Abstract
Objectives
Cervical cancer is the leading cause of cancer-related mortality among women in sub-Saharan Africa (SSA). Data on HPV epidemiology in adolescent girls in SSA are essential to inform HPV vaccine policy recommendations for cervical cancer prevention. We assessed the burden of HPV infection, and risk factors for infection, among adolescent girls around the time of sexual debut.
Methods
Cross-sectional study of secondary school girls aged 17-18 years in Tanzania. Consenting participants provided samples for HPV and STI testing. Vaginal swabs were tested for 37 HPV genotypes by Roche Linear Array®. Logistic regression was used to identify factors associated with HPV infection. Y-chromosome was tested as a marker of recent condom-less sex.
Results
163/385 girls (42.3%) reported previous penetrative sex. HPV was detected in 125/385 (32.5%) girls, including 84/163 (51.5%) girls reporting previous sex and 41/222 (18.5%) reporting no previous sex. High-risk (HR) genotypes were detected in 70/125 (56.0%) girls with HPV infection. The most common HR genotype was HPV-16 (15/385; 3.9%). The prevalence of other HR HPV vaccine genotypes was between 0.8%-3.1%. Among 186 girls who reported no previous sex, were negative for Y-chromosome and had no sexually transmitted infections, 32 (17%) had detectable HPV. Lactobacillus species and bacterial vaginosis (BV)-associated bacteria were negatively and positively associated, respectively, with HPV.
Conclusions
HPV prevalence among adolescent girls around the time of sexual debut was high. However, prevalence of most vaccine genotypes was low, indicating that extending the age range of HPV vaccination in this region may be cost-effective.
Introduction
Cervical cancer is the leading cause of cancer mortality among women in sub-Saharan Africa (SSA), and East Africa bears one of the highest burdens, with an age-standardised incidence of 40/100,000 and mortality rate of 30/100,000.[1] Almost all cervical cancers can be attributed to persistent infection with one of 13 high-risk (HR) genotypes of human papillomavirus (HPV).[2] In addition to its oncogenic potential, HPV may also be an important co-factor in HIV acquisition.[3]
Infection with up to 7 HR and 2 low risk (LR) HPV genotypes can be prevented with HPV vaccination.[4] However the vaccine offers less protection once an HPV genotype has been acquired. Vaccination is recommended before first sex since the predominant mechanism of HPV acquisition is thought to be through penetrative sex.[5] HPV incidence increases rapidly after first sex and with changes of sexual partner, although most infections are cleared within 12 months.[6,7] Reviews of global age-specific HPV prevalence show the highest prevalence in women aged <25 years.[8] Most women are assumed to be HPV negative before first sex; however, some studies have detected HPV in girls and young women who report no previous penetrative sex.[9,10]
The few published studies on HPV infection in adolescent girls in SSA suggest that HPV prevalence may be very high at a young age. A study in Tanzania found HPV prevalences of 73% in sexually active girls aged 14-18 years, and one in Uganda in girls aged 12-24 found a prevalence of 75%.[11,12] A recent study in Tanzanian girls aged 15-16 years who reported no previous penetrative sex found an HPV prevalence of 8%.[13] The prevalence of non-optimal vaginal microbiota, including bacterial vaginosis (BV), is particularly high in SSA.[14] The vaginal microbiome may modulate susceptibility to HPV infection, as well as other sexually transmitted infections (STI) and HIV.[15]
There is an important need for data on HPV epidemiology in adolescent girls in SSA in order to inform HPV vaccine policy recommendations, to help allocate scarce public-health resources efficiently and achieve the greatest public health gains, especially as vaccine supplies are currently constrained.[16] Policymakers in SSA may not be able to draw conclusions from other settings because of differences in demographic structure, sexual behaviour, HPV genotype distribution, and co-factors such as HIV infection. As part of a cross-sectional study of the vaginal microbiota of girls aged 17-18 years in secondary schools in Tanzania, over half of whom reported no previous sex, we measured the burden of HPV infection and risk factors for infection.
Methods
Study design
The study design and procedures have been reported previously.[17] Briefly, this was a cross sectional survey in Mwanza, north-western Tanzania. Girls were enrolled between November 2013-June 2014 from government-funded secondary schools; selected schools had at least 25 girls in the target age range. Inclusion criteria were being aged 17-18 years, resident in Mwanza, and planning to stay in Mwanza for 1 month post-enrolment. Exclusion criteria were being outside the age range; being unwilling/unable to provide informed assent/consent (or parent unable/unwilling to provide informed consent, if aged 17); being temporarily in Mwanza, or planned travel within 1 month post-enrolment.
Girls were interviewed about socio-demographics, hygiene practices, and sexual behaviours. Participants provided 5 self-administered vaginal swabs in the presence of a nurse who assisted them if needed. Blood and urine samples were collected. Participants were offered HIV voluntary counselling and testing, with referral for care if positive. Laboratory results for treatable STI and free treatment as required were provided to participants within 2 weeks.
Laboratory methods
Laboratory procedures have been described previously.[17] HPV genotyping used the Linear Array HPV Genotyping assay (Roche Molecular Systems, USA), which detects 37 genotypes. DNA was extracted using the AmpliLute Liquid Media Extraction kit (Roche Molecular Systems), and amplified using the Linear Array HPV Genotyping Test. Generated amplicons were detected using the Linear Array Detection Kit. PCR reaction in this assay is based on a multiplex system, including human β-globin amplification primers, as an internal control for specimen quality. Specimens consistently negative for β-globin amplification were excluded since it was assumed that vaginal sampling was unsuccessful, or the extraction or amplification failed. DNA extraction, amplification and typing were performed in different rooms and included negative processing controls.
Vaginal swabs were tested for Neisseria gonorrhoeae (NG), Chlamydia trachomatis (CT) and Mycoplasma genitalium (MG) by in-house real-time PCR.[17] Concentrations of Lactobacillus crispatus, L. gasseri, L. jensenii, L. iners, L. vaginalis, Gardnerella vaginalis, Atopobium vaginae were measured using quantitative (q)PCR as previously described,[18] with DNA extraction by the QIAmp DNA mini kit. Primers and probes were from Eurogentec S.A. (Belgium) and PCRs were run on the QIAGEN Rotorgene. Gram-stained vaginal smears were examined for BV using the Nugent score. Trichomonas vaginalis (TV) was diagnosed by culture (InPouch TV, BioMed Diagnostics, USA). Serum samples were tested for HSV-2 antibodies by ELISA (Kalon Biological, UK). Syphilis was determined by Immutrep Rapid Plasma Reagin (Omega Diagnostics, Scotland) and Treponema pallidum particle agglutination assay (SERODIA, Fujirebio, Japan). Blood samples were tested with Determine HIV1/2 rapid test (Alere, Japan), then Uni-Gold HIV (Trinity Biotech, Ireland) if reactive. If both tests were reactive, the result was deemed positive. If tests were discordant, the sample was tested with HIV1/2 Stat-Pak (Chembio, USA), and deemed positive if reactive. Swabs from girls who reported no previous sex were tested for Y-chromosome using an in-house real time PCR, as a marker of recent condom-less sex.[19] Y-chromosome can be detected for up to 15 days, so may provide a rough measure of reporting bias.
Except for Y-chromosome testing, all tests were done at the National Institute for Medical Research (NIMR) laboratory, Mwanza. Quality assurance (QA) and Y-chromosome testing were performed by the STI Reference Laboratory at ITM Antwerp.
Statistical methods
Questionnaire data were double-entered into OpenClinica (Akaza Research, USA), and analysed using STATA V14.0 (StataCorp, USA).
Participant characteristics, and the number of infections of each HPV genotype, were tabulated among girls who reported no previous penetrative sex and those who had passed sexual debut (termed ‘sexually active’). Socioeconomic status was measured using a deprivation score, based on household ownership of 3 items: 1=car (least deprived); 2=television, without car; 3=mobile phone, without car or television; 4=none of the 3 items (most deprived). We used logistic regression to estimate odds ratios (OR) and 95% confidence intervals (CI) for factors associated with any HPV infection among all girls. Potential determinants of HPV infection were examined using a conceptual framework with 3 levels: sociodemographic, behavioural, and biological factors. Age was considered an a priori confounder and included in all models. First, sociodemographic factors whose age-adjusted association with HPV infection was significant at p<0.10 were included in a multivariable model; those remaining associated at p<0.10 were retained. Behavioural factors were then added to this model one by one. Those that were associated with HPV at p<0.10, after adjusting for sociodemographic factors, were retained if they remained significant at p<0.10. Associations with biological factors were determined in a similar way. This strategy allowed us to assess the effects of variables at each level of the framework, adjusted for more distal variables. We used a similar approach to examine sexual behaviour factors associated with HPV infection among girls who reported being sexually active.
Ethical considerations
The Institutional Review Board of the Institute of Tropical Medicine in Antwerp (867/13), the Ethics Committee of the University Teaching Hospital in Antwerp (13/14/147), the Lake Zone Institutional Review Board in Mwanza (MR/53/100/86) and the National Ethics Committee of the NIMR Coordinating Committee (NIMR/HQ/R.8a/Vol.IX/1544) approved the study protocol. All participants provided written informed consent/assent; written parental consent was required for participants <18 years.
Results
Characteristics of study participants
We identified 26 eligible secondary schools; 24 participated in the study. 1210 girls aged 17-18 years were registered on the school lists; 802 (66%) were located and their parents invited to a meeting about the study. 439 parents (55%) attended the meeting and 421 (96%) agreed to their daughter’s participation. 401/421 (95%) girls consented/assented and were enrolled (50% of those located; 33% of those on the school lists). Of these, 385 (97%) had HPV results and were included in the analysis.
Overall, 222 (58%) participants reported never having had penetrative sex. Of those who reported previous sex, 61% (99) had passed sexual debut in the past year. Sexually active girls were older than those who reported no previous sex (51% vs. 39% aged 18 years, respectively; p=0.02); however, there was no evidence of a difference in other sociodemographic characteristics (Table 1). Nearly all participants (381; 99%) had passed menarche, at a median age of 14 years (IQR 14-15). Among girls who reported no previous sex, 20 (9%) reported non-penetrative sexual contact with a male partner (e.g. kissing, genital touching).
Table 1. Characteristics at enrolment of 385 adolescent girls attending secondary school in Mwanza, Tanzania.
| Report no previous penetrative sex (N=222) n (column%) | Report previous penetrative sex (N=163) n (column%) | All girls (N=385) n (column%) | |
|---|---|---|---|
| Sociodemographic | |||
| Age | |||
| 17 years | 135 (61%) | 80 (49%) | 215 (56%) |
| 18 years | 87 (39%) | 83 (51%) | 170 (44%) |
| Tribe | |||
| Sukuma | 99 (45%) | 70 (43%) | 169 (44%) |
| Non-Sukuma | 123 (55%) | 93 (57%) | 216 (56%) |
| Religion | |||
| Catholic | 97 (44%) | 87 (53%) | 184 (48%) |
| Other Christian | 88 (40%) | 54 (33%) | 142 (37%) |
| Muslim | 32 (14%) | 20 (12%) | 52 (14%) |
| Other | 5 (2%) | 2 (1%) | 7 (2%) |
| Secondary school form | |||
| Form 1 | 2 (1%) | 1 (1%) | 3 (1%) |
| Form 2 | 47 (21%) | 21 (13%) | 68 (18%) |
| Form 3 | 127 (57%) | 99 (61%) | 226 (59%) |
| Form 4 | 46 (21%) | 42 (26%) | 88 (23%) |
| Who lives with | |||
| Mother in household | 146 (66%) | 98 (60%) | 244 (63%) |
| Father but not mother | 12 (5%) | 12 (7%) | 24 (6%) |
| Neither mother or father | 64 (29%) | 53 (33%) | 117 (30%) |
| Deprivation score | |||
| 1 (least deprived) | 16 (7%) | 8 (5%) | 24 (6%) |
| 2 | 88 (40%) | 76 (47%) | 164 (43%) |
| 3 | 110 (50%) | 73 (45%) | 183 (48%) |
| 4 (most deprived) | 8 (4%) | 6 (4%) | 14 (4%) |
| Behavioural | |||
| Ever drink alcohol? | |||
| Yes | 6 (3%) | 5 (3%) | 11 (3%) |
| Ever kissed with tongues | |||
| Yes | 19 (9%) | 76 (47%) | 95 (25%) |
| Ever engaged in genital touching1 | |||
| Yes | 3 (1%) | 40 (25%) | 43 (11%) |
| Ever had oral sex2 | |||
| Yes | 1 (<1%) | 9 (6%) | 10 (3%) |
| Ever had anal sex | |||
| Yes | 0 | 2 (1%) | 2 (1%) |
| Passed menarche | |||
| Yes | 218 (98%) | 163 (100%) | 381 (99%) |
| Ever cleanse inside vagina | |||
| Yes | 16 (7%) | 42 (26%) | 58 (15%) |
Sexual touching with a man/boy where girl touched his penis with her hand, he touched her vagina with his hand, or he rubbed his penis on her legs/buttocks/genitals but did not have vaginal sex.
Ever had man/boy put his penis in girl’s mouth, or he licked/sucked the girl’s genitals.
The overall prevalence of any STI was 21% among girls who reported previous sex (TV 9%, CT 5%, NG 2%, MG 4%, HSV2 3%, HIV 0%), and 7% among girls who reported no previous sex (TV 1%, CT <1%, NG 1%, MG 1%, HSV2 2%, HIV 1%). BV prevalence among girls who reported previous sex was 33%, and 19% among those reporting no previous sex.
Prevalence of HPV genotypes
387/401 girls provided vaginal swabs for HPV testing (6 were pregnant, 8 refused); β-globin was detected in 385/387 specimens. The prevalence of any HPV infection was 32.5% (125/385); 64/125 girls (51.2%) with HPV infection had >1 genotype. The most prevalent HR genotypes were HPV-16 (3.9%), HPV-39 and HPV-52 (both 3.1%), and HPV-58 (2.9%) (Figure 1). HPV-18 was detected in 3 girls (0.8%). 17 girls (4.4%) were infected with HPV-16 and/or HPV-18, the HR genotypes targeted by all HPV vaccines. 53 girls (13.8%) were infected with a genotype targeted by the new 9-valent HPV vaccine, Gardasil-9.
Figure 1.
HPV genotype-specific prevalence among 385 girls attending secondary school in Mwanza, Tanzania. Vertical lines indicate 95% confidence intervals and numbers are raw frequencies.
HPV prevalence varied by self-reported sexual behaviour (online supplementary material Table S1; Figures S1 and S2). HPV was detected in 84/163 (51.5%) sexually active girls and 41/222 (18.5%) girls who reported no penetrative sex. HR HPV was detected in 47 (28.8%) sexually active girls, and 23 (10.4%) who reported no penetrative sex. Among those with HR HPV, 31.9% of sexually active girls and 13.0% of those who reported no penetrative sex were infected with >1 HR genotype. The most common HR genotypes among sexually active girls were HPV-52 (6.7%), HPV-16 (5.5%), HPV-39 and HPV-68 (both 4.9%). Among girls who reported no penetrative sex, the most common HR genotypes were HPV-16 and HPV-58 (both 2.7%).
Among the 222 girls who reported no penetrative sex, 19 (9%) were either positive for Y-chromosome (N=7) or had laboratory-confirmed STI other than HPV (N=12). HPV prevalence among the 186 girls who reported no sexual contact nor penetrative sex, and had no evidence of Y chromosome nor any STI, was 17.2% (N=32).
Factors associated with HPV infection
In the unadjusted analysis among all girls, there was some evidence of an association of HPV with increasing deprivation score, and strong evidence of an association with sexual behaviour (penetrative sex, kissing, engaging in genital touching; Table 2). There was also evidence of an association with vaginal cleansing and menstrual hygiene. After adjusting for age, deprivation score and penetrative sex, there was still strong evidence of an association with menstrual hygiene (p=0.004), with participants who used cloths instead of commercial pads having the lowest odds of HPV infection, and those who used only underwear having the highest odds of infection. There was also weak evidence of an association with vaginal cleansing (aOR=1.70, 95%CI=0.92-3.16, p=0.09). After adjusting for age, deprivation score, penetrative sex, menstrual hygiene and vaginal cleansing, there was evidence of an association with gonorrhoea (aOR=5.70, 95%CI=0.91-35.6), MG (aOR=6.01, 95%CI=1.08-33.6), HIV (aOR=10.4, 95%CI=0.90-121.5) and BV (aOR=1.93, 95%CI=1.13-3.29).
Table 2. Factors associated with any HPV infection among 385 adolescent girls attending secondary school in Mwanza, Tanzania.
| N with HPV / total N (%) |
Crude OR (95% CI) |
||
|---|---|---|---|
|
Sociodemographic | |||
| Age | P=0.40 | ||
| 17 years | 66 / 215 (30.7%) | 1 | |
| 18 years | 59 / 170 (34.7%) | 1.20 (0.78–1.84) | |
| Tribe | P=0.85 | ||
| Sukuma | 54 / 169 (32.0%) | 1 | |
| Non-Sukuma | 71 / 216 (32.9%) | 1.04 (0.68–1.60) | |
| Religion | P=0.60 | ||
| Catholic | 59 / 184 (32.1%) | 1 | |
| Other Christian | 51 / 142 (35.9%) | 1.19 (0.75–1.88) | |
| Muslim | 15 / 52 (28.8%) | 0.86 (0.44–1.69) | |
| Other | 0 / 7 (0.0 %) | – | |
| Who lives with | P=0.86 | ||
| Mother in household | 79 / 244 (32.4%) | 1 | |
| Father but not mother | 9 / 24 (37.5%) | 1.25 (0.53–2.99) | |
| Neither mother or father | 37 / 117 (31.6%) | 0.97 (0.60–1.55) | |
| Deprivation score | |||
| 1 (least deprived) | 6 / 24 (25.0%) | ||
| 2 | 51 / 164 (31.1%) | P=0.09 | |
| 3 | 59 / 183 (32.2%) | 1.32 (0.95–1.82) | |
| 4 (most deprived) | 9 / 14 (64.3%) | ||
| Behaviour |
Adjusted OR (95% CI)1 |
||
| Ever drink alcohol? | P=0.28 | P=0.24 | |
| No | 123 / 374 (32.9%) | 1 | 1 |
| Yes | 2 / 11 (18.2%) | 0.45 (0.10–2.13) | 0.41 (0.08 -2.05) |
| Ever kissed with tongues | P=0.006 | P=0.82 | |
| No | 83 / 290 (28.6%) | 1 | 1 |
| Yes | 42 / 95 (44.2%) | 1.98 (1.23–3.19) | 0.94 (0.54 -1.63) |
| Ever engaged in genital touching2 | P=0.003 | P=0.64 | |
| No | 102 / 342 (29.8%) | 1 | 1 |
| Yes | 23 / 43 (53.5%) | 2.71 (1.42–5.14) | 1.19 (0.59 -2.40) |
| Ever had oral sex3 | P=0.07 | P=0.55 | |
| No | 119 / 375 (31.7%) | 1 | 1 |
| Yes | 6 / 10 (60.0%) | 3.23 (0.89–11.65) | 1.51 (0.39 -5.87) |
| Ever had vaginal sex | P<0.001 | P<0.001 | |
| No | 41 / 222 (18.5%) | 1 | 1 |
| Yes | 84 / 163 (51.5%) | 4.69 (2.97–7.42) | 4.81 (3.02 -7.66) |
| Menstrual hygiene4 | P=0.002 | P=0.004 | |
| Pads only (+/- pants) | 68 / 202 (33.7%) | 1 | 1 |
| Cloths only (+/- pants) | 12 / 71 (16.9%) | 0.40 (0.20–0.80) | 0.45 (0.22 -0.94) |
| Pants only | 15 / 29 (51.7%) | 2.11 (0.96–4.63) | 2.75 (1.17 -6.46) |
| Cloth & pads (+/- pants) | 30 / 79 (38.0%) | 1.21 (0.70–2.07) | 1.16 (0.65 -2.08) |
| Ever cleanse inside vagina | P=0.001 | P=0.09 | |
| No | 95 / 327 (29.1%) | 1 | 1 |
| Yes | 30 / 58 (51.7%) | 2.62 (1.48–4.62) | 1.70 (0.92 -3.16) |
|
Biological | |||
| HSV-2 | P=0.45 | P=0.48 | |
| Negative | 121 / 376 (32.2%) | 1 | 1 |
| Positive | 4 / 9 (44.4%) | 1.69 (0.44–6.39) | 1.70 (0.40 -7.29) |
| Chlamydia trachomatis | P=0.15 | P=0.76 | |
| Negative | 120 / 376 (31.9%) | 1 | 1 |
| Positive | 5 / 9 (55.6%) | 2.67 (0.70–10.11) | 1.25 (0.31 -5.02) |
| Neisseria gonorrhoeae | P=0.03 | P=0.05 | |
| Negative | 120 / 378 (31.7%) | 1 | 1 |
| Positive | 5 / 7 (71.4%) | 5.37 (1.03–28.10) | 5.70 (0.91 -35.56) |
| Trichomonas vaginalis | P=0.02 | P=0.79 | |
| Negative | 115 / 368 (31.3%) | 1 | 1 |
| Positive | 10 / 17 (58.8%) | 3.14 (1.17–8.46) | 1.16 (0.39 -3.45) |
| Active syphilis (RPR+/TPPA+) | |||
| Negative | 125 / 385 (32.9%) | – | – |
| Positive | 0 (–) | – | – |
| HIV | P=0.24 | P=0.06 | |
| Negative | 123 / 382 (32.2%) | 1 | 1 |
| Positive | 2 / 3 (66.7%) | 4.21 (0.38 -46.89) | 10.44 (0.90 -121.5) |
| Mycoplasma genitalium | P=0.005 | P=0.02 | |
| Negative | 118 / 376 (31.4%) | 1 | 1 |
| Positive | 7 / 9 (77.8%) | 7.65 (1.57–37.39) | 6.01 (1.08 -33.56) |
| Bacterial vaginosis5 | P=0.002 | P=0.05 | |
| Normal | 71 / 261 (27.2%) | 1 | 1 |
| Intermediate | 9 / 28 (32.1%) | 1.27 (0.55–2.93) | 0.94 (0.36 -2.44) |
| BV | 45 / 95 (47.4%) | 2.41 (1.48–3.92) | 1.93 (1.13 -3.29) |
Behavioural factors adjusted for age (a priori), deprivation score, and vaginal sex. Biological factors adjusted for age, deprivation score, vaginal sex, menstrual hygiene and vaginal cleansing.
Sexual touching with a man/boy where girl touched his penis with her hand, he touched her vagina with his hand, or he rubbed his penis on her legs/buttocks/genitals but did not have vaginal sex.
Ever had man/boy put his penis in girl’s mouth, or he licked/sucked the girl’s genitals.
Sanitary products used for menstrual hygiene; most girls who reported using pads or cloths also reported wearing pants (underwear). Restricted to girls who have passed menarche (N=381).
Missing data for 1 girl.
Among sexually active girls, after adjusting for age and deprivation score, there was evidence of an association of HPV with having >1 lifetime partner (aOR=2.63, 95%CI=1.22-5.69), an older first partner, and a first partner who had concurrent partners (online supplementary material Table S2). There was also evidence of an association with time since sexual debut, with HPV infection highest among those whose sexual debut was 1-2 years ago (aOR=2.29, 95%CI=1.01-5.21, relative to those with sexual debut <1 year ago).
Association with vaginal microbiota
Among all girls, after adjusting for age, deprivation score and penetrative sex, HPV infection was positively associated with A. vaginae (aOR=2.19, 95%CI=1.32-3.64; p=0.002) and G. vaginalis (aOR=1.79, 95%CI=1.05-3.04) (Table 3). In contrast, HPV infection had an inverse association with several Lactobacillus species, including L. crispatus (aOR=0.48, 95%CI=0.29-0.80; p=0.005) and L. jensenii (aOR=0.44, 95%CI=0.27-0.73; p=0.001), and, to a lesser extent, L vaginalis. The same trends were seen for the association with high levels (>1 million cells/mL) of each bacterial species.
Table 3. Association of bacterial species with any HPV infection among 385 adolescent girls attending secondary school in Mwanza, Tanzania.
| N with HPV / total N (%) | Crude OR (95% CI) | Adjusted OR (95% CI)1 | Adjusted OR (95% CI)2 | |
|---|---|---|---|---|
| Presence/absence | ||||
| A. vaginae 3 | P<0.001 | P<0.001 | P=0.002 | |
| Absent | 49 / 220 (22.3%) | 1 | 1 | 1 |
| Present | 63 / 139 (45.3%) | 2.89 (1.82-4.59) | 2.30 (1.40-3.76) | 2.19 (1.32-3.64) |
| G. vaginalis 4 | P<0.001 | P=0.02 | P=0.03 | |
| Absent | 31 / 145 (21.4%) | 1 | 1 | 1 |
| Present | 87 / 209 (41.6%) | 2.62 (1.62-4.25) | 1.87 (1.12-3.14) | 1.79 (1.05-3.04) |
| L. vaginalis 5 | P<0.001 | P=0.03 | P=0.04 | |
| Absent | 59 / 131 (45.0%) | 1 | 1 | 1 |
| Present | 64 / 242 (26.4%) | 0.44 (0.28-0.69) | 0.58 (0.36-0.93) | 0.60 (0.37-0.98) |
| L. crispatus 6 | P<0.001 | P=0.005 | P=0.005 | |
| Absent | 55 / 120 (45.8%) | 1 | 1 | 1 |
| Present | 60 / 238 (25.2%) | 0.40 (0.25-0.63) | 0.49 (0.30-0.81) | 0.48 (0.29-0.80) |
| L. gasseri 7 | P=0.91 | P=0.86 | P=0.76 | |
| Absent | 98 / 300 (32.7%) | 1 | 1 | 1 |
| Present | 24 / 75 (32.0%) | 0.97 (0.56-1.67) | 1.06 (0.59-1.89) | 1.10 (0.61-1.99) |
| L. iners 8 | P=0.08 | P=0.20 | P=0.37 | |
| Absent | 16 / 68 (23.5%) | 1 | 1 | 1 |
| Present | 106 / 311 (34.1%) | 1.68 (0.92-3.08) | 1.51 (0.79-2.89) | 1.35 (0.69-2.63) |
| L. jensenii 5 | P<0.001 | P<0.001 | P=0.001 | |
| Absent | 83 / 198 (41.9%) | 1 | 1 | 1 |
| Present | 37 / 175 (21.1%) | 0.37 (0.23-0.59) | 0.44 (0.27-0.71) | 0.44 (0.27-0.73) |
| >1,000,000/mL | ||||
| A. vaginae 3 | P<0.001 | P<0.001 | P=0.001 | |
| No | 55 / 239 (23.0%) | 1 | 1 | 1 |
| Yes | 57 / 120 (47.5%) | 3.03 (1.90-4.83) | 2.37 (1.44-3.92) | 2.33 (1.39-3.91) |
| G. vaginalis 4 | P<0.001 | P=0.02 | P=0.02 | |
| No | 47 / 195 (24.1%) | 1 | 1 | 1 |
| Yes | 71 / 159 (44.7%) | 2.54 (1.61-4.00) | 1.83 (1.12-2.98) | 1.83 (1.10-3.03) |
| L. vaginalis 5 | P=0.57 | P=0.75 | P=0.68 | |
| No | 91 / 269 (33.8%) | 1 | 1 | 1 |
| Yes | 32 / 104 (30.8%) | 0.87 (0.53-1.41) | 0.92 (0.54-1.55) | 0.89 (0.52-1.53) |
| L. crispatus 6 | P<0.001 | P=0.009 | P=0.01 | |
| No | 59 / 132 (44.7%) | 1 | 1 | 1 |
| Yes | 56 / 226 (24.8%) | 0.41 (0.26-0.64) | 0.52 (0.32-0.85) | 0.52 (0.32-0.87) |
| L. gasseri 7 | P=0.57 | P=0.41 | P=0.34 | |
| No | 106 / 331 (32.0%) | 1 | 1 | 1 |
| Yes | 16 / 44 (36.4%) | 1.21 (0.63-2.34) | 1.35 (0.67-2.73) | 1.42 (0.69-2.91) |
| L. iners 8 | P=0.05 | P=0.24 | P=0.52 | |
| No | 20 / 85 (23.5%) | 1 | 1 | 1 |
| Yes | 102 / 294 (34.7%) | 1.73 (0.99-3.01) | 1.43 (0.79-2.59) | 1.22 (0.66-2.25) |
| L. jensenii 5 | P<0.001 | P<0.001 | P<0.001 | |
| No | 94 / 238 (39.5%) | 1 | 1 | 1 |
| Yes | 26 / 135 (19.3%) | 0.37 (0.22-0.60) | 0.40 (0.24-0.68) | 0.38 (0.22-0.65) |
Adjusted for age (a priori), deprivation score, and vaginal sex.
Adjusted for age (a priori), deprivation score, vaginal sex, menstrual hygiene and vaginal cleansing.
Missing data for 26 girls.
Missing data for 31 girls.
Missing data for 12 girls.
Missing data for 27 girls.
Missing data for 10 girls.
Missing data for 6 girls.
Discussion
We found a high prevalence of HPV infection among adolescent Tanzanian girls around the time of reported sexual debut. Over half of sexually active participants and one-fifth of those who reported no penetrative sex had HPV infection, and 56% of those infected had an HR genotype. Multiple HPV genotype infections were also very common, with 51% of HPV-infected girls having >1 genotype. These findings are consistent with previous studies of HPV in sexually active adolescents in the USA and in Africa.[11,12,20]
HPV-16 was the most prevalent HR genotype (3.9%). The prevalence of HPV-18 was <1% and of other vaccine genotypes was <3%, except for HPV-52 (targeted by Gardasil-9), which was 3.1%. Similarly, a study in Mozambique found HPV-52, 58 and 16 were the most common HR genotypes in young women.[21] Also, a large meta-analysis of HPV prevalence worldwide found HPV-16 to be the most common genotype among women in SSA, followed by HPV-52.[8]
The Director General of the World Health Organization recently announced a goal to eliminate cervical cancer.[22] Currently vaccination strategies target girls in an age range considered to be pre-sexual debut, typically 9-14 years. The optimum upper age limit for vaccination will depend on different factors including HPV vaccine genotype prevalence, age of sexual debut, cost per dose of vaccine, availability of cervical cancer screening, and HPV transmission dynamics. Screening for cervical cancer is extremely limited in Tanzania. Previous studies of HPV epidemiology in Tanzania have shown one of the highest reported HPV incidences, and a high prevalence of vaccine-related genotypes among young women aged 20-25 years, so the cost-effectiveness of vaccinating girls up to 17 years is likely to be high.[11] Over half the girls in our study reported no previous penetrative sex, and most were not yet infected with vaccine-related genotypes. These findings suggest that a catch-up strategy that goes beyond the multiyear cohort approach of vaccinating 9-14 year olds,[23] by offering vaccination to girls aged 15-17 years, could help reduce HPV acquisition at a critical time after sexual debut. Furthermore, this would contribute towards the goal of cervical cancer elimination by decreasing the proportion of females in the population who would otherwise acquire persistent HPV infection, a necessary prerequisite for cervical cancer.
The Tanzanian national programme is currently vaccinating with Gardasil, which covers 2/13 (15%) circulating HR genotypes in our study population, but only one of the more common ones (HPV-16). In contrast, Gardasil-9 would cover 7/13 (54%) circulating HR genotypes, including the three most common ones. Although Gardasil and Cervarix offer some cross-protection for other genotypes (Gardasil against HPV-31, and Cervarix against HPV-31, 33, and 45), these cross-protected genotypes were less common in our population. Therefore, Gardasil-9 may offer the best coverage given the distribution of HR genotypes in our setting.
HPV transmission appears extremely efficient in the early years of sexual activity in our setting, with >60% of girls whose sexual debut was 1-2 years ago being infected with HPV. HPV prevalence among girls who report no penetrative sex was also high (18.5%), and substantially higher than in studies in industrialised countries among women who reported no previous sex. A cross-sectional study in Sweden in women aged 10-25 years who reported no previous sex found a prevalence of 1.5%.[10] A longitudinal study in USA found HPV in only 1.7% of samples from women aged 18-20 who never had sex.[9] Our prevalence is also higher than in a longitudinal study in Tanzanian girls aged 15-16 years who reported no penetrative sex over 18 months, which found HPV in only 11.6% of samples.[24] Lack of disclosure is likely to be part of the explanation for the high HPV prevalence in girls in our study who denied previous sexual activity. This is supported by our finding that 9% of girls who reported no previous sex were positive for Y-chromosome and/or an STI (excluding HPV). In Tanzania, girls who are still in school may be particularly reluctant to disclose sexual activity, since potential consequences include expulsion, physical punishment or social exclusion.[25]
Alternative explanations for HPV infection in girls who report no previous sex include mother-to-child transmission, non-penetrative sexual contact, or transmission via fomites.[26,27] A recent study in Mwanza showed a high prevalence of HPV DNA in oral washes and fingertip samples from adolescent girls, and on surfaces in their bathrooms.[28] Nevertheless, even with potential reporting errors, our findings of a very high HPV prevalence in girls who report no penetrative sex is important.
After adjusting for potential confounders, we found a strong inverse association between HPV and D-lactic-acid and H2O2-producing Lactobacillus species, including L. crispatus and L. jensenii, key constituents of optimal vaginal microbiota. Furthermore, there was a strong positive association between HPV infection and anaerobic bacterial species G. vaginalis and A. vaginae. These species are characteristic of BV, which has been associated with increased susceptibility to STI and HIV.[29] A recent meta-analysis found that disturbance in the vaginal microbiota away from a Lactobacillus-dominated environment was associated with increased risk of HPV acquisition and persistence, and related cervical disease.[30]
Strengths of our study include detailed interviews given by trained nurses experienced in adolescent sexual behaviour research. Collection of vaginal swabs, although self-administered, was observed by nurses; all but 2 specimens contained β-globin, indicating successful sampling and specimen processing. The Roche Linear Array used for HPV genotyping has a high sensitivity and specificity; all laboratory assays were conducted according to SOPs with external QA at an internationally-recognised reference laboratory.
Limitations include the cross-sectional design, which makes it difficult to assess causality or to measure past HPV infection, since individual genotype-specific infections may be rapidly cleared.[7] Face-to-face interviews may have increased social desirability bias in responses; the inclusion of parents and recruitment from schools may have compounded this issue. We enrolled girls who were still in school; many girls in this age range in Tanzania are no longer in school. Unpublished demographic and health survey (DHS) data from the Mwanza region in 2017 showed 41% of girls aged 17-18 years were still in school. Furthermore, only 33% of 1210 girls on the school lists were enrolled, mostly because they were not found at the school or their parents could not be located, which suggests possible selection bias. National DHS data show that young women with secondary education have an older age at first sex, and later age at first birth.[31] Therefore, we may have underestimated HPV prevalence among all women in this age group. However, our findings of the distribution of HPV genotypes, and factors associated with HPV, are consistent with other studies in SSA, and may be more broadly generalisable.
In conclusion, we found a high prevalence of HPV infection, and HR genotypes, among adolescent girls in the early years after becoming sexually active, and among girls who reported no penetrative sex. HPV vaccination in Tanzania is currently offered to 14 year old girls through a national vaccination programme. The prevalence of most vaccine-related genotypes was low, indicating that extending the age range of HPV vaccination through a catch-up campaign in this region, with one of the highest rates of cervical cancer worldwide and limited facilities for screening, may be cost-effective.
Supplementary Material
HPV genotype-specific prevalence1 among 165 girls attending secondary school in Mwanza, Tanzania, who report having passed sexual debut. Vertical lines indicate 95% confidence intervals and numbers are raw frequencies.
HPV genotype-specific prevalence among 222 girls attending secondary school in Mwanza, Tanzania, who report not having passed sexual debut Vertical lines indicate 95% confidence intervals and numbers are raw frequencies.
Key Messages.
HPV infection among adolescent girls attending secondary school in Tanzania was high (32.5%), and high-risk (HR) oncogenic genotypes were detected in over half the girls with HPV infection
HPV infection was inversely associated with Lactobacillus species, key constituents of optimal vaginal microbiota.
The Tanzanian national programme is vaccinating with Gardasil, which protects against 2/13 (15%) HR genotypes circulating in our study population, including the most prevalent one (HPV-16).
Extending the age range of vaccination in this region, where cervical cancer screening is extremely limited, may be cost-effective.
Acknowledgments
We would like to express our gratitude to the parents and participants for who gave up their time and provided samples for this study, and to our dedicated study nurses and clinical officers: Fidelis Malifedha, Beatrice Kamala, Justa Lucas, Esther Mwinuka, Herijema Yona, and AnnaMaria King, and our field tracing team: Deborah Byabato, Scholastica Magucha, Cecilia Paul, Malilo Juma, Dickson Rubirya, Enos Mbata, Mathias Godda and Damian Dotto. We also thank the laboratory team, and Simon Buhalata for technical assistance in Nugent scoring, and Rama Hashim for data management. Finally, we thank administrative and support staff at the Mwanza Interventional Trials Unit and the Institute of Tropical Medicine for their contribution to this study.
Financial Support
This work was supported by the European and Developing Countries Clinical Trials Partnership (project code: SP.2011.41304.066). Additionally, KJB, SCF and RJH received salary support through a jointly funded award by the UK Medical Research Council (MRC) and the UK Department for International Development (DFID) under the MRC/DFID Concordat agreement which is also part of the EDCTP2 programme supported by the European Union (MR/R010161/1). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Footnotes
Competing Interests
DWJ has received research grants from GSK Biologicals for HPV vaccine-related research. All other authors declare that they have no conflicts of interest.
Author Contributions
Conception or design of the work: DWJ, AB, JC, TC, SN, KJB, SCF, AA, RJH. Data collection: JI, SN, AA. Data analysis and interpretation: KJB, CHH, RJH, AB, DWJ. Drafting the article: KB, DWJ. Critical revision of the article: KJB, AA, SCF, TC, AB, DWJ. Final approval of the version to be published: DWJ, AB, RJH, CHH, SCF, TC, JC, SN, JI, AA, KJB.
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Associated Data
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Supplementary Materials
HPV genotype-specific prevalence1 among 165 girls attending secondary school in Mwanza, Tanzania, who report having passed sexual debut. Vertical lines indicate 95% confidence intervals and numbers are raw frequencies.
HPV genotype-specific prevalence among 222 girls attending secondary school in Mwanza, Tanzania, who report not having passed sexual debut Vertical lines indicate 95% confidence intervals and numbers are raw frequencies.