillbirth HIV PCR testing; and placental pathology (blinded to HIV and treatment status) were performed. Results: Ninety-nine stillbirths were evaluated, including 62 from HIV-infected women (34% on HAART from conception, 8% on HAART started in pregnancy, 23% on zidovudine started in pregnancy, and 35% on no antiretrovirals) and 37 from a comparator group of HIV-uninfected women. Only 2 (3.7%) of 53 tested stillbirths from HIV-infected women were HIV PCR positive, and both were born to women not receiving HAART. Placental insufficiency associated with hypertension accounted for most stillbirths. Placental findings consistent with chronic hypertension were common among HIV-infected women who received HAART and among HIV-uninfected women (65% vs. 54%, p = 0.37), but less common among HIV-infected women not receiving HAART (28%, p = 0.003 vs. women on HAART). Conclusions: In utero HIV infection was rarely associated with stillbirths, and did not occur among women receiving HAART. Hypertension and placental insufficiency were associated with most stillbirths in this tertiary care setting

Strengthening Healthcare Capacity Through a Responsive, Country-Specific, Training Standard: The KITSO AIDS Training Program’s Sup-port of Botswana’s National Antiretroviral Therapy Rollout

In parallel with the rollout of Botswana’s national antiretroviral therapy (ART) program, the Botswana Ministry of Health established the KITSO AIDS Training Program by entering into long-term partnerships with the Botswana–Harvard AIDS Institute Partnership for HIV Research and Education and others to provide standardized, country-specific training in HIV/AIDS care. The KITSO training model has strengthened human capacity within Botswana’s health sector and been indispensable to successful ART rollout. Through core and advanced training courses and clinical mentoring, different cadres of health care workers have been trained to provide high-quality HIV/AIDS care at all ART sites in the country. Continuous and standardized clinical education will be crucial to sustain the present level of care and successfully address future treatment challenges

Linear Growth Faltering Among HIV-Exposed Uninfected Children

Background: HIV-exposed uninfected (HEU) children experience increased mortality compared wit their HIV-unexposed uninfected (HUU) peers. It is unclear whether HEU children are also at increased risk for undernutrition, a modifiable risk factor for mortality. Methods: We conducted a cross-sectional, population-based survey of children under 5 years of age in five health districts in Botswana. Linear mixed-effects models were used to assess continuous outcomes while generalized estimating equations were used to estimate relative risks of stunting, wasting, and underweight between HEU (n=396) and HUU (n=1,109) children. Secondary analyses examined potential mediation by low birthweight. Results: The association between maternal HIV-exposure and child stunting varied significantly by child age (p<0.01). HEU children <1 year and ≥2 years of age had 1.85 (95% CI: 1.03-3.31; p=0.04) and 1.41 (95% CI: 1.06-1.88; p=0.02) times the risk of stunting compared with HUU children after multivariate adjustment, respectively. During the period of 1-2 years of age, when breastfeeding cessation occurred among HUU children, HUU children had increased risk of stunting compared with HEU children who were predominantly formula fed (RR: 1.56; 95% CI: 1.05-2.32; p=0.03). A mediation analysis estimated 67% of the excess risk of stunting among HEU children ≥2 years was attributable to low birthweight (p=0.02). There was no difference in risk of wasting or underweight. Conclusion: HEU children are at increased risk of stunting compared with their HUU peers; however, interventions to increase birthweight may significantly ameliorate this excess risk. Interventions to support optimal growth during weaning are needed for all breastfed children

Maternal weight and birth outcomes among women on antiretroviral treatment from conception in a birth surveillance study in Botswana.

Antiretrovirals such as dolutegravir (DTG) and tenofovir alafenamide (TAF) have been associated with excessive weight gain. The objective of this study was to understand the potential impact of ART-associated weight gain on pregnancy outcomes among women living with HIV. Using data from the Tsepamo birth outcomes surveillance study in Botswana, we evaluated the relationship between maternal weight (and weight gain) and severe birth outcomes (very preterm delivery  4000 g) and maternal hypertension. We estimated the relative risk of each outcome by baseline weight (first weight in pregnancy 90 kg) was associated with increased risk of macrosomia (aRR 3.24, 95% CI 2.36, 4.44) and maternal hypertension (aRR 1.79, 95% CI 1.62, 1.97). Baseline weight was not associated with stillbirth or early neonatal death. For all outcomes, second trimester weight gain showed weaker associations than did baseline weight. Duration of pre-pregnancy ART (years) was associated with higher baseline weight for DTG but not for EFV, and the risk of maternal hypertension by baseline weight category was higher for DTG than EFV for all strata. ART regimens associated with weight gain may reduce the number of women at risk for certain severe adverse pregnancy outcomes associated with low weight but increase the number at risk of macrosomia and maternal hypertension. Further research could determine whether weight-based ART treatment strategies improve maternal and child health. [Abstract copyright: © 2021 The Authors. Journal of the International AIDS Society published by John Wiley & Sons Ltd on behalf of International AIDS Society.

Evaluation of Phylogenetic Methods for Inferring the Direction of Human Immunodeficiency Virus (HIV) Transmission: HIV Prevention Trials Network (HPTN) 052

Background: Phylogenetic analysis can be used to assess human immunodeficiency virus (HIV) transmission in populations. We inferred the direction of HIV transmission using whole-genome HIV sequences from couples with known linked infection and known transmission direction. Methods: Complete next-generation sequencing (NGS) data were obtained for 105 unique index-partner sample pairs from 32 couples enrolled in the HIV Prevention Trials Network (HPTN) 052 study (up to 2 samples/person). Index samples were obtained up to 5.5 years before partner infection; partner samples were obtained near the time of seroconversion. The bioinformatics method, phyloscanner, was used to infer transmission direction. Analyses were performed using samples from individual sample pairs, samples from all couples (1 sample/person; group analysis), and all available samples (multisample group analysis). Analysis was also performed using NGS data from defined regions of the HIV genome (gag, pol, env). Results: Using whole-genome NGS data, transmission direction was inferred correctly (index to partner) for 98 of 105 (93.3%) of the individual sample pairs, 99 of 105 (94.3%) sample pairs using group analysis, and 31 of the 32 couples (96.9%) using multisample group analysis. There were no cases where the incorrect transmission direction (partner to index) was inferred. The accuracy of the method was higher with greater time between index and partner sample collection. Pol region sequences performed better than env or gag sequences for inferring transmission direction. Conclusions: We demonstrate the potential of a phylogenetic method to infer the direction of HIV transmission between 2 individuals using whole-genome and pol NGS data

Timing Constraints of In Vivo Gag Mutations during Primary HIV-1 Subtype C Infection

Background: Aiming to answer the broad question “When does mutation occur?” this study examined the time of appearance, dominance, and completeness of in vivo Gag mutations in primary HIV-1 subtype C infection. Methods: A primary HIV-1C infection cohort comprised of 8 acutely and 34 recently infected subjects were followed frequently up to 500 days post-seroconversion (p/s). Gag mutations were analyzed by employing single-genome amplification and direct sequencing. Gag mutations were determined in relation to the estimated time of seroconversion. Time of appearance, dominance, and completeness was compared for different types of in vivo Gag mutations. Results: Reverse mutations to the wild type appeared at a median (IQR) of 62 (44;139) days p/s, while escape mutations from the wild type appeared at 234 (169;326) days p/s (p&lt;0.001). Within the subset of mutations that became dominant, reverse and escape mutations appeared at 54 (30;78) days p/s and 104 (47;198) days p/s, respectively (p&lt;0.001). Among the mutations that reached completeness, reverse and escape mutations appeared at 54 (30;78) days p/s and 90 (44;196) days p/s, respectively (p = 0.006). Time of dominance for reverse mutations to and escape mutations from the wild type was 58 (44;105) days p/s and 219 (90;326) days p/s, respectively (p&lt;0.001). Time of completeness for reverse and escape mutations was 152 (100;176) days p/s and 243 (101;370) days p/s, respectively (p = 0.001). Fitting a Cox proportional hazards model with frailties confirmed a significantly earlier time of appearance (hazard ratio (HR): 2.6; 95% CI: 2.3–3.0), dominance (4.8 (3.4–6.8)), and completeness (3.6 (2.3–5.5)) of reverse mutations to the wild type Gag than escape mutations from the wild type. Some complex mutational pathways in Gag included sequential series of reversions and escapes. Conclusions: The study identified the timing of different types of in vivo Gag mutations in primary HIV-1 subtype C infection in relation to the estimated time of seroconversion. Overall, the in vivo reverse mutations to the wild type occurred significantly earlier than escape mutations from the wild type. This shorter time to incidence of reverse mutations remained in the subsets of in vivo Gag mutations that reached dominance or completeness

Can Directionality of HIV Transmission be Predicted by Next-Generation Sequencing Data?

Background: We evaluated use of phylogenetic methods to predict the direction of human immunodeficiency virus (HIV) transmission. Methods: For 33 pairs of HIV-infected patients (hereafter, "index patients") and their partners who acquired genetically linked HIV infection during the study, samples were collected from partners and index patients close to the time when the partner seroconverted (hereafter, "SC samples"); for 31 pairs, samples collected from the index patient at an earlier time point (hereafter, "early index samples") were also available. Phylogenies were inferred using env next-generation sequences (1 tree per pair/subtype). The direction of transmission (DoT) predicted from each tree was classified as correct or incorrect on the basis of which sequences (those from the index patient or the partner) were closest to the root. DoT was also assessed using maximum parsimony to infer ancestral node states for 100 bootstrap trees. Results: DoT was predicted correctly for both single-pair and subtype-specific trees in 22 pairs (67%) by using SC samples and in 23 pairs (74%) by using early index samples. DoT was predicted incorrectly for 4 pairs (15%) by using SC or early index samples. In the bootstrap analysis, DoT was predicted correctly for 18 pairs (55%) by using SC samples and for 24 pairs (73%) by using early index samples. DoT was predicted incorrectly for 7 pairs (21%) by using SC samples and for 4 pairs (13%) by using early index samples. Conclusions: Phylogenetic methods based solely on the tree topology of HIV env sequences, particularly without consideration of phylogenetic uncertainty, may be insufficient for determining DoT

Comparison of Hepatitis B Virus Infection in HIV-Infected and HIV-Uninfected Participants Enrolled in a Multinational Clinical Trial: HPTN 052

Objective: Data comparing hepatitis B virus (HBV) infection in HIV-infected [HIV(+)], and HIV-uninfected [HIV(2)] individuals recruited into the same study are limited. HBV infection status and chronic hepatitis B (cHB) were characterized in a multinational clinical trial: HIV Prevention Trials Network (HPTN 052). Method: HBV infection status at enrollment was compared between HIV(+) (N = 1241) and HIV(-) (N = 1232) from 7 HBV-endemic countries. Hepatitis B e antigen and plasma HBV DNA were determined in cHB. Median CD4, median plasma HIV RNA, and prevalence of transaminase elevation were compared in HIV(+) with and without cHB. Significance was assessed with x2 Fisher exact and median tests. Results: Among all participants, 33.6% had HBV exposure without cHB (8.9% isolated HBV core antibody, "HBcAb"; 24.7% HBcAb and anti-HB surface antibody positive, "recovered"), 4.3% had cHB, 8.9% were vaccinated, and 53.5% were uninfected. Data were similar among HIV(+) and HIV(2) except for isolated HBcAb, which was more prevalent in HIV(+) than HIV(2) [10.1% vs. 7.7%, P = 0.046]. Median HBV DNA trended higher in HIV(+) than in HIV(2). In HIV (+) with cHB versus those without cHB, transaminase elevations were more prevalent (alanine aminotransferase # grade 2, 12% vs. 5.2%, P = 0.037; aspartate aminotransferase # grade 2, 26% vs. 6.0%, P, 0.001), CD4 trended lower, and HIV RNA was similar. Conclusions: HBV infection status did not differ by HIV infection status. HIV co-infection was associated with isolated HBcAb and a trend of increased HBV DNA. In HIV, cHB was associated with mild transaminase elevations and a trend toward lower CD4