Viral sequence analysis of HIV-positive women and their infected children: insight on the timing of infection and on the transmission network.
ABSTRACT: We used high-resolution phylogenetic methods in the context of mother-to-child transmission to obtain information on the timing of the infection and on the transmission network. A total of 33 pol sequences (from maternal peripheral blood, from breast milk, and from plasma of children) belonging to five cases of HIV infant transmission were studied. Using time-scaled phylogeny we were able to estimate that in two cases the transmission occurred after the recommended duration of breastfeeding, supporting a longer, not reported, duration of breastfeeding as a significant factor associated with HIV infant acquisition in this cohort. Among the postnatal infections we were also able to demonstrate that the cell-free virus in breast milk was the most likely population associated with the event of transmission. Our study showed that a coalescent-based model within a Bayesian statistical framework can provide important information that can contribute to optimizing preventive strategies.
Project description:We quantified the relationship between human immunodeficiency virus (HIV) RNA shedding in breast milk, cumulative RNA exposure, and postnatal transmission, relating timing of infection in the infant to estimated total volume of milk exposure.Nested case-control study of 36 infants of HIV-infected mothers. Case patients were infants who acquired HIV infection through breastfeeding from age 6 through 28 weeks, and control subjects were uninfected infants matched on age at obtainment of a breast milk sample. Mothers and infants received peripartum single-dose nevirapine prophylaxis. Feeding data were collected daily; breast milk samples were collected and infant anthropometry was performed at 6 weeks and monthly thereafter. Volume of milk ingested was estimated using infant weight and feeding pattern.Before HIV acquisition in case patients, feeding pattern (exclusive breastfeeding; median duration, 65 vs 70 days; P = .6) and daily milk intake (mean volume, 638 vs 637 mL; P = .97) did not differ significantly between case patients and control subjects. Case mothers were more likely to shed virus (64% vs 9% always, 22% vs 20.5% intermittently, 14% vs 70.5% never shed; overall, P < .001). Case patients ingested ~15 times more HIV-1 RNA particles than did control subjects (196.5 vs 13 × 10? copies; P < .001). Allowing for maternal antenatal CD4 cell count and plasma HIV-1 load, child sex and duration of mixed breastfeeding, the association between HIV RNA exposure and infection remained statistically significant (P < .001).Postnatal acquisition of HIV-1 is more strongly associated with cumulative exposure to cell-free particles in breast milk than with feeding mode. Reducing breast milk viral load through antiretroviral therapy to mother or child can further decrease postnatal transmission in exclusively breastfed infants.
Project description:HIV-1 transmission rates have been reduced over the last decade, an estimated 2 million new infections per year arise, including 220,000 paediatric cases. The main post-natal HIV exposure is through breastfeeding, where both its duration and modality (exclusive or not) are associated with postnatal transmission. The ANRS 12174 trial compared HIV-1 postnatal transmission of 2 prophylaxis drugs for infants during lactation (lamivudine and lopinavir-ritonavir). Our objective has been to examine the feeding practices and the determinants of exclusive/ predominant (EPBF) or any breastfeeding among the participants of this trial in Burkina Faso, South Africa, Uganda and Zambia.Mothers infected with HIV-1 and their uninfected offspring were followed from day 7 after birth for 50 weeks, keeping monthly records of their feeding patterns. Feeding was classified into 3 categories: 1) exclusive breastfeeding during the first six months, only breast-milk being given to infant for 6 months, 2) predominant breastfeeding, breast-milk with liquid-based items being given, and 3) mixed feeding, other non-breast milk or solid food being given in addition to breast milk with or without liquid-based items. The categories were merged into 2 groups: EPBF applying to infants aged <6 months and mixed feeding applying to infants of any age. The feeding patterns have been given as Kaplan-Meier curves. A flexible parametric multiple regression model was used to identify the determinants of the mothers' feeding behaviour.A total of 1,225 mother-infant pairs provided feeding data from Burkina Faso (N?=?204), South Africa (N?=?213), Uganda (N?=?274) and Zambia (N?=?534) between November 2009 and March 2013. The mean maternal age was 27.4 years and the mean BMI was 24.5. 57.7 and 93.9% of mothers initiated breastfeeding within the first hour and first day, respectively. Overall, the median durations of any form of breastfeeding and EPBF were 40.6, and 20.9 weeks, respectively. Babies randomized to the lopinavir/ritonavir group in South Africa tended to do less EPBF than those in the lamivudine group. Overall the group of mothers aged between 25 and 30 years, those married, employed or multiparous tended to stop early EPBF. Mothers living in Uganda or Zambia, those aged between 25 -30 years, better educated (at least secondary school level), employed or having undergone C-section stopped any breastfeeding early.There is a need to improve breastfeeding and complementary feeding practices of children, particularly those exposed to HIV and anti-retrovirals, taking into account context and socio-demographic factors.Clinical trial registration: NCT00640263.
Project description:To estimate the rates and timing of mother to infant transmission of HIV associated with breast feeding in mothers who seroconvert postnatally, and their breast milk and plasma HIV loads during and following seroconversion, compared with women who tested HIV positive at delivery.Prospective cohort study.Urban Zimbabwe.14 110 women and infants enrolled in the Zimbabwe Vitamin A for Mothers and Babies (ZVITAMBO) trial (1997-2001).Mother to child transmission of HIV, and breast milk and maternal plasma HIV load during the postpartum period.Among mothers who tested HIV positive at baseline and whose infant tested HIV negative with polymerase chain reaction (PCR) at six weeks (n=2870), breastfeeding associated transmission was responsible for an average of 8.96 infant infections per 100 child years of breast feeding (95% CI 7.92 to 10.14) and varied little over the breastfeeding period. Breastfeeding associated transmission for mothers who seroconverted postnatally (n=334) averaged 34.56 infant infections per 100 child years (95% CI 26.60 to 44.91) during the first nine months after maternal infection, declined to 9.50 (95% CI 3.07 to 29.47) during the next three months, and was zero thereafter. Among women who seroconverted postnatally and in whom the precise timing of infection was known (≤90 days between last negative and first positive test; n=51), 62% (8/13) of transmissions occurred in the first three months after maternal infection and breastfeeding associated transmission was 4.6 times higher than in mothers who tested HIV positive at baseline and whose infant tested HIV negative with PCR at six weeks. Median plasma HIV concentration in all mothers who seroconverted postnatally declined from 5.0 log(10) copies/mL at the last negative enzyme linked immunosorbent assay (ELISA) to 4.1 log(10) copies/mL at 9-12 months after infection. Breast milk HIV load in this group was 4.3 log(10) copies/mL 0-30 days after infection, but rapidly declined to 2.0 log(10) copies/mL and <1.5 log(10) copies/mL by 31-90 days and more than 90 days, respectively. Among women whose plasma sample collected soon after delivery tested negative for HIV with ELISA but positive with PCR (n=17), 75% of their infants were infected or had died by 12 months. An estimated 18.6% to 20.4% of all breastfeeding associated transmission observed in the ZVITAMBO trial occurred among mothers who seroconverted postnatally.Breastfeeding associated transmission is high during primary maternal HIV infection and is mirrored by a high but transient peak in breast milk HIV load. Around two thirds of breastfeeding associated transmission by women who seroconvert postnatally may occur while the mother is still in the "window period" of an antibody based test, when she would test HIV negative using one of these tests. Trial registration Clinical trials.gov NCT00198718.
Project description:OBJECTIVE:Estimate association between postpartum antiretroviral adherence and breast milk HIV-1 transmission. DESIGN:Prospective cohort study. METHODS:Mother-infant pairs were randomized after delivery to immediately begin receiving 28 weeks of either triple maternal antiretrovirals (zidovudine, lamivudine, and either nevirapine, nelfinavir, or lopinavir-ritonavir) or daily infant nevirapine as part of the Breastfeeding, Antiretrovirals, and Nutrition (BAN) study. Associations between postpartum antiretroviral adherence and rate of breast milk HIV-1 transmission were estimated using Cox models. We measured adherence over four postpartum time intervals using pill count, suspension bottle weight, and maternal self-report. Adherence was categorized and lagged by one interval. Missing adherence measures were multiply imputed. Infant HIV-1 infection was determined by DNA PCR every 2-6 weeks. The primary endpoint was infant HIV-1 infection by 38 weeks of age among infants alive and uninfected at 5 weeks. RESULTS:Analyses included 1479 mother-infant pairs and 45 transmission events. Using pill count and bottle weight information, 22-40% of mother-infant pairs at any given interval were less than 90% adherent. Having at least 90% adherence was associated with a 52% [95% confidence interval (CI) 3-76] relative reduction in the rate of breast milk HIV-1 transmission, compared with having less than 90% adherence when controlling for study arm, breastfeeding status, and maternal characteristics. Complete case analysis rendered similar results (n?=?501; relative reduction 59%, 95% CI 6-82). CONCLUSION:Nonadherence to extended postpartum antiretroviral regimens in 'real world' settings is likely to be higher than that seen in BAN. Identifying mothers with difficulty adhering to antiretrovirals, and developing effective adherence interventions, will help maximize benefits of antiretroviral provision throughout breastfeeding.
Project description:UNLABELLED:Despite the nutritional and health benefits of breast milk, breast milk can serve as a vector for mother-to-child HIV transmission. Most HIV-infected infants acquire HIV through breastfeeding. Paradoxically, most infants breastfed by HIV-positive women do not become infected. This is potentially attributed to anti-HIV factors in breast milk. Breast milk of HIV-negative women can inhibit HIV infection. However, the HIV-inhibitory activity of breast milk from HIV-positive mothers has not been evaluated. In addition, while significant differences in breast milk composition between transmitting and nontransmitting HIV-positive mothers have been correlated with transmission risk, the HIV-inhibitory activity of their breast milk has not been compared. This knowledge may significantly impact the design of prevention approaches in resource-limited settings that do not deny infants of HIV-positive women the health benefits of breast milk. Here, we utilized bone marrow/liver/thymus humanized mice to evaluate the in vivo HIV-inhibitory activity of breast milk obtained from HIV-positive transmitting and nontransmitting mothers. We also assessed the species specificity and biochemical characteristics of milk's in vivo HIV-inhibitory activity and its ability to inhibit other modes of HIV infection. Our results demonstrate that breast milk of HIV-positive mothers has potent HIV-inhibitory activity and indicate that breast milk can prevent multiple routes of infection. Most importantly, this activity is unique to human milk. Our results also suggest multiple factors in breast milk may contribute to its HIV-inhibitory activity. Collectively, our results support current recommendations that HIV-positive mothers in resource-limited settings exclusively breastfeed in combination with antiretroviral therapy. IMPORTANCE:Approximately 240,000 children become infected with HIV annually, the majority via breastfeeding. Despite daily exposure to virus in breast milk, most infants breastfed by HIV-positive women do not acquire HIV. The low risk of breastfeeding-associated HIV transmission is likely due to antiviral factors in breast milk. It is well documented that breast milk of HIV-negative women can inhibit HIV infection. Here, we demonstrate, for the first time, that breast milk of HIV-positive mothers (nontransmitters and transmitters) inhibits HIV transmission. We also demonstrate that breast milk can prevent multiple routes of HIV acquisition and that this activity is unique to human milk. Collectively, our results support current guidelines which recommend that HIV-positive women in resource-limited settings exclusively breastfeed in combination with infant or maternal antiretroviral therapy.
Project description:BACKGROUND: The choice of infant feeding method is important for HIV-positive mothers in order to optimise the chance of survival of their infants and to minimise the risk of HIV transmission. The aim of this study was to investigate feeding practices, including breastfeeding, in the context of PMTCT for infants and children under two years of age born to HIV-positive mothers in Uganda. METHODS: In collaboration with The Aids Support Organisation Mbale, we conducted a cross-sectional survey involving 235 HIV-positive mothers in Uganda. Infant feeding practices, reasons for stopping breastfeeding, and breast health problems were studied. Breastfeeding duration was analysed using the Kaplan-Meier method based on retrospective recall. RESULTS: Breastfeeding was initiated by most of the mothers, but 20 of them (8.5%) opted exclusively for replacement feeding. Pre-lacteal feeding was given to 150 (64%) infants and 65 (28%) practised exclusive breastfeeding during the first three days. One-fifth of the infants less than 6 months old were exclusively breastfed, the majority being complementary fed including breast milk. The median duration of breastfeeding was 12 months (95% confidence interval [CI] 11.5 to 12.5). Adjusted Cox regression analysis indicated that a mother's education, socio-economic status, participation in the PMTCT-program and her positive attitude to breastfeeding exclusively, were all associated with a reduction in breastfeeding duration. Median duration was 3 months (95% CI 0-10.2) among the most educated mothers, and 18 months (95% CI 15.0-21.0) among uneducated mothers. Participation in the PMTCT program and being socio-economically better-off were also associated with earlier cessation of breastfeeding (9 months [95% CI 7.2-10.8] vs. 14 months [95% CI 10.8-17.2] and 8 months [95% CI 5.9-10.1] vs. 17 months [95% CI 15.2-18.8], respectively). The main reasons for stopping breastfeeding were reported as: advice from health workers, maternal illness, and the HIV-positive status of the mother. CONCLUSION: Exclusive breastfeeding was uncommon. Exclusive replacement feeding was practised by few HIV-positive mothers. Well-educated mothers, mothers who were socio-economically better-off and PMTCT-attendees had the shortest durations of breastfeeding. Further efforts are needed to optimise infant feeding counselling and to increase the feasibility of the recommendations.
Project description:Breast milk HIV-1 transmission is currently the predominant contributor to pediatric HIV infections. Yet, only ~10% of breastfeeding infants born to untreated HIV-infected mothers become infected. This study assessed the protective capacity of natural HIV envelope-specific antibodies isolated from the milk of HIV-infected women in an infant rhesus monkey (RM), tier 2 SHIV oral challenge model. To mimic placental and milk maternal antibody transfer, infant RMs were i.v. infused and orally treated at the time of challenge with a single weakly neutralizing milk monoclonal antibody (mAb), a tri-mAb cocktail with weakly neutralizing and ADCC functionalities, or an anti-influenza control mAb. Of these groups, the fewest tri-mAb-treated infants had SHIV detectable in plasma or tissues (2/6, 5/6, and 7/8 animals infected in tri-mAb, single-mAb, and control-mAb groups, respectively). Tri-mAb-treated infants demonstrated significantly fewer plasma transmitted/founder variants and reduced peripheral CD4+ T cell proviral loads at 8 weeks post-challenge compared to control mAb-treated infants. Abortive infection was observed as detectable CD4+ T cell provirus in non-viremic control mAb- and single mAb-, but not in tri-mAb-treated animals. These results suggest that polyfunctional milk antibodies contribute to the natural inefficiency of HIV-1 transmission through breastfeeding and infant vaccinations eliciting non-neutralizing antibody responses could reduce postnatal HIV transmission.
Project description:Intrapartum administration of single-dose nevirapine (sdNVP) reduces perinatal HIV-1 transmission in resource-limiting settings by half. Yet this strategy has limited effect on subsequent breast milk transmission, making the case for new treatment approaches to extend maternal/infant antiretroviral prophylaxis through the period of lactation. Maternal and transmitted infant HIV-1 variants frequently develop NVP resistance mutations following sdNVP, complicating subsequent treatment/prophylaxis regimens. However, it is not clear whether NVP-resistant viruses are transmitted via breastfeeding or arise de novo in the infant.We performed a detailed HIV genetic analysis using single genome sequencing to identify the origin of drug-resistant variants in an sdNVP-treated postnatally-transmitting mother-infant pair. Phylogenetic analysis of HIV sequences from the child revealed low-diversity variants indicating infection by a subtype C single transmitted/founder virus that shared full-length sequence identity with a clonally-amplified maternal breast milk virus variant harboring the K103N NVP resistance mutation.In this mother/child pair, clonal amplification of maternal NVP-resistant HIV variants present in systemic and mammary gland compartments following intrapartum sdNVP represents one source of transmitted NVP-resistant variants that is responsible for the acquisition of drug resistant virus by the breastfeeding infant. This finding emphasizes the need for combination antiretroviral prophylaxis to prevent mother-to-child HIV transmission.
Project description:Exposure of the infant's gut to cell-associated and cell-free HIV-1 trafficking in breast milk (BM) remains a primary cause of mother-to-child transmission (MTCT). The mammary gland represents a unique environment for HIV-1 replication and host-virus interplay. We aimed to explore the origin of the virus transmitted during breastfeeding, and the link with quasi-species found in acellular and cellular fractions of breast-milk (BM) and in maternal plasma. The C2-V5 region of the env gene was amplified, cloned and sequenced from the RNA and DNA of BM, the RNA from the mother's plasma (PLA) and the DNA from infant's dried blood spot (DBS) in 11 post-natal mother-infant pairs. Sequences were assembled in Geneious, aligned in ClustalX, manually edited in SeAL and phylogenetic reconstruction was undertaken in PhyML and MrBayes. We estimated the timing of transmission (ETT) and reconstructed the time for the most recent common ancestor (TMRCA) of the infant in BEAST. Transmission of single quasi-species was observed in 9 of 11 cases. Phylogenetic analysis illustrated a BM transmission event by cell-free virus in 4 cases, and by cell-associated virus in 2 cases but could not be identified in the remaining 5 cases. Molecular clock estimates, of the infant ETT and TMRCA, corresponded well with the timing of transmission estimated by sequential infant DNA PCR in 10 of 11 children. The TMRCA of BM variants were estimated to emerge during gestation in 8 cases. We hypothesize that in the remaining cases, the breast was seeded with a long-lived lineage latently infecting resting T-cells. Our analysis illustrated the role of DNA and RNA virus in MTCT. We postulate that DNA archived viruses stem from latently infected quiescent T-cells within breast tissue and MTCT can be expected to continue, albeit at low levels, should interventions not effectively target these cells.
Project description:A significant number of infants acquire HIV-1 through their infected mother's breast milk, primarily due to limited access to antiretrovirals. Passive immunization with neutralizing antibodies (nAbs) may prevent this transmission. Previous studies, however, have generated conflicting results about the ability of nAbs to halt mother-to-child transmission (MTCT) and their impact on infant outcomes. This study compared plasma neutralizing activity in exposed infants and the infected mothers (n = 63) against heterologous HIV-1 variants and the quasispecies present in the mother. HIV-exposed uninfected infants (HEU) (n = 42), compared to those that eventually acquired infection (n = 21), did not possess higher nAb responses against heterologous envelopes (P = 0.46) or their mothers' variants (P = 0.45). Transmitting compared to nontransmitting mothers, however, had significantly higher plasma neutralizing activity against heterologous envelopes (P = 0.03), although these two groups did not have significant differences in their ability to neutralize autologous strains (P = 0.39). Furthermore, infants born to mothers with greater neutralizing breadth and potency were significantly more likely to have a serious adverse event (P = 0.03). These results imply that preexisting anti-HIV-1 neutralizing activity does not prevent breast milk transmission. Additionally, high maternal neutralizing breadth and potency may adversely influence both the frequency of breast milk transmission and subsequent infant morbidity.IMPORTANCE Passive immunization trials are under way to understand if preexisting antibodies can decrease mother-to-child HIV-1 transmission and improve infant outcomes. We examined the influence of preexisting maternal and infant neutralizing activity on transmission and infant morbidity in a breastfeeding mother-infant cohort. Neutralization was examined against both the exposure strains circulating in the infected mothers and a standardized reference panel previously used to estimate breadth. HIV-exposed uninfected infants did not possess a broader and more potent response against both the exposure and heterologous strains compared to infants that acquired infection. Transmitting, compared to nontransmitting, mothers had significantly higher neutralization breadth and potency but similar responses against autologous variants. Infants born to mothers with higher neutralization responses were more likely to have a serious adverse event. Our results suggest that preexisting antibodies do not protect against breast milk HIV-1 acquisition and may have negative consequences for the baby.