In 2015, Botswana introduced the quadrivalent human papillomavirus (HPV) vaccine as a two-dose schedule in girls aged 9-13 years. We sought to establish a baseline HPV prevalence in unvaccinated young adults in Botswana. HIV-uninfected men and women aged 18-22 years were recruited from the University of Botswana in Gaborone during October 2019-February 2021. Demographic and behavioural characteristics were self-reported during structured interviews. Self-collected vaginal and penile swabs were tested for 28 HPV types using Seegene Anyplex II HPV28. We compared any HPV type, quadrivalent vaccine (HPV 6, 11, 16, 18)-type and non-quadrivalent vaccine-type prevalence in men and women and evaluated the risk factors for prevalence of any HPV type. A total of 493 men and 500 women were included in the analysis. Compared to men, women had higher prevalence of any HPV type (63.0% versus 31.4%, P < 0.001), vaccine-type HPV (21% versus 9.7%, P < 0.001) and non-vaccine-type HPV (60.4% versus 28.4%, P < 0.001). Higher prevalence of any HPV type in men and women was associated with having >/=2 sex partners in the past 12 months; always using condoms in the past 3 months was associated with a lower HPV prevalence. These data provide baseline information for future evaluation of the population impact of the HPV vaccination programme, including potential herd effects in men.

INTRODUCTION: In 2015, Botswana introduced quadrivalent human papillomavirus (HPV) vaccine for girls aged 9-13 years. To establish a baseline HPV prevalence for future HPV vaccine impact monitoring, we evaluated HPV prevalences among the youngest unvaccinated women in Botswana and compared HPV prevalences among women living with HIV (WLHIV) and without HIV. METHODS: Women aged 18-22 years were recruited from the University of Botswana and HIV clinics in Gaborone from October 2019-January 2021. Demographic and behavioral characteristics were self-reported during structured interviews; HIV clinical characteristics were abstracted from medical charts. Self-collected vaginal swabs were tested for 28 HPV types using Seegene Anyplex II HPV28. We compared prevalence of any HPV, high risk (HR)-HPV, and quadrivalent HPV vaccine types (HPV6/11/16/18) among WLHIV and women without HIV and evaluated risk factors for prevalence of HR-HPV. RESULTS: A total of 306 WLHIV and 500 women without HIV were recruited. Compared to women without HIV, WLHIV were more likely to be sexually experienced (86.6% versus 74.4%) and have ≥ 3 lifetime sex partners (55.3% versus 27.8%). All HPV type prevalences were significantly higher among WLHIV compared to women without HIV, including prevalence of any HPV (82.7% versus 63.0%), HR-HPV (72.9% versus 53.8%), and quadrivalent vaccine HPV types (34.3% versus 21.0%). Among WLHIV, there were no differences between those perinatally and non-perinatally infected for HPV prevalences, number of HPV types detected, CD4 count, or viral load. CONCLUSIONS: Over one-third of WLHIV and nearly a quarter of those without HIV had vaccine-type HPV detected. This study supports need for the national HPV vaccination program in Botswana and provides important baseline data for future evaluation of impact of the program.

Aberrant promoter methylation of biologically relevant genes in cervical cancer and uneven CpG distribution within the human papillomavirus 16 (HPV 16) enhancer region have been reported. Cervical samples and questionnaires from 151 women screened for cervical cancer in Appalachian Ohio were analyzed. Methylation was measured by bisulfite sequencing in candidate gene sites in ESR1, DCC, p16, and LINE1 elements. Among 89 HPV 16-positive women, CpG sites in the E6 promoter and enhancer regions and the L1 region of the HPV 16 genome were measured. Methylation levels were compared by cervical cytology and HPV 16 status. HPV methylation was low regardless of cytology status, however E6 methylation was significantly higher in women with normal cytology. ESR1 and DCC methylation were significantly higher in HPV 16-positive women. Increased methylation at sites in the E6 promoter region was associated with lower odds of abnormal cytology. Increased methylation in candidate genes was associated with higher odds of abnormal cytology, particularly DCC region 2.4, DCC region 2.6, ESR1 region 3.2, and LINE1 site 1.2. HPV 16 genome CpG methylation was low except for the L1 region. In general, lower HPV 16 methylation and higher candidate gene methylation levels were associated with higher odds of abnormal cytology.

BACKGROUND: Human papillomavirus (HPV) 52 is one of the most frequent high risk HPV types found in cervical and anal infections. Reliable and well characterized methods for HPV 52 detection are therefore of great importance. Unfortunately one of the most widely used commercially available HPV typing assays, the Roche Linear Array (LA), detects type 52 only through its XR probe which cross-reacts with types 33, 35 and 58 and fails to give unambiguous detection of HPV 52. METHODS: To address this problem a real-time TaqMan PCR assay for HPV 52 targeting the E6/E7 region was developed and validated, which can be applied as robust duplex assay simultaneously detecting beta-globin as genomic control and reference or as highly sensitive single target detection assay. RESULTS: Optimized primer and probe concentrations produced linear PCR amplifications over seven logs of targets (10(1) - 10(7)). The detection limit for HPV 52 was reproducibly at 10 copies per reaction for the duplex assay format and 5 copies for the single-plex format. The assay was very type-specific and no amplification signal was observed with 10(7) copies of the related HPV33, 35, and 58 DNA. Of 89 samples that tested unambiguously positive for HPV 52 in the LA, 75 were confirmed in the duplex format and 88 in the single-plex format. An additional 100 samples negative for HPV 52 in LA were all negative in both HPV 52 real-time PCR assay formats. CONCLUSIONS: These results indicate 92.6% and 99.5% accuracy relative to LA for the duplex and single-plex formats, respectively. In ongoing testing of 18,484 from various studies, 10.8% required the HPV52 TaqMan assay to unequivocally determine the status. Including the HPV 52 duplex assay provides the ability to monitor variations in the cell yield in various methods of sample collection and processing. This additional information is useful in QC monitoring of epidemiologic studies.

The performance of three line blot assays (LBA) - Linear Array HPV genotyping assay (LA; Roche Diagnostics), INNO-LiPA HPV Genotyping Extra (LiPA; Innogenetics) and the Reverse Hybridization assay (RH; Qiagen) was evaluated using quantitated whole genomic HPV plasmids (types 6, 11, 16, 18, 31, 33, 35, 39, 51, 52, 56, 58, 59, 68b) as well as epidemiologic samples. In a plasmid titration series LiPA and RH did not detect 50 international units (IU) of HPV 18 in the presence of 5 x 10(4) or more IU HPV16. HPV DNA (1-6 types) in the plasmid challenges at 50 (IU) or genome equivalents (GE) were identified with an accuracy of 99.9% by LA, 97.3% by LiPA and 95.4% by RH with positive reproducibility of 99.8% (Kappa=0.992), 88.2% (Kappa=0.928) and 88.1% (Kappa=0.926) respectively. Two instances of mis-typing occured with LiPA. Of the 120 epidemiologic samples 76 were positive for high-risk types by LA, 90 by LiPA and 69 by RH with a positive reproducibility of 87.3% (Kappa=0.925), 83.9% (Kappa=0.899) and 90.2% (Kappa=0.942) respectively. Although all the assays had good concordance in the clinical samples, the greater accuracy and specificity in the plasmid panel suggest that the LA assay has an advantage for internationally comparable genotyping studies.

OBJECTIVES: To characterize the epidemiology of genital human papillomavirus (HPV) infection in children without previous consensual sexual activity, comparing HPV prevalence by certainty of child sexual abuse (CSA). PATIENTS AND METHODS: Patients presenting for evaluation of CSA in 8 sites in Atlanta, Houston, Harrisburg, and New York City were recruited along with patients presenting for unrelated health visits. CSA certainty was classified as definite, probable, possible, or no evidence following published guidelines and the results of history, physical examination, and laboratory tests. Urine and swabs of external genitalia were tested for HPV using L1 consensus polymerase chain reaction. RESULTS: The study included 576 participants (89.9% female) aged 6 months to 13 years (mean: 7.9); 534 of whom were evaluated for CSA and 42 for unrelated reasons. Of those evaluated for CSA, 14 had genital warts. One or more HPV types were detected in 11.8% (61 of 517) of participants with adequate samples. HPV detection was more likely among abused participants (definite, probable, or possible) than among participants without evidence of CSA (13.7% and 1.3%, respectively; P < .0001) and increased with certainty of abuse (8.4%, 15.6%, and 14.5% in participants with possible, probable, and definite CSA, respectively; P < .0001). Participants aged 10 years or older had a higher prevalence of HPV (20.6%) than others (5.6%) (P < .0001). CSA, anogenital warts, and age were independently associated with HPV detection. CONCLUSIONS: HPV detection was associated with CSA and increased with CSA certainty. In this population, genital HPV seemed to behave as a sexually transmitted infection.

BACKGROUND: Human papillomavirus (HPV) infection of the cervix and related abnormal cervical cytology in HIV-infected women has been well described. Little is known about anal HPV infection in HIV-infected women. METHODS: The SUN Study is a prospective cohort study of 700 HIV-infected patients including 167 women. At baseline, patients completed a behavioral questionnaire and provided, among other samples, cervical and anal swabs for HPV detection and genotyping and for cytologic examination. Here, we present the available baseline data on the 167 women in the SUN study. RESULTS: Baseline results were available for 120 women (median age: 38 years, 57% non-Hispanic black, median CD4 cell count 444.5 cells/mm), of whom, 77% were taking antiretroviral therapy. The prevalences in the anus and cervix of any HPV were 90% and 83%, respectively (P = 0.039), and of high-risk (HR) types 85% and 70%, respectively, (P = 0.001). There was no significant difference in the prevalences of abnormal cytology between the anus and cervix: 38% and 33%, respectively (P = 0.217). Although the presence of abnormal cervical cytology was associated with the presence of abnormal anal cytology (relative risk: 1.7, P = 0.024), its sensitivity (52.5%) and positive predictive value positive (45.6%) for identifying women with abnormal anal cytology were poor. A history of anal sex was not associated with anal HPV infection or abnormal anal cytology. CONCLUSIONS: In this cohort of HIV-infected women, anal HPV infection was more prevalent and diverse than cervical HPV infection. Anal cytologic abnormalities were as prevalent as cervical cytologic abnormalities, and although abnormal cervical cytology was predictive of abnormal anal cytology, results were not highly concordant. These data support the need for studies of anal cytologic screening of HIV-infected women.

HPV16 genotype variants have been the subject of several investigation, but study participants were rarely sampled more than ones. Among a cohort of Human immunodeficiency virus (HIV)-infected adults, we investigated HPV16 variants in samples collected concurrently from anus and cervix, as well as in serial samples collected from the same anatomical site at twelve-month intervals. We determined HPV16 variants in stored extracts of cervical and anal samples from subjects with multiple visits and at least one sample positive for HPV16. Seven polymorphic nucleotide positions within the E6 region were analyzed by pyrosequencing to determine genotype variants. Of 364 samples examined, 176 anal and 39 cervical swabs from 84 different subjects yielded unequivocal sequences of eight major HPV16 variants. Eight samples contained probable novel HPV16 variants and in 1 sample two variants were detected. In eight of 29 (27.6%) anal-cervical sample pairs positive for HPV 16, discordant variants were found. From 57 anal and 9 cervical sample series of HPV 16 positive samples a change in HPV16 variant status over time was seen in nine (15.8%) instances (7 anal, 2 cervical) from eight different participants. Changes of HPV 16 variants in HIV-infected adults was most frequently seen when different anatomic sites were sampled, but was also observed over time.