Intra-host sequence variability in human papillomavirus.

Human papillomaviruses (HPVs) co-evolve slowly with the human host and each HPV genotype displays epithelial tropisms. We assessed the evolution of intra HPV genotype variants within samples, and their association to anogenital site, cervical cytology and HIV status. Variability in the L1 gene of 35 HPV genotypes was characterized phylogenetically using maximum likelihood, and portrayed by phenotype. Up to a thousand unique variants were identified within individual samples. In-depth analyses of the most prevalent genotypes, HPV16, HPV18 and HPV52, revealed that the high diversity was dominated by a few abundant variants. This suggests high intra-host mutation rates. Clades of HPV16, HPV18 and HPV52 were associated to anatomical site and HIV co-infection. Particularly, we observed that one HPV16 clade was specific to vaginal cells and one HPV52 clade was specific to anal cells. One major HPV52 clade, present in several samples, was strongly associated with cervical neoplasia. Overall, our data suggest that tissue tropism and HIV immunosuppression are strong shapers of HPV evolution.

Geographical distribution and risk association of human papillomavirus genotype 52-variant lineages.

Human papillomavirus (HPV) genotype 52 is commonly found in Asian cases of cervical cancer but is rare elsewhere. Analysis of 611 isolates collected worldwide revealed a remarkable geographical distribution, with lineage B predominating in Asia (89.0% vs 0%-5.5%; P(corrected) < .001), whereas lineage A predominated in Africa, the Americas, and Europe. We propose that the name "Asian lineage" be used to denote lineage B, to signify this feature. Preliminary analysis suggested a higher disease risk for lineage B, although ethnogeographical confounders could not be excluded. Further studies are warranted to verify whether the reported high attribution of disease to HPV52 in Asia is due to the high prevalence of lineage B.

Geographical distribution and oncogenic risk association of human papillomavirus type 58 E6 and E7 sequence variations.

Human papillomavirus (HPV) 58 accounts for a notable proportion of cervical cancers in East Asia and parts of Latin America, but it is uncommon elsewhere. The reason for such ethnogeographical predilection is unknown. In our study, nucleotide sequences of E6 and E7 genes of 401 HPV58 isolates collected from 15 countries/cities across four continents were examined. Phylogenetic relationship, geographical distribution and risk association of nucleotide sequence variations were analyzed. We found that the E6 genes of HPV58 variants were more conserved than E7. Thus, E6 is a more appropriate target for type-specific detection, whereas E7 is more appropriate for strain differentiation. The frequency of sequence variation varied geographically. Africa had significantly more isolates with E6-367A (D86E) but significantly less isolates with E6-203G, -245G, -367C (prototype-like) than other regions (p ≤ 0.003). E7-632T, -760A (T20I, G63S) was more frequently found in Asia, and E7-793G (T74A) was more frequent in Africa (p < 0.001). Variants with T20I and G63S substitutions at E7 conferred a significantly higher risk for cervical intraepithelial neoplasia grade III and invasive cervical cancer compared to other HPV58 variants (odds ratio = 4.44, p = 0.007). In conclusion, T20I and/or G63S substitution(s) at E7 of HPV58 is/are associated with a higher risk for cervical neoplasia. These substitutions are more commonly found in Asia and the Americas, which may account for the higher disease attribution of HPV58 in these areas.

Predictors of attrition among children born in a PMTCT programme in Zimbabwe followed up over 5 years.

Eliminating of paediatric HIV within prevention of mother to child transmission (PMTCT) interventions rests on complete follow-up of all children. We report on predictors of child attrition in the PMTCT cascade over 5 years where 1050 pregnant women were enrolled at 36 gestational weeks. Mother and child pairs were followed up at birth, 6 weeks, 4 months, 9 months, and every 6 months thereafter for 60 months. Higher attrition was observed for children of economically advantaged, socially stable mothers regardless of HIV status, whereas compliance was observed for children whose mothers tested positive for HIV-1, HSV-2 and Syphilis. Low birthweight was associated with attrition regardless of maternal HIV status. Five years predictors of attrition did not differ by maternal HIV status, as HIV-exposed children succumbed to mortality and those not exposed were loss to follow-up (LFU). Child follow-up is influenced more by maternal lifestyle and health risks leading to retention of high-risk children in PMTCT programmes.

Identification of human papillomavirus type 58 lineages and the distribution worldwide.

BACKGROUND: Human papillomavirus type 58 (HPV-58) accounts for a much higher proportion of cervical cancers in East Asia than other types. A classification system of HPV-58, which is essential for molecular epidemiological study, is lacking. METHODS AND RESULTS: This study analyzed the sequences of 401 isolates collected from 15 countries and cities. The 268 unique concatenated E6-E7-E2-E5-L1-LCR sequences that comprised 57% of the whole HPV-58 genome showed 4 distinct clusters. L1 and LCR produced tree topologies that best resembled the concatenated sequences and thus are the most appropriate surrogate regions for lineage classification. Moreover, short fragments from L1 (nucleotides 6014-6539) and LCR (nucleotides 7257-7429 and 7540-52) were found to contain sequence signatures informative for lineage identification. Lineage A was the most prevalent lineage across all regions. Lineage C was more frequent in Africa than elsewhere, whereas lineage D was more prevalent in Africa than in Asia. Among lineage A variants, sublineage A2 dominated in Africa, the Americas, and Europe, but not in Asia. Sublineage A1, which represents the prototype that originated from a patient with cancer, was rare worldwide except in Asia. CONCLUSIONS: HPV-58 can be classified into 4 lineages that show some degree of ethnogeographic predilection in distribution. The evolutionary, epidemiological, and pathological characteristics of these lineages warrant further study.

Human Immunodeficiency Virus (HIV) types Western blot (WB) band profiles as potential surrogate markers of HIV disease progression and predictors of vertical transmission in a cohort of infected but antiretroviral therapy naïve pregnant women in Harare, Zimbabwe.

BACKGROUND: Expensive CD4 count and viral load tests have failed the intended objective of enabling access to HIV therapy in poor resource settings. It is imperative to develop simple, affordable and non-subjective disease monitoring tools to complement clinical staging efforts of inexperienced health personnel currently manning most healthcare centres because of brain drain. Besides accurately predicting HIV infection, sequential appearance of specific bands of WB test offers a window of opportunity to develop a less subjective tool for monitoring disease progression. METHODS: HIV type characterization was done in a cohort of infected pregnant women at 36 gestational weeks using WB test. Student-t test was used to determine maternal differences in mean full blood counts and viral load of mothers with and those without HIV gag antigen bands. Pearson Chi-square test was used to assess differences in lack of bands appearance with vertical transmission and lymphadenopathy. RESULTS: Among the 64 HIV infected pregnant women, 98.4% had pure HIV-1 infection and one woman (1.7%) had dual HIV-1/HIV-2 infections. Absence of HIV pol antigen bands was associated with acute infection, p = 0.002. All women with chronic HIV-1 infection had antibody reactivity to both the HIV-1 envelope and polymerase antigens. However, antibody reactivity to gag antigens varied among the women, being 100%, 90%, 70% and 63% for p24, p17, p39 and p55, respectively. Lack of antibody reactivity to gag p39 antigen was associated with disease progression as confirmed by the presence of lymphadenopathy, anemia, higher viral load, p = 0.010, 0.025 and 0.016, respectively. Although not statistically significant, women with p39 band missing were 1.4 times more likely to transmit HIV-1 to their infants. CONCLUSION: Absence of antibody reactivity to pol and gag p39 antigens was associated with acute infection and disease progression, respectively. Apart from its use in HIV disease diagnosis, WB test could also be used in conjunction with simpler tests like full blood counts and patient clinical assessment as a relatively cheaper disease monitoring tool required prior to accessing antiretroviral therapy for poor resource settings. However, there is also need to factor in the role of host-parasite genetics and interactions in disease progression.

The incidence of HIV among women recruited during late pregnancy and followed up for six years after childbirth in Zimbabwe.

BACKGROUND: HIV incidence is a useful tool for improving the targeting of populations for interventions and assessing the effectiveness of prevention strategies. A study in Harare, Zimbabwe reported cumulative incidences of 3.4% (3.0-3.8) and 6.5% (5.7-7.4) among post-partum women followed for 12 and 24 months respectively between 1997 and 2001. According to a Government report on HIV the prevalence of HIV fell from about 30% in 1999 to 14% in 2008. The purpose of this study was to determine the incidence of HIV-1 among women enrolled during late pregnancy and followed for six years after childbirth and to identify risk factors associated with acquisition of HIV. METHODS: HIV-uninfected pregnant women around 36 weeks gestation were enrolled from primary health care clinics in peri-urban settlements around Harare and followed-up for up to six years after childbirth. At every visit a questionnaire was interview-administered to obtain socio-demographic data and sexual history since the previous visit. A genital examination was performed followed by the collection of biological samples. RESULTS: Of the 552 HIV-uninfected women 444 (80.4%) were seen at least twice during the six years follow-up and 39 acquired HIV, resulting in an incidence (95% CI) of 2.3/100 woman-years-at-risk (wyar) (1.1-4.1). The incidence over the first nine months post-partum was 5.7/100 wyar (3.3-8.1). A greater proportion of teenagers (15.3%) contributed to a high incidence rate of 2.9/100 (0.6-8.7) wyar. In multivariate analysis lower education of participant, RR 2.1 (1.1-4.3) remained significantly associated with HIV acquisition. Other risk factors associated with acquisition of HIV-1 in univariate analysis were young age at sexual debut, RR 2.3, (1.0-5.6) and having children with different fathers, RR 2.7(1.3-5.8). Women that knew that their partners had other sexual partners were about four times more likely to acquire HIV, RR 3.8 (1.3-11.2). CONCLUSION: The incidence of HIV was high during the first nine months after childbirth. Time of seroconversion, age and educational level of seroconverter are important factors that must be considered when designing HIV intervention strategies.

Sexual behaviour does not reflect HIV-1 prevalence differences: a comparison study of Zimbabwe and Tanzania.

BACKGROUND: Substantial heterogeneity in HIV prevalence has been observed within sub-Saharan Africa. It is not clear which factors can explain these differences. Our aim was to identify risk factors that could explain the large differences in HIV-1 prevalence among pregnant women in Harare, Zimbabwe, and Moshi, Tanzania. METHODS: Cross-sectional data from a two-centre study that enrolled pregnant women in Harare (N = 691) and Moshi (N = 2654) was used. Consenting women were interviewed about their socio-demographic background and sexual behaviour, and tested for presence of sexually transmitted infections and reproductive tract infections. Prevalence distribution of risk factors for HIV acquisition and spread were compared between the two areas. RESULTS: The prevalence of HIV-1 among pregnant women was 26% in Zimbabwe and 7% in Tanzania. The HIV prevalence in both countries rises constantly with age up to the 25-30 year age group. After that, it continues to rise among Zimbabwean women, while it drops for Tanzanian women. Risky sexual behaviour was more prominent among Tanzanians than Zimbabweans. Mobility and such infections as HSV-2, trichomoniasis and bacterial vaginosis were more prevalent among Zimbabweans than Tanzanians. Reported male partner circumcision rates between the two countries were widely different, but the effect of male circumcision on HIV prevalence was not apparent within the populations. CONCLUSIONS: The higher HIV-1 prevalence among pregnant women in Zimbabwe compared with Tanzania cannot be explained by differences in risky sexual behaviour: all risk factors tested for in our study were higher for Tanzania than Zimbabwe. Non-sexual transmission of HIV might have played an important role in variation of HIV prevalence. Male circumcision rates and mobility could contribute to the rate and extent of spread of HIV in the two countries.

Antenatal HIV-1 RNA load and timing of mother to child transmission; a nested case-control study in a resource poor setting.

OBJECTIVE: To determine HIV-1 RNA load during the third trimester of pregnancy and evaluate its effect on in utero and intra-partum/postpartum transmissions in a breastfeeding population. DESIGN: A nested case-control study within a PMTCT cohort of antiretroviral therapy naive pregnant women and their infants. METHODS: A case was a mother who transmitted HIV-1 to her infant (transmitter) who was matched to one HIV-1 positive but non-transmitting mother (control). RESULTS: From a cohort of 691 pregnant women, 177 (25.6%) were HIV-1 positive at enrollment and from these 29 (23%) transmitted HIV-1 to their infants, 10 and 19 during in utero and intra-partum/postpartum respectively. Twenty-four mothers sero-converted after delivery and three transmitted HIV-1 to their infants. Each unit increase in log10 viral load was associated with a 178 cells/mm(3) and 0.2 g/dL decrease in TLC and hemoglobin levels, p = 0.048 and 0.021 respectively, and a 29% increase in the risk of transmission, p = 0.023. Intra-partum/postpartum transmitters had significantly higher mean viral load relative to their matched controls, p = 0.034. CONCLUSION: Antenatal serum HIV-1 RNA load, TLC and hemoglobin levels were significantly associated with vertical transmission but this association was independent of transmission time. This finding supports the rationale for preventive strategies designed to reduce vertical transmission by lowering maternal viral load.

The burden and risk factors of Sexually Transmitted Infections and Reproductive Tract Infections among pregnant women in Zimbabwe.

BACKGROUND: Sexually transmitted infections (STIs) and Reproductive tract infections (RTIs) are responsible for high morbidity among women. We aim to quantify the magnitude of the burden and risk factors of STI/RTI s among pregnant women in Zimbabwe. METHODS: A cross sectional study of pregnant women enrolled at 36 weeks of gestation from the national PMTCT program. Study was conducted from three peri-urban clinics around Harare Zimbabwe offering maternal and child health services. RESULTS: A total of 691 pregnant women were enrolled. Prevalence of HSV was (51.1%), HIV (25.6%) syphilis (1.2%), Trichomonas vaginalis (11.8%), bacterial vaginosis (32.6%) and Candidiasis (39.9%). Seven percent of the women had genital warts, 3% had genital ulcers and 28% had an abnormal vaginal discharge. Prevalence of serological STIs and vaginal infections were 51% and 64% respectively. Risk factors for a positive serologic STI were increasing age above 30 years, polygamy and multigravid; adjusted OR (95% CI) 2.61(1.49-4.59), 2.16(1.06-4.39), 3.89(1.27-11.98) respectively, partner taking alcohol and number of lifetime sexual partners. For vaginal infections it was age at sexual debut; OR (95% CI) 1.60(1.06-2.42). More than 25% of the women reported previous STI treatment. Fifty two percent reported ever use of condoms and 65% were on oral contraceptives. Mean age gap for sexual partners was 6.3 years older. CONCLUSIONS: There is a high morbidity of STI/RTIs in this cohort. There is need to continuously screen, counsel, treat and monitor trends of STI/RTIs to assess if behaviour changes lead to reduction in infections and their sustainability.

Type-specific cervico-vaginal human papillomavirus infection increases risk of HIV acquisition independent of other sexually transmitted infections.

BACKGROUND: Sexually transmitted infections (STIs) such as herpes simplex virus (HSV)-2 are associated with an increased risk of HIV infection. Human papillomavirus (HPV) is a common STI, but little is know about its role in HIV transmission. The objective of this study was to determine whether cervico-vaginal HPV infection increases the risk of HIV acquisition in women independent of other common STIs. METHODS AND FINDINGS: This prospective cohort study followed 2040 HIV-negative Zimbabwean women (average age 27 years, range 18-49 years) for a median of 21 months. Participants were tested quarterly for 29 HPV types (with L1 PCR primers) and HIV (antibody testing on blood samples with DNA or RNA PCR confirmation). HIV incidence was 2.7 per 100 woman-years. Baseline HPV prevalence was 24.5%, and the most prevalent HPV types were 58 (5.0%), 16 (4.7%), 70 (2.4%), and 18 (2.3%). In separate regression models adjusting for baseline variables (including age, high risk partner, positive test for STIs, positive HSV-2 serology and condom use), HIV acquisition was associated with having baseline prevalent infection with HPV 58 (aHR 2.13; 95% CI 1.09-4.15) or HPV 70 (aHR 2.68; 95% CI 1.08-6.66). In separate regression models adjusting for both baseline variables and time-dependent variables (including HSV-2 status, incident STIs, new sexual partner and condom use), HIV acquisition was associated with concurrent infection with any non-oncogenic HPV type (aHR 1.70; 95% CI 1.02-2.85), any oncogenic HPV type (aHR 1.96; 95% CI 1.16-3.30), HPV 31 (aHR 4.25; 95% CI 1.81-9.97) or HPV 70 (aHR 3.30; 95% CI 1.50-7.20). Detection of any oncogenic HPV type within the previous 6 months was an independent predictor of HIV acquisition, regardless of whether HPV status at the HIV acquisition visit was included (aHR 1.95; 95% CI 1.19-3.21) or excluded (aHR 1.96; 95% CI 1.02-2.85) from the analysis. CONCLUSIONS/SIGNIFICANCE: Cervico-vaginal HPV infection was associated with an increased risk of HIV acquisition in women, and specific HPV types were implicated in this association. The observational nature of our study precludes establishment of causation between HPV infection and HIV acquisition. However, given the high prevalence of HPV infection in women, further investigation of the role of HPV in HIV transmission is warranted.

The prevalence, incidence and risk factors of herpes simplex virus type 2 infection among pregnant Zimbabwean women followed up nine months after childbirth.

BACKGROUND: Herpes simplex virus type 2 (HSV-2) is the leading cause of genital ulcer disease worldwide. The virus can be transmitted to neonates and there are scarce data regarding incidence of HSV-2 among women in pregnancy and after childbirth. The aim of this study is to measure the incidence and risk factors for HSV-2 infection in women followed for 9 months after childbirth. METHODS: Pregnant women were consecutively enrolled late in pregnancy and followed at six weeks, four and nine months after childbirth. Stored samples were tested for HSV-2 at baseline and again at nine months after childbirth and HSV-2 seropositive samples at nine months after childbirth (seroconverters) were tested retrospectively to identify the seroconversion point. RESULTS: One hundred and seventy-three (50.9%) of the 340 consecutively enrolled pregnant women were HSV-2 seronegative at baseline. HSV-2 incidence rate during the 10 months follow up was 9.7 (95% CI 5.4-14.4)/100 and 18.8 (95% CI 13.9-26.1)/100 person years at risk (PYAR) at four months and nine months after childbirth respectively. Analysis restricted to women reporting sexual activity yielded higher incidence rates. The prevalence of HSV-2 amongst the HIV-1 seropositive was 89.3%. Risk factors associated with HSV-2 seropositivity were having other sexual partners in past 12 months (Prevalence Risk Ratio (PRR) 1.8 (95% CI 1.4-2.4) and presence of Trichomonas vaginalis (PRR 1.7 95% CI 1.4-2.1). Polygamy (Incidence Rate Ratio (IRR) 4.4, 95% CI 1.9-10.6) and young age at sexual debut (IRR 3.6, 95% CI 1.6-8.3) were associated with primary HSV-2 infection during the 10 months follow up. CONCLUSIONS: Incidence of HSV-2 after childbirth is high and the period between late pregnancy and six weeks after childbirth needs to be targeted for prevention of primary HSV-2 infection to avert possible neonatal infections.

Feasibility and potential acceptability of three cervical barriers among vulnerable young women in Zimbabwe.

BACKGROUND: We explored the potential acceptability of three cervical barriers (CB) (Ortho All-Flex diaphragm, SILCS diaphragm, FemCap cervical cap) among sexually experienced Zimbabwean young women. METHODS: Forty-five young women (aged 16-21 years) received an individual CB educational session. Participants were then randomly assigned to one of the three CBs in a 1:1:1 ratio, and practised insertion and removal of their device at the clinic. Next, participants were interviewed on their practice experiences, and their post-practice attitudes towards CB. RESULTS: All 45 young women were willing and able to insert their assigned device. The majority reported "easy" insertion and removal and 93% "liked" the device they tried. All showed interest in participating in future CB studies: when asked which device they would like to try in the future, over half (58%) chose SILCS, regardless of the device they had tried. The majority felt comfortable touching their genitals to insert/remove the CB and most participants favoured methods' attributes associated with female-control and non-interference with sex. Over half the participants said they would prefer to use a CB continuously compared to episodic use. Two-thirds of them expressed interest in CB for dual protection. CONCLUSION: The concept of CB, and initial insertion experience, were well accepted in this selected, small group of Zimbabwean young women. Evaluating CB in larger studies seems feasible in this population.