There is conclusive evidence that human papillomavirus (HPV) infections of the cervix are a necessary cause of cervical cancer. In Italy there are consistent data of HPV prevalence in women aged 25 - 64 years, but there is limited data for younger women. The objective of this on-going 3-year prospective cohort study is to investigate the prevalence, acquisition, clearance and persistence of HPV infections in young Tuscan women and the risk factors correlated with such events.One thousand and sixty-six women aged between 18 and 24 years were enrolled and received an initial HPV test. They were asked to return to the clinic over the study period for further tests every 12 months, if their HPV HR result was negative, or every 6 months, if positive. Additionally, women with an HPV positive result were given a cytological examination and if the cytological diagnosis was ASC-US or more severe, only women with HPV HR, were referred for colposcopy.We present here data for the enrolment phase of the study. At baseline, within the study sample, just under 30% of women were infected by HPV and 19.3% of women were infected with oncogenic types. A relationship was highlighted between HPV infection, number of sexual partners (in particularly in the last 3 years) and the lifetime number of partner's partners. Condom use showed a slight protective effect in univariate analysis but these data were not statistically significant in multivariate analysis. The association between HPV infection and demographic and behavioural variables were tested by crude odds ratio (OR). Multivariate logistic regression was applied to compute the adjusted odds ratios.The prevalence of oncogenic HPV types was high in young Tuscan women. The 3-year follow-up of this cohort may provide a better understanding of the processes of acquisition, clearance and persistence of infection and the correlated risk factors. Cervical cancer is the second most common cancer in women worldwide and knowledge regarding its cause and pathogenesis is rapidly expanding. Although there is conclusive evidence that infection of the cervix by some types of human papillomavirus (HPV) is a necessary cause of cervical cancer ,2 the diThe accumulated evidence that high risk types (HPV HR) are a necessary cause of cervical cancer has led to the design of prophylactic vaccines. The currently licensed vaccines are Gardasil  and Cervarix (GlaxoSmithKline) and protect against infection by HPV types 6, 11, 16 & 18 and types 16 & 18, respectively. The vaccines, both of which have been approved for girls and young women, are designed to prevent 90% of all genital warts and 70% of all cervical cancers (Gardasil) and 70% of all cervical cancers (Cervarix). Studies have shown that types 6 & 11 are responsible for 90% of genital warts and types 16 & 18 for 70% of cervical cancers. In Italy, there are consistent HPV prevalence data based on NTCC studies for women aged 25 - 64 years [The study sample was recruited from three areas of Tuscany  each with good experience in delivering population-based cervical cancer screening programmes which, following Italian national guidelines , includeWith the intention to enrol at least 1000 women and on the assumption of a 10% and 15% compliance to invitation, approximately 8000 women aged 18 - 24 years were randomly selected from Tuscan population registries, and were sent a letter with a request that they participate by calling and fixing an appointment. In cases of no response, a reminder was sent.The study protocol is briefly summarized in the following lines and shown graphically in Figure The study was approved by the local ethics committee in each area.At enrolment, trained midwives explained the study aims and procedures. A written informed consent was given by each participating woman. The cervical cell samples, obtained by using a plastic Ayre's spatula and a cytobrush, were put in PreservCyt solution  and were used for HPV testing and for cytological examination.Participants provided information on socio-demographic characteristics and sexual history. In a face-to-face, standardized interview participants gave information on age, race/ethnicity, education, occupation, reproductive and menstrual factors, number of pregnancies and smoking habits, and in a self-administered questionnaire at the clinic participants provided information on sexual history - age at first intercourse, age of the first partner at time of first intercourse, number of sexual partners in participant's lifetime and in the last year, lifetime number of partner's partners and use of contraceptive methods and condoms. During subsequent visits to the clinic, information on changes in smoking and sexual habits since baseline examination was requested, and endo- and ectocervical cells from the uterine cervix were collected.The Hybrid Capture 2 assay  was used for HPV testing. The two groups of probes designed to detect high-risk HPV types  and low-risk types  were used. Four mL of PreservCyt sample was processed with the Sample Conversion Kit  followed by an HC2 assay, according to the manufacturer's instructions. HC2 results were expressed in Relative Light Units (RLU) as the ratio of the specimen's light emission to that of three concurrently tested 1 pg/mL HPV DNA controls. An RLU is an (indirect) measure of the specimen's viral concentration relative to 1 pg/mL.All HPV HR positive samples were typed using the 'Consensus High Risk HPV genotyping' kit . The test is based on the reverse hybridization principle. Denaturated biotynilated amplicons, resulting from the amplification of a part of the L1 region with GP5+/GP6+ primers, were hybridized with specific oligonucleotide probes, which were immobilized as parallel lines on membrane strips. After hybridization and stringent washing, streptavidin-conjugated alkaline phosphatase was added and bound to any previously formed biotinylated hybrid. Incubation with chromogen yielded a purple precipitate and the results could be visually interpreted. The kit allows an easy and reliable identification of 18 HPV HR types .The HPV HR positive samples that were negative in the PCR enzyme-linked immunoadsorbent assay were typed with cycle sequencing and analyzed on an ABI PRISM 310 DNA Genetic Analyzer . All nucleotide sequences were analyzed and compared with HPV type sequences in GenBank using BLAST 2.0 . A sequence was considered a match if it had a 90% nucleotide identity to an HPV sequence in GenBank.In order to identify the HPV LR types, samples positive for HC2 probe A (specific for LR types) were amplified using specific primers for five HPV LR types .The amplified products were analyzed by gel electrophoresis in a 2% agarose gel containing ethidium bromide, and observed under ultraviolet light.HPV infections were defined by oncogenic risk ( i.e. high or low risk) and phylogenetic species .Liquid-based cytology was performed using the ThinPrep system  for all women who were HPV positive ,11. One The association between HPV infection and demographic and behavioural variables was tested by crude odds ratio (OR). Multivariate logistic regression was applied to compute the adjusted odds ratios.All variables were analyzed in the univariate analysis, but in the multivariate analysis we adjusted the model for the statistically significant variables and those variables that, though not statistically significant, play an important biological role.All tests were two-sided. Stata 10 software was used for analysis.Recruitment commenced in June 2007 and was completed by June 2008 with the enrolment of 1066 women. Seven women had been excluded as they did not meet the eligibility criteria. In Florence, Valdarno Aretino and Viareggio, respectively, 542, 289, and 235 women were enrolled. The overall compliance to invitation was 15.7% with no statistically relevant difference between areas . The compliance by age is shown in Table Of the study sample, 206 19.32%) women were HPV HR positive and 107 (10.03%) were HPV LR positive. Table 9.32% womFigure The association between HPV infection and the presence of cervical cytological abnormalities was examined in all HPV positive participants. Table Compliance to colposcopy was 82.76% (48/58): 18 had negative results, 24 had histological diagnoses lower than CIN2, 5 had a histological diagnosis CIN 2 and 1 had a histological diagnosis CIN 3.Table Some 45.1% of participants reported no current use of oral contraception (8 missing). The current use of oral contraceptives in the univariate analysis showed no effect on prevalence of infection.Four hundred and twenty two women were smokers . Smoking increased the risk of infection .Women reporting an occasional use of condoms (28.6%) had a similar risk to those who never used them (24.6%) , whereas an increasing protection was observed for women who used condoms most of the time (23.0%)  or always (22.3%) .No previous pregnancies showed a protective effect  as did not reporting previous genital disease . No partner's genital disease showed a not statistically significant protective effect . A lower level of education was associated with a not statistically significant increased risk of infection.Table After adjustment of the other covariates condom use, no prior pregnancies, no previous genital disease and greater age at first intercourse showed a very small and not statistically significant protection.This paper reports the baseline results at enrolment for an on-going study.The compliance to the invitation (15%) was low in comparison with the compliance to invitations for the cervical screening programs for women aged 25 - 64 years in the same area (approximately 40%). It was particularly low for women aged 18 and 19 years.The difference may be due, in part, to the lower age of the study population and their being unaccustomed to gynaecological examination, and in part, to the test offered (the HPV test) which was still little known in Italy when the study began. However, the participation rate is very similar to that obtained in similar studies conducted in Italy [Nevertheless, the participating women do not seem to differ from the general population in terms of age at first intercourse and the number of partners. In fact these characteristics were similar to those found in other Italian surveys. Ammatuna et al. in a study of 1000 Sicilian women reported a mean age of 21.6, a mean age of first sexual intercourse of 17.4, and mean number of lifetime partners of 2.22 .Our baseline result that almost 30% of sexually active women aged 18 - 24 years were infected by HPV is comparable with the range 19.7% - 39%) reported worldwide for women in the same age group ,14. Infe% - 39% rCervical infection with multiple HPV types was relatively common among Italian cohort participants, ,17. GoodHPV 16 (8.53%) and HPV 18 (1.88%) were present in 10.41% of the study sample. This prevalence of vaccine types would suggest that the young population being studied, especially the 18 & 19 year-olds, is an appropriate target for the prophylactic vaccination. Amongst 18 year-olds the prevalence of HPV 16 was 1.94% (4/206) and of HPV 18 was 0% (0/206), whereas amongst 19 year-olds the prevalence of HPV 16 was 2.43% (5/206) and of HPV 18 was 0.97% (2/206).Our baseline results confirm the roles of the number of sexual partners (in particularly in the last 3 years) and of the lifetime number of partner's partners  as deterAt baseline smoking habits act as an independent factor of increased acquisition of HPV infections in both the univariate or multivariate models. This association has been observed in the pooled analysis of the International Agency for Research on smoking and human papillomavirus infection where the risk of being HPV positive increased with smoking intensity, after allowing for lifetime number of sexual partners .The use of condoms has an slight protective effect in the univariate analysis, but is not statistically significant in the multivariate analysis, suggesting that use of condoms is associated to low risk sexual behaviours. In fact, our data shows that women who said they frequently use condoms, also indicated that they have had a low number of sexual partners.This point is heavily debated and has important consequences. Many previous papers, in which a retrospective approach has been adopted (as in this study) ,27-31, hAt least two different explanations might account for these conflicting figures:a) Information bias: information collected retrospectively could determine a misclassification bias. If this were the case, the relationship between condom use and HPV would move towards the null value.b) The probability of developing HPV infection depends on from the time passed since the first intercourse and as protection is not absolute for condom users, in the long term, they also could be become infected. In particular it is worth mentioning that, as time passes, HPV infection tends to spread to areas not protected by condoms and HPV infections can still be transmitted through non-penetrative sexual contact or contact with areas of unprotected genital skin.This study design will permit a prospective analysis, in a relatively short interval, of the role of different risk factors  in the acquisition of new HPV infections (or with a different HPV type) for women negative at baseline. Furthermore, it might permit an evaluation of the role of these risk factors in the clearance of infections for women positive at baseline.In the follow-up of this study we will analyze data from the yearly self-administered questionnaires regarding only the year to date. In this manner we aim to reduce reporting biases related to the large interval since the beginning of sexual activity and also with a temporal sequence of its progress.Financial competing interests· In the past five years have you received reimbursements, fees, funding, or salary from an organization that may in any way gain or lose financially from the publication of this manuscript, either now or in the future? Is such an organization financing this manuscript (including the article-processing charge)?NO· Do you hold any stocks or shares in an organization that may in any way gain or lose financially from the publication of this manuscript, either now or in the future?NO· Do you hold or are you currently applying for any patents relating to the content of the manuscript? Have you received reimbursements, fees, funding, or salary from an organization that holds or has applied for patents relating to the content of the manuscript?NO· Do you have any other financial competing interests?NOThe authors declare that they have not competing interests.Non-financial competing interestsAre there any non-financial competing interests  to declare in relation to this manuscript?NOThe authors declare that they have not competing interests.MC conceived the study and participated in its design and coordination and helped to draft the manuscript. FC conceived the study and participated in its design and coordination and helped to draft the manuscript. MZ performed the statistical analysis and helped to draft the manuscript. LV performed the statistical analysis and helped to draft the manuscript. AI carried out the protocol organization in Florence. PC carried out the cytological analysis in Florence. LB performed the sampling collection in Florence. MF performed the colposcopic examination. FM carried out the protocol organization and the cytological analysis in Arezzo. PV carried out the protocol organization in Lucca. AS carried out the cytological analysis in Lucca. DB performed the sampling collection in Florence. CS carried out the molecular analysis, participated in the study coordination and helped to draft the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/10/157/prepub


The female lower genital tract is a gateway for pathogens entering the host through the mucous membrane. One of the prevalent human viruses is Torque teno virus (TTV). The major reported routes of TTV transmission are fecal-oral and parenteral. Furthermore, other modes of transmission, e.g. sexual contact, are suggested. To investigate the sexual route of TTV transmission, cervical smears of healthy women and those with cervical lesions were screened for the presence of TTV DNA.TTV DNA was studied in cervical smears of 95 patients with cervical lesions and 55 healthy women. Paired serum samples were available from 55 and 42 women, respectively. All healthy women had normal cytology while 44 patients had histologically confirmed low-grade lesion (LGL) and 51 high-grade lesion (HGL). TTV DNA was detected with primers specific for the non-coding region. In 40 paired cervical smears and serum samples, the phylogenetic group of TTV isolates was determined. The presence of HPV DNA in cervical smears was detected by means of PCR with MY09/11 primers.The prevalence of TTV DNA in cervical smears of healthy women was 52.7% and was comparable with that in paired serum samples (50%). Symptomatic women had significantly higher prevalence of TTV DNA in cervical smears (74.7%) than healthy controls. The TTV DNA prevalence in patient serum samples was 51%. The phylogenetic groups of TTV serum isolates were concordant with those of TTV from cervical smears of the same subjects. In cervical smears, a wider variety of TTV isolates was found. The viral loads in cervical smears were 10 to 1000 times as high as in sera. The HPV-positive study subjects had significantly higher TTV DNA prevalence than HPV negatives. The prevalence of TTV was not associated with disease severity.High prevalence of TTV in cervical smears suggests that sexual transmission is another mode of expansion of TTV infection among the population. The higher viral load in cervical smears than in the respective serum samples might indicate active TTV replication in the female genital tract. Nevertheless, cooperation between TTV and HPV needs to be further investigated. The mucosal surface of the female lower genital tract is the first line of defense against pathogens. The physical barrier created by epithelial cells prevents the penetration of microbes and together with dendritic cells, macrophages, NK cells and chemical elements, they are the main components of the innate immune system. However, many common infectious disease pathogens succeed in entering the host through the mucous membrane after which they spread systematically. One of the most prevalent viral pathogens of cervical epithelial cells is human papillomavirus (HPV). HPV is a very common sexually transmitted infection with prevalence rates ranging from 2% to 44% among healthy women around the world . HPV plaAnellovirus [The human population is frequently infected with Torque teno virus (TTV), a small, non-enveloped DNA virus with a single stranded circular genome, recently classified into a floating family called llovirus . TTV shollovirus . To datellovirus . NeverthThe major routes of transmission are parenteral and fecal-oral. Since TTV DNA was detected in semen and cervical smears, sexual transmission has also been considered as a possible mode of infection -9. A stuTo investigate the possibility of sexual transmission of TTV, cervical smears from healthy women and those with cervical lesions were screened for the presence of TTV DNA. For most study subjects, parallel serum samples were also available for comparison and TTV subtyping. Co-infection with TTV and HPV was investigated to support the hypothesis of the sexual route of TTV transmission.Cervical smears and sera samples were obtained from the bank of the National Reference Laboratory for Papillomaviruses in Prague. All samples were collected during a prospective study of HPV in patients treated for cervical lesions in 1996–1997 . CervicaCervical brushes were transported in tubes containing phosphate-buffered saline solution (PBS) with 5 mM ethylene diamine tetraacetic acid (EDTA), pH 8.0, and stored at +4°C. The tubes were vortexed and cells were collected by centrifugation at 1,000 rpm for 10 min. Forty microlitres of each cell pellet were digested in 100 μl of lysis buffer  containing 100 μg/ml proteinase K  at 37°C overnight. Proteinase K was inactivated at 95°C for 10 minutes and the samples were stored at -20°C. DNA was extracted from 200 μl of serum by QIAamp Blood kit  and dissolved in 100 μl of the elution buffer . The control plasmid with a part of the human beta-globin gene was added to serum sample for the evaluation of DNA quality. Extracted DNA was stored at -20°C.The integrity and quality of DNA was assessed by amplification of the human beta-globin gene with primer PC03/PC04. All tested samples were positive for the human beta-globin gene .The cervical smears were analyzed for the presence of HPV DNA by means of PCR with MY09/11 primers followed by Southern blot hybridization and identification of HPV types on dot blot hybridization with oligonucleotide type specific probes or by sequencing .TTV DNA detection in cervical smears and serum samples was done by nested PCR directed at the non-coding region of TTV (PCR NCR) with NG133/147 and NG132/134  and withTen microlitres of the PCR product were separated electrophoretically on a 3% agarose gel .N) testing positive was used as a relative titer for determining the viral titer per 1 ml of TTV DNA in serum samples and cervical smears.DNA extracted from the sera and cervical smears from 5 patients with cervical abnormalities and 5 healthy women was serially diluted (in 10-fold steps) in distilled water and the presence of TTV DNA was detected by PCR NCR. The highest dilution  with 95% confidence intervals (CI) and two-tailed P values were calculated in 2 × 2 tables using the EPI INFO statistical package (2002 version) and Graph Pad InStat (version 3.05) . In all tests, the basic significance level was P = 0.05.In total, 150 cervical smears and 97 serum samples were screened by PCR for the presence of TTV. The prevalence rates of TTV in the study groups of healthy women and patients with cervical lesions are shown in Table The prevalence rates of TTV DNA were 52.7% in cervical smears of healthy women (29/55) and 50% in the parallel serum samples (21/42). The difference between the TTV DNA prevalence rates from the patient cervical smears and serum samples was statistically significant (74.7% (71/95) versus 51% (28/55), OR = 2.9, CI 1.4–5.8, P = 0.004). Comparable prevalence rates of TTV DNA were found in the sera from patients and healthy women, while patients showed significantly higher TTV DNA rates in cervical smears than healthy women .Overall, 98.9% (94/95) of cervical smears from patients with cervical abnormalities and 27.3% (15/55) cervical smears from healthy women were HPV DNA positive. The HPV-positive study subjects showed a significantly higher prevalence of TTV DNA, i.e. 71.6% (78/109), than the HPV-negatives with 48.8% (20/41) . The patients were divided into two groups according to cervical lesion severity. The TTV prevalence rate was slightly higher in the group with high-grade lesions, i.e. 76.5% (39/51), than in that with low-grade lesions, i.e. 68.2% (30/44), but the difference was not statistically significant.The relative viral loads in cervical smears and in the respective serum samples were compared for five patients and five healthy women. All of these subjects had 10 to 1.000 times higher TTV DNA titers in cervical smears than in serum samples.TTV genotype was determined in 25 patients and 15 healthy women with both tested specimens positive for TTV. Any of TTV genotypes was recognized in 86% of patient cervical smears, 50% of patient serum samples, 77% of healthy women's cervical smears and 34% of the healthy women's serum samples. The most prevalent phylogenetic group was TTV 1 (45%), followed by TTV 3 (40%), the phylogenetic group TTV 5 was found rarely (6%) and TTV 2 only in the group of patients (4%). Concordant phylogenetic groups of TTV isolates were found in serum samples and the parallel cervical smears from the same subject; however, in cervical smears more diverse TTV genotypes were present  in subjects with less than three sexual partners and 64.5% (20/31) in those with three or four sexual partners.TTV infects the majority of the general population worldwide but is not associated with any clinical evidence of disease. The high prevalence of TTV in the general population might be explained by the existence of multiple transmission routes. Several studies have reported the parenteral and fecal-oral routes of TTV transmission and other modes of transmission have also been considered. The presence of TTV DNA in cervical smears and semen suggests that TTV may be spread by sexual contact .In our study, in agreement with Calcaterra et al., Chan et al., and Fornai et al -8, we obThe cervical smears from patients with cervical lesions were more frequently infected by TTV in comparison with those from healthy women. The co-presence of HPV and TTV in the patient cervical smears suggests that both viruses have a common transmission route. The co-infection with TTV and other viruses such as HBV, HCV, and HIV l has been reported by others ,19,21. TOriginally, liver tissue had been thought to be a site of TTV replication since the viral loads in hepatocytes were 10 to 100 times higher than those in the respective plasma samples. This hypothesis was confirmed by the detection of a double-stranded variant of TTV DNA in the liver and hepatocytes by in situ hybridization. The viral titer was 10 to 10.000 times higher in saliva than in serum. The assumption that TTV might replicate in oropharyngeal tissues and/or salivary glands was supported by the detection of TTV in the gingival tissue . In our The reported higher viral loads of TTV in HIV infected patients might be associated with a poorer prognosis in AIDS. A higher viral load can be explained by the immune stimulation due to HIV infection. CIN biopsies in HIV-positive women have shown higher lymphocyte and macrophage counts in the squamous epithelium of the cervix and an increased intralesional concentration of proinflammatory cytokines modulating HIV replication, with consequently increased HIV genital shedding . SimilarTo conclude, high TTV prevalence in cervical smears suggests that sexual transmission is another mode of expansion of TTV infection among the population. The higher viral loads in cervical smears than in the respective serum samples might indicate active TTV replication in the female genital tract or might be the result of plasma secretion. The TTV shedding is possibly mediated by the immune upregulation in the cervix and vagina due to HPV infection. Nevertheless, natural cooperation between TTV and HPV needs to be further investigated.The authors declare that they have no competing interests.MS projected the study, carried out the experimental work, did all statistical analyses and evaluation of the results. MS wrote the first draft of the paper and other coauthors contributed to the final draft. RT and VN participated in the study design and data interpretation. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/9/118/prepub


Human Papillomavirus (HPV) is a well-known pathogen for lower genital tract neoplasias, yet little is known regarding HPV prevalence in Turkey. The aim of this study was to investigate the prevalence of HPV DNA and to determine HPV types distribution among women with normal and abnormal cytology.A total of five hundred seven (n = 507) women were retrospectively evaluated between 2004-2008. Conventional polymerase chain reaction was used to detect the presence of HPV types in cervicovaginal samples obtained from patients during gynecologic examination.One hundred four (n = 104) of the women were excluded from the study because of the incomplete data and a total of 403 women were used for the final analysis. There were, 93 (23%) women with cytologic abnormality and 310 (77%) women with normal cytology. Overall, 23% of the women was HPV positive. The overall prevalence of HPV in women with abnormal Pap smears was 36% (93/403), of which in ASCUS 22%, LSIL 51% and HSIL 60%. Also, HPV DNA was positive in 20% of the women with normal cervical cytology. The most common HPV types in cytologically normal women were as follows; HPV 16 (36%), HPV 6 (22%) and HPV 18 (13%). The rate of other HPV types were as follows; HPV11 4.4%, HPV45 4.4%, HPV90 4.4%, HPV35 2.2%, HPV67 2.2%, HPV81 2.2%, and multiple type HPVs 8.9%. The most common HPV types in cytologically abnormal women were HPV 16 (35%), HPV6 (19%) and HPV18 (8%). The rate of multiple HPV infections in women with normal Pap test was 2.2%.HPV prevalence and type distribution in this study were similar to that reported worldwide at least in our study population. Hovewer, HPV prevalence was more common compared with previous studies reported from Turkey. This might be related with methodology and hospital based patient accrual and high rate of women with abnormal cytology. Further population based prospective studies are needed to eliminate the drawbacks of our study and to determine nonhospital based HPV prevalence in Turkish women. Cervical cancer is the one of the leading cause of cancer-related deaths among women worldwide. It is e estimated 500,000 cases occurring annually and a 50% case fatality rate .According to statistics of Ministry of Health and GLOBOCAN 2002, cervical cancer is the 9th most common cancer among women in Turkey and ranks 13th among cancer-related deaths, with an estimated 1364 cases occurring annually and more than 50% of fatality rate [Today, it is widely accepted that Human papillomavirus virus (HPV) is the main causal factor of cervical carcinoma. More than 100 HPV genotypes have been described and nearly 20 of them have been associated with cervical carcinoma. It has been reported that HPV prevalence range between 1.4% to 25.6% in the world . PreviouBetween January 2004 and August 2008, 503 consecutive sample of women attending gynecological outpatient clinics for regular gynecologic control in Baskent university School of Medicine, Departments of Obstetrics and Gynecology were included to this retrospective analysis. Women were eligible if they had no previous diagnosis or treatment for cervical, vulvar or vaginal cancer and ever sexually active. Also, patients with previous history of chemoradiotherapy for cervical carcinoma were excluded from the analysis. Information on sociodemographic characteristics, medical history and sexual and reproductive behaviour was obtained at the time of the gynecological visit which was filled during the gynecological visit. Samples of exfoliated cervical cells were obtained using a cytobrush. After the preparation of a standard cervical smear, the remaining cells were placed in tubes with 0.9% saline and stored at - 201C until shipment to the our microbiology laboratory. Pap smears were classified according to the Bethesda system by pathology department. HPV DNA detection and genotyping performed by PCR in our microbiology labaratory as described previously ,11 StatiHPV DNA detection in cervical swabs was conducted by using real-time polymerase chain reaction (PCR) with a commercial kit . For DNA extraction the QIAamp DNA Mini Kit  was used in accordance with the instructions of the manufacturer. A 150 bp fragment of the L1 gene was amplified using GP5 and GP6 primers. An amplified gene product was identified via melting curve analysis and visualized by incorporation of Sybr Green dye during amplification. HPV genotyping was performed with DYEnamic ET Terminator Cycle Sequencing Kit  and ABI PRISM 310 Genetic Analyzer at Iontek Ltd, Turkey.The study protocol was approved by at Ethical Board of Baskent University, School of Medicine.During the four year period, 503 women who attend for regular gynecological examination were analyzed for HPV in our departments. One hundred four of these women were excluded from the analysis because of the incomplete data and a total of 403 women were used for the final analysis.Mean age of the patients was 37.5 ± 9.3 (range 19-67 y). Ninety percent of the population was premenaposal while 10% was menopausal. There were significant differences with respect to HPV positivity between women under 30 years old and women older than 30 years old  but, there was no significant differences with respect to decads of life and HPV positivity. Also, menopausal status has no effect on HPV positivity. HPV was positive in 24% of the premenopausal women and 17% of the menopausal women (p = 0.323).There were, 93 23%) women with cytologic abnormality and 310 (77%) women with normal cytology. Cervical cytologic abnormalities were diagnosed as follows; 42% atypical squamous cell of undetermined significance (ASCUS), 46% low-grade squamous intraepithelial lesion (LSIL), and 11% high-grade SIL (HSIL). Overall, 23% of the women was HPV positive. On the other hand, HPV was positive in the 36% of the women with cytologic abnormalities and this figure was 20% in women without cytologic abnomality. HPV was positive in 22%, 51%, 60% of the ASCUS, LSIL and HSIL respectively , at least 12% of oropharyngeal cancers and 3% of oral cancers . FurtherAlthough Turkey has historically had low incidence rates of invasive cervical cancer, cervical cancer ranks among the 9th most common female malignancies with an age-standardized incidence rate of 4.5 per 100.000. Our country has a more than 70 million population, but, unfortunately, there is no population based studies which investigating HPV prevalence in Turkish women. Previous studies reported that HPV prevalence between 2%-6% among the low risk women in Turkey (Table In 2007, Sanjosé et al. reported that overall HPV prevalence in 157.879 women with normal cervical cytology was estimated to be 10.4% in a meta-analysis of 78 studies . In 2008In women with abnormal cytology results, the presence of HPV infection has been reported in 28.8%-61.3% of cases -22. In oSome epidemiologic studies reported that a significant reduction in HPV prevalence throughout the third decade of life . It has In conclusion, although geographicly Turkey is located between Europe and Asia, HPV prevalence and genotypes are similar to those reported from western countries in Turkish women with normal and abnormal cytology. Hovewer, HPV prevalence was more common compared with previous studies reported from Turkey. This might be related with methodology and hospital based patient accrual and high rate of women with abnormal cytology. Further population based prospective studies are needed to eliminate the drawbacks of our study and to determine nonhospital based HPV prevalence in Turkish women.The authors declare that they have no competing interests.PD, conceived of the study, performed analysis and interpretation of data, drafted the manuscript and revised the manuscript. SSS performed the data analysis, revised the manuscript. HA, performed the data analysis, revised the manuscript. EK supervised the study, revised the manuscript. AA, supervised the study and revised the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/9/191/prepub


P<0.001), and was less common in those with a higher personal income . The risk of conjunctival cancer increased with increasing time spent in cultivation and therefore in direct sunlight , but decreased with decreasing age at leaving home , perhaps reflecting less exposure to sunlight consequent to working in towns, although both results were of borderline statistical significance. To reduce confounding, sexual and reproductive variables were examined among HIV seropositive individuals only. Cases were more likely than controls to report that they had given or received gifts for sex , but this may have been a chance finding as no other sexual or reproductive variable was associated with conjunctival cancer, including the number of self-reported lifetime sexual partners (P=0.4). The seroprevalence of antibodies against HPV-18 and -45 was too low to make reliable conclusions. The presence of anti-HPV-16 antibodies was not significantly associated with squamous cell carcinoma of the conjunctiva  and nor were anti-KSHV antibodies . The 10-fold increased risk of conjunctival cancer in HIV infected individuals is similar to results from other studies. The role of other oncogenic viral infections is unclear.As part of a larger investigation of cancer in Uganda, we conducted a case–control study of conjunctival squamous cell carcinoma in adults presenting at hospitals in Kampala. Participants were interviewed about social and lifestyle factors and had blood tested for antibodies to HIV, KSHV and HPV-16, -18 and -45. The odds of each factor among 60 people with conjunctival cancer was compared to that among 1214 controls with other cancer sites or types, using odds ratios, estimated with unconditional logistic regression. Conjunctival cancer was associated with HIV infection  87, 301–308. doi:10.1038/sj.bjc.6600451www.bjcancer.comCancer Research UK© 2002  Symptoms can range from none, to severe pain and visual loss. Lesions generally arise on exposed areas of the eye, particularly on the nasal side, and treatment involves local excision, or in more severe cases, orbital clearance. Metastases are rare and the prognosis is usually favourable.Squamous cell carcinoma of the conjunctiva is thought to be an extreme form of a spectrum of clinical conditions, collectively known as ‘ocular surface squamous neoplasias’, which range in severity from mild dysplasia, to carcinoma Although relatively rare everywhere, conjunctival carcinoma is more frequent in parts of sub-Saharan Africa. Uganda offers a good setting in which to investigate the epidemiology of squamous cell carcinoma of the conjunctiva, because the tumour was relatively frequent there, even before the onset of the HIV epidemic  with a provisional new diagnosis of incident cancer were eligible for recruitment into the study from all the wards and out-patient clinics of the four main hospitals in Kampala, Uganda: Mulago (including the Uganda Cancer Institute), Nsambya, Mengo and Rubaga. Further details of the methods can be found elsewhere . The conInterviewers  approached ward or clinic staff to gain permission to interview potential recruits. If permission was granted, the patient was approached by one of four interviewers and invited to participate in the study. A HIV test was requested and appropriate counselling was given. Patients who had been HIV tested within a month of recruitment and had a medical certificate indicating the test result, were not re-tested. The patient was interviewed about social and demographic factors, and sexual and reproductive history. Where possible, the patient was interviewed by a counsellor of the same sex, and in their native language. Blood was drawn, primarily for the HIV test, but also for storage of serum and leukocytes. Interviewers reported the HIV test results back to the patient, together with post-test counselling. The study was approved by the Committee on Human Research  and by the Uganda National Council for Science and Technology.Diagnoses of cancer were established by histology or other laboratory investigation, where possible. Diagnoses made on clinical grounds alone, were reviewed by the investigators . Among the cases, 36 (60%) underwent histological review  and the remainder were diagnosed clinically. Among controls, 63% of tumour diagnoses were verified by laboratory investigation .HIV-1 serostatus was determined using a single ELISA, the Cambridge Bioscience Recombigen enzyme linked immunosorbent assay . Periodic laboratory quality control assays using blinded standards from the United States Centers for Disease Control  revealed test sensitivity and specificity of 99%. In the initial months of the study, 40–50% of adults declined to have venepuncture, mostly because they were too ill. Therefore, a GACELISA saliva test for HIV was introduced as an option (with sensitivity and specificity similar to the blood test), and the refusal rate dropped to 10%. In total, HIV test results were available on 57 cases and 826 controls. However, for one case and seven controls the result was indeterminate. Any remaining aliquots of sera were stored at −80°C and were later shipped on dry ice to the Laboratoire de Virologie Moléculaire in Tours, France, for HPV testing and to University College London, UK, for KSHV testing.et al; 1998; BglII sites. The amplified product was then inserted into pFastBacI to generate HPV L1 VLPs. VLPs were produced according to a procedure used for HPV-16 VLPs (All assays were performed by a single investigator (LB), who was unaware of each patient's personal characteristics, and diagnosis. HPV VLPs were produced in Sf21 insect cells using recombinant baculoviruses encoding the L1 gene of HPV-16, -18, and -45, according to previously described procedures  were read at 492 nm with an automated plate reader . For each serum sample the background reactivity found in the BSA coated wells was subtracted from the OD found in each of the HPV–VLP coated wells. Negative values were adjusted to zero. The cut-off values for positivity to HPV-16, -18 and -45 were set up at 0.2 (OD test well minus OD control well) and those with a value of 0.4 or greater were considered to have a high titre of anti-HPV antibodies. In total, 457 samples were available for testing for antibodies against HPV-16 (39 cases and 418 controls). In addition, there were sufficient sera for 453 of those to be tested for antibodies against HPV-18 and -45 (39 cases and 414 controls).Flat-bottomed wells of 96-well microplates  were coated overnight at 4°C with 200–400 ng of VLPs (test well) or 200 ng of BSA (control well) in PBS, pH 7.4. Each serum sample was tested twice against each of the seven VLP types and BSA at the same time on the same plate. After washing with PBS, 0.1% Tween 20 and 200 μl of PBS containing 1% newborn bovine serum  were added (2 h at 37°C). The blocking solution was replaced by 100 μl of sera diluted 1 : 20 in 5xPBS-10% NBS and 2% Tween 20, and plates were incubated at 45°C for 60 min. After four washes, bound antibodies were detected with a goat anti-human IgG immunoglobulin (diluted 1 : 5000) conjugated to horseradish peroxidase (Sigma). Following incubation at 45°C for 1 h and four washes, 100 μl of a substrate solution containing ortho-phenylenediamine and HAll assays were performed by a single investigator (DB), who was unaware of each patient's personal characteristics, and diagnosis. A body-cavity-related B-cell lymphoma cell, BCP-1, which is positive for KSHV and negative for Epstein–Barr virus, was used for an indirect immunofluorescence assay to detect IgG antibodies against KSHV antigen . In order to avoid confounding, these factors were examined among HIV seropositive individuals only . All P values are two-sided. Note that numbers of cases and controls in the tables do not always add to the total, because of missing values.Data were computerised by trained clerks using EPI-INFO-5 software, and statistical analyses were conducted using STATA 7.0 . The oddP=0.7) and, 41% of cases and 23% of controls reported their current residence as being in Kampala (Table 1P=0.13). The seroprevalence of anti-HIV-1 antibodies was 70% among cases and 15% among controls . The risk of conjunctival carcinoma was significantly lower among those with a high personal income . For those who left home at ages 21+ years (including those who never left), 15–20 years and 1–14 years, the odds ratio was 1.0 (reference group), 0.7 (0.4–1.5) and 0.4 (0.2–1.0) respectively (Ptrend=0.05). Study participants were asked how long each week they spent cultivating, 0–9 h, 10–19 h or 20+ h. The risk of conjunctival carcinoma increased significantly with increasing time spent cultivating .Among those with conjunctival cancer, 43% (26 out of 60) were men and 57% (34 out of 60) were women. The proportion of all cancers comprising conjunctival carcinoma declined from 9% in those aged 15–24 years to 2% in those over the age of 45 years. Seven per cent of cases and 5% of controls were born in Kampala, the remainder being born outside the capital city (Ptrend=0.2). Only 15 people had anti-HPV antibodies to more than one tested HPV subtype (two cases and 13 controls) and there was no significant excess risk of the tumour in these individuals, as compared to those who were considered to be negative for all three subtypes . In relation to Kaposi's sarcoma-associated herpesvirus, the seroprevalence of anti-KSHV antibodies was 47% (15 out of 32) among cases and 49% (188 out of 384) among controls .Table 2P=0.02). Results for sexual and reproductive variables among HIV seropositive people only, are shown in Appendix 4. A self reported history of having either given or received gifts in exchange for sex was associated with an increased risk of the tumour , although this may have been a chance finding as there was no statistically significant association with any other sexual or reproductive variable, including the reported number of lifetime sexual partners (P=0.4).Further results are provided in Appendices 1–4. Results for other social and demographic factors, none of which was associated with an increased risk of conjunctival carcinoma, are shown in Appendix 1. Factors, other than income, that might characterise wealth and social status in Uganda, such as ownership of livestock and travel away from home, are shown in Appendix 2. None of these variables were associated with conjunctival carcinoma. Appendix 3 shows results for other possible exposures. Those who reported having had a blood transfusion in the past were less likely to be cases  was available on 60% of cases  and 63% of controls. Typical of studies in developing countries where laboratory services are limited, this proportion compares favourably with other cancer series reported from Africa . Also, tAnother potential source of bias arises from the fact that most  cases had been identified from out-patient clinics, which tend to draw patients from the local area, whereas controls had been hospital in-patients, who are from a wider area. Despite adjusting for region of residence in the analyses, the possible impact of this on the results, is difficult to assess.Case reports of squamous cell carcinoma of the conjunctiva in human immunodeficiency virus (HIV) infected men in the USA and France, coupled with a marked increase in the numbers of tumours being seen by ophthalmologists in at least two African centres (which mirrored the increases seen for Kaposi's sarcoma), led to the suggestion of an association with HIV . HoweverThe mechanism whereby HIV infection increases the risk of conjunctival cancer is not clear. There is no evidence that it is directly oncogenic and the impact on cancer risk is most likely to be mediated via immunosuppression, as is the case for other HIV-associated cancers . In geneSeveral types of squamous carcinoma are associated with human papillomavirus (HPV) infection, most notably cancer of the uterine cervix, induced primarily by HPV-16, -18 and others. Squamous carcinoma of the skin has also been associated with HPV-5 and -8 in immunosuppressed individuals . EvidencThe HPV assays used in this study are based on the expression of L1 major capsid proteins of HPV-16, -18 and -45 in insect cells by using recombinant baculoviruses (Although relatively rare everywhere, conjunctival carcinoma is more frequent in sub-Saharan Africa and other tropical areas than in temperate countries, leading to the hypothesis that it may be associated with exposure to solar ultraviolet radiation . GeograpIn summary, the current study gives independent support for the strong epidemiological evidence that solar ultraviolet radiation is an important cause of squamous cell carcinoma of the conjunctiva. Another established risk factor is HIV infection, although the mechanism whereby it increases the risk of conjunctival cancer is not clear. We find little evidence supporting the possible role of sexually transmitted forms of HPV in the aetiology of conjunctival cancer, but larger studies are required.


This paper aimed at studying the transplacental transmission of HPV and looking at the epidemiological factors involved in maternal viral infection. The following sampling methods were used: (1) in the pregnant woman, (a) genital; (b) peripheral blood; (2) in the newborn, (a) oral cavity, axillary and inguinal regions; (b) nasopharyngeal aspirate, and (c) cord blood; (3) in the placenta. The HPV DNA was identified using two methods: multiplex PCR of human β-globin and of HPV using the PGMY09 and PGMY11 primers; and nested-PCR, which combines degenerated primers of the E6/E7 regions of the HPV virus, that allowed the identification of genotypes 6/11, 16, 18, 31, 33, 42, 52 and 58. Transplacental transmission was considered when type-specific HPV concordance was found between the mother, the placenta and the newborn or the mother and cord blood. The study included 49 HPV DNA-positive pregnant women at delivery. Twelve placentas  had a positive result for HPV DNA. Eleven newborn were HPV DNA positive in samples from the nasopharyngeal or buccal and body or cord blood. In 5 cases  there was HPV type-specific agreement between genital/placenta/newborn samples. In one case  there was type specific HPV concordance between genital/cord blood and also suggested transplacental transmission. A positive and significant correlation was observed between transplacental transmission of HPV infection and the maternal variables of immunodepression history . In conclusion the study suggests placental infection in 23.3% of the cases studied and transplacental transmission in 12.2%. It is suggested that in future HPV DNA be researched in the normal endometrium of women of reproductive age. The possible consequence of fetal exposure to HPV should be observed. Human papillomavirus (HPV), the most common sexually transmitted infection, has been recognized as a cause of anogenital warts HPV type 6 and 11) and cervical cancer -6. and 11 aAlthough it has been established that HPV is sexually transmitted,8, thereThis cross-sectional, prospective study aimed at evaluating transplacental transmission of HPV and enhancing understanding of the maternal epidemiologic features involved.Between April 2005 and April 2007, a cross-sectional, prospective study was performed on 71 pregnant women  with a prior history of HPV infection (n = 22), or who had abnormal Papanicolaou smear (n = 20) or genital warts (n = 29), due to the high probability that they could have HPV infection. The women were referred from the Obstetrical Service of the University of Caxias do Sul and by the Basic Health Units of the Single Health System in Caxias do Sul. This study was performed with the approval of the Ethics in Research Committee at the University of Caxias do Sul, and of the Editorial and Scientific Board of the General Hospital of Caxias do Sul, and did not present a conflict of interest. The Letter of Free and Informed Consent and the epidemiological evaluation tool were obtained from all the women by individual interviews during the obstetrical examinations. Sixty-three (79.7%) of the 71 pregnant women selected who entered the study underwent delivery and 16 (20.3%) dropped out of the study.The epidemiological study was performed taking the following variables into account: age, race, level of education, smoking, marital status, age at first sexual   intercourse, parity, number of sexual partners in lifetime, number of sexual  partners in past year, frequency of condom use with sexual partners in lifetime, frequency of condom use with sexual partners in past year, marital stability in years, history of immunodepression (HIV – acquired immunodeficiency syndrome), type of HPV lesion , site of HPV lesion , type of HPV infection , gestational age at the time HPV infection was diagnosed (weeks), duration of labor (minutes), time of amniotic membrane rupture (minutes), type of delivery  and HPV lesion at delivery .The maternal genital samples were obtained during pregnancy, at the first visit, when the woman was recruited. The sample was obtained using a special brush for cytopathological sampling of the cervix, which was used for genital brushing in the following order: cervix and possible clinical and subclinical lesions of the vagina, vulva and perineal region. The brush was placed in a TE solution , and the material collected was kept frozen at -20°C, until the desoxyribonucleic acid (DNA) was extracted.Immediately before delivery (pre-partum period), a sample of peripheral blood was obtained from the woman using a 3 ml disposable syringe (27/5 needle), retrieving about 1 ml of blood which was placed in a KMA type tube with EDTA. The blood collected was kept frozen at -20°C, until DNA was extracted.In newborns, in the first minutes of the life, buccal, body, nasopharyngeal aspirates and arterial blood from the umbilical cord samples were obtained.The swabs were collected in the first minutes of life, using the special brush for cytopathological sampling of the cervix, with which brushing was performed in the following order: buccal cavities, axillary and inguinal regions of the newborn. The brush was placed in a TE solution  and kept frozen at -20°C, until DNA was extracted.® Linha Sondas Descartáveis Mercosul®, Empresa CPL Medical's Produtos Médicos LTDA), used to aspirate the upper airways  of the newborn immediately after birth, was removed. The distal extremity of the catheter (about 4 cm long) was cut and placed in TE solution , keeping it frozen at -20°C, until DNA was extracted.The distal extremity of the tracheal aspiration catheter  to obtain about 1 ml of fetal blood. The collection was performed after clamping the cord and complete delivery of the placenta and fetal membranes. The fetal blood was placed in a KMA type tube with EDTA and frozen at -20°C, until DNA was extracted.The placental sampling methods were performed immediately after complete delivery and cleaning of the placental disk sides, using surgical compresses.The swabs were obtained using special brushes for the cytopathological collection from the cervix, by brushing in the following order: initially on the fetal side of the placenta, and later with a new brush, on the maternal side of the placenta. The brushes were placed individually in a TE solution , keeping them frozen at -20°C, until DNA was extracted.Two biopsies were performed on the sides of the placental disk: one in the more central portion; another in the more peripheral portion . The biopsies were performed with the help of the rat-tooth forceps, and the curved iris scissors. The fragments collected were placed individually in a TE solution  and kept frozen at -20°C, until DNA was extracted.Wizard Genomic DNA Purification Kit (Promega), according to the manufacturer's specifications. In the brush samples, DNA was extracted using 600 μl of NaOH 50 mM stirred in a vortex for 5–10 seconds and later incubated at 95°C for 5 minutes. The solution was then neutralized with 60 μl of Tris HCl pH 8.0 and kept in a freezer at -20°C, until it was submitted to the next stages.DNA was extracted from the blood and tissue samples using the After the DNA extraction methodology, the products were submitted to two different PCR methods to identify and type the HPV DNA: multiplex PCR and type specific nested multiplex-PCR.Human Papillomavirus maThe positive and significant correlation between placental HPV infection and the maternal epidemiological variable "history of immunodepression" , may be related to the special characteristics of the gravid cycle, especially the changes in the hormonal and immunological balance prevailing during this period, which might favor placental HPV infection.Although the nested multiplex PCR methodology is used to identify and type only 9 types of HPV shown as the most prevalent in the city of Caxias do Sul, it performed very well in identifying maternal HPV DNA, and also considerably increased the number of pregnant women with multiple virus infections. In the newborn and placental samples the nested multiplex PCR method showed its great sensitivity and specificity to identify HPV. The use of that method was also crucial to evaluate the concordance of type specific HPV DNA among the maternal/placental/newborn samples, thus defining the vertical and transplacental transmission rates of the virus.Concluding, the HPV DNA detection rate in the placenta was 24.5% (n = 12/49) and the transplacental transmission rate was 12.2% (n = 6/49). A transplacental transmission rate of 54.5% (n = 6/11) was observed when only the cases of vertical transmission were analyzed. These results were achieved in analyses of the placentas and newborns of mothers with genital warts or intraepithelial lesions of the uterine cervix. Thus, different forms of management can be adopted for each of these different stages , both from the diagnostic and therapeutic perspective. The mother and the newborn must be observed clinically and educational preventive measures must be established concerning the forms of HPV transmission, besides effective strategies for specific immunization.In future, the HPV DNA rates in must be observed in the normal endometrium of women of reproductive age, in order to explain the possible route of infection by continuity and/or contiguity between the endometrium and conception products.


Although human papillomavirus (HPV) infection in heterosexual couples has been sparsely studied, it is relevant to understand disease burden and transmission mechanisms. The present study determined the prevalence and concordance of type-specific HPV infection as well as the determinants of infection in heterosexual couples in a rural area of Mexico.A cross-sectional study was conducted in 504 clinically healthy heterosexual couples from four municipalities in the State of Mexico, Mexico. HPV testing was performed using biotinylated L1 consensus primers and reverse line blot in cervical samples from women and in genital samples from men. Thirty-seven HPV types were detected, including high-risk oncogenic types and low-risk types. Multivariate logistic regression models were utilized to evaluate factors associated with HPV.The prevalence of HPV infection was 20.5% in external male genitals and 13.7% in cervical samples. In 504 sexual couples participating in the study, concordance of HPV status was 79%; 34 partners (6.7%) were concurrently infected, and 21 out of 34 partners where both were HPV positive (61.8%) showed concordance for one or more HPV types. The principal risk factor associated with HPV DNA detection in men as well as women was the presence of HPV DNA in the respective regular sexual partner . In men, having a history of 10 or more sexual partners over their lifetime  and having had sexual relations with prostitutes  increased the likelihood of detecting HPV DNA.In heterosexual couples in rural regions in Mexico, the prevalence of HPV infection and type-specific concordance is high. High-risk sexual behaviors are strong determinants of HPV infection in men. Although there is clear evidence for the influence of the male factor in the development of cervical neoplasia,2, HPV tThe goal of the present study was to determine genital HPV prevalence in sexual couples, evaluate HPV type-specific concordance and the association of known risk factors of HPV infection in a low-risk, predominantly monogamous population.We conducted an HPV DNA prevalence and type-specific concordance study in 504 heterosexual couples attending first-level health centers for medical attention in four municipalities belonging to the Texcoco Sanitary District in the State of Mexico, in the central region of the Mexican Republic. Three of the municipalities had rural characteristics  and one was semi-urban . The study period was November 2002 to September 2003.The study partners were identified and selected using convenience sampling in healthcare centers. First, women who consistently sought care in health centers for diverse reasons were identified and invited to participate. The women who were accepted into the study were asked to invite their regular sexual partner to participate . Subjects were invited to participate after a talk about HPV infection and its association with cervical cancer given at the participating healthcare centers. Since the professional occupation for 65% of this population is agriculture, the study was conducted during the morning hours to increase the likelihood that the male sexual partner would attend.Sexual couples were included when both partners were available to participate in the study; excluded were sexual couples in which the female partner was pregnant or had a hysterectomy.Male partners were instructed not to wash their genitals for at least 12 hours prior to the examination and to be sexually abstinent for three days. Female partners were asked to be sexually abstinent and not menstruating.After receiving written informed consent guaranteeing confidentiality, and in complete privacy, a self-administered questionnaire was completed to obtain information about socioeconomic variables, educational level, smoking habits, reproductive history, use of contraceptive methods and sexual behavior factors. The couples answered the questionnaire in separate locations. The questionnaires and collection of biological samples for each partner were carried out on the same day at the corresponding health center. The study was approved by the ethical and research committees at the institutions that participated in the study. The overall response rate was 60%.The methods used to collect the samples from the male genital area have previously been described. BrieflyIn women, a sample of epithelial cells was taken from the exocervix and endocervical canal using a nylon cytobrush, which was rotated 360°C to assure sampling of the cervical transformation zone. All genital samples were collected by a trained doctor. All brushes containing the collected material were placed in a 5 ml aliquot of phosphate-buffered saline (PBS)/merthiolate 0.01% (v/v). The samples were maintained at -20°C for an average of 30 days, until their delivery to the laboratory. The samples were also stored at -20°C in the laboratory prior to DNA extraction.Previous to DNA extraction all samples which arrived at the lab were centrifuged at 4500 rpm for 6 minutes, after pellet was suspended in 1 ml of 0.01 M TRIS HCL pH 7.4. Briefly, genital samples were treated with proteinase K (170 ug/ml). The DNA extraction was performed with phenol-chlorophorm/isoamyl alcohol 24:1, then NaCl 5 M was added and precipitated with isopropanol. Finally pellet was suspended in 50 μl of buffer TE pH 7.6 and stored at -70°C .Was performed using DNA hybridization test as described by Gravitt et al . After sHPV detection and genotyping was performed on the products of PCR (inverse hybridization) which utilized the nylon membranes that were used for the hybridization. Each membrane contained 39 test lines, 37 of which correspond to type-specific HPV and two to the quantification of low and high concentrations of β-globin. HPV types that were considered high-risk for the development of cervical neoplasia are 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66; low-risk are 6, 11, 26, 40, 42, 53, 54, 55, 61, 62, 64, 67, 68, 69, 70, 71, 72, 73, 81, 82, 83, 84, IS39, CP6108. The hybA stratified analysis was conducted by sex and a socioeconomic level index (SLI) was created using principal component analysis with the variables education, floor material in the dwelling, availability of potable water, drainage availability, owning a vehicle and owning domestic electronic equipment such as a television, video cassette recorder, gas stove and water heater. The index obtained was categorized in tertiles to define low, medium, and high SLI. The McNemar's test was used to compare the prevalence of HPV among men and their sexual partners.The concordance of HPV status between sexual partners and between HPV risk groups (high- and low-risk) was evaluated based on contingency tables using the Kappa statistic. Concurrence of HPV infection in heterosexual couples was defined as the presence of infection in both of the sexual partners, independently of whether or not there was HPV type concordance. For couples where both partners presented with at least one type of HPV infection, percentages for type-specific positive concordance were obtained and, for those with type-specific concordance, it was determined whether the partners were concordant for one type, two types, or three types. The association of potential determinants of HPV positivity in both men and women was evaluated using logistic regression modeling adjusted for age and SLI, obtaining odds ratios (OR) with 95% confidence intervals. All p-values were two-sided.The prevalence of HPV infection was 20.4% in men and 13.7% in women. The most frequently detected high-risk types were HPVs 59, 18, 39 and 16 among men, and HPVs 59, 16, 31, 52 and 58 among women. The most common low-risk types were HPVs 61, 62, 53, 84 and 81 among men, and HPVs 62, 71, 81 and 54 among women. Overall, the pattern of HPV type distribution was similar among men and women . In 69(50%) of these only the man presented some type of infection, in 35(25.4%) only the woman, and in 34(24.6%) both were infected. Among heterosexual couples in which both partners were infected, 21(61.8%) showed type-specific concordance in one or more HPV types. Overall concordance was statistically significant . The most frequent HPV types found in both partners who presented type-specific concordant infection were HPVs 59, 62, 54 and 39  compared to living in a rural area. In addition, being single  and having less than 7 years of schooling  were variables significantly associated with an increased risk for penile HPV infection. For women, not having a stable partner was associated with a statistically significant increase in the risk of cervical HPV infection , as was being a current smoker . Age, years of schooling, SLI, and religion were not associated with the presence of cervical HPV infection in women   as was having had 10 or more lifetime sexual partners , a history of having had sexual relations with prostitutes  Among women, 4% had never been pregnant, 37.3% never had a cervical cytology, and 29.4% did not use any contraceptive method. An increased risk of cervical HPV infection was observed  in women whose male partners had sexual relations with prostitutes while living together, as was for those whose partners did not use condoms while having relations with prostitutes . In women, no statistically significant associations were found between sexual behavior characteristics and HPV detection (data not shown).The presence of any HPV type infection in men was strongly associated with the presence of HPV infection in their female sexual partners  Table .This work describes one of the first studies in a Mexican population that evaluates HPV type-specific concordance among heterosexual couples in a rural area in central Mexico. Of 138 couples where at least one partner was infected, approximately 25% (34/138 = 24.6) of the respective partners were simultaneously infected by HPV. Among these couples, type-specific concordance was high (61.8%). The principal predictors of HPV in men were factors related to high-risk sexual behavior. The presence of HPV in both men and women was strongly associated with the detection of HPV in their respective partners.Studies over the past 20 years evaluating HPV infection concordance among heterosexual partners have shown many inconsistencies, reporting concordances of type-specific infection of between 2 and 87%,8,16-21.An early report on HPV concordance in heterosexual partners documented that 75% of women whose male partners had HPV were also HPV positive, while only 39% of men with HPV-positive female partners were HPV positive in semen. Female Other studies have recently shown concordance findings to be similar to those found in this study; 76% of male partners of infected women have been shown to be HPV positive. Three oAn association between the presence of lesions in the sexual partner and the presence of HPV infection was not demonstrated in the current study as the large proportion of infections in this population was subclinical. It is possible that the combined sampling of sites of the scrotum and penile shaft, balano-preputial lamina and urinary meatus has increased the type-specific concordance value found in this study. It has been shown in a previous study that use of combined samples increases HPV DNA detection.The 13.7% prevalence of HPV infection found in women is less than the prevalence of 20.4% found in men; this lower prevalence in women compared to their male partners has been observed in other studies that evaluate both men and women. The natThe above is a reflection of the fact that HPV prevalence of and concordance among couples is not only highly variable but also depends on the sexual behavior of the couples, the sensitivity of the tests employed, and more importantly on the differences between acquisition rates among men and women.The identification of risk factors associated with HPV detection in our study in both men and women is consistent with that of other reports-48. In mThe results of this study of couples are singularly important, in part, because this is one of the first studies to quantify type-specific HPV concordance for a female population with a pattern of sexual conduct that is predominantly monogamous. In addition, because HPV prevalence estimates in the external male genitalia was found to be two times higher than that previously reported for Mexican men with low-risk sexual behavior. These findings are bolstered by an external quality control mechanism for the determination of HPV DNA in the study population that included a determination of HPV blind to knowledge of gender and of the HPV results of the corresponding partner. Therefore, information bias with regard to the characterization of the presence of HPV DNA is improbable.Prospective cohort studies among different populations are warranted to confirm these estimates as well as to quantify the probability of HPV transmission patterns in men and women and explore the role of potentially associated cofactors.The authors declare that they have no competing interests.RP participated in the design of the study, supervised the trial and data acquisition process, RM performed the statistical analysis, interpreted the data and wrote this paper, ARG and XC analyzed and interpreted the data, AC participated in the design of the study, ELP conceived of the study, participated in its design, coordination and wrote this paper. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/10/223/prepub


Education campaigns seeking to raise awareness of human papillomavirus (HPV) and promoting HPV vaccination depend on accurate surveys of public awareness and knowledge of HPV and related sexual behavior. However, the most recent population-based studies have relied largely on computer-assisted telephone interviews (CATI) as opposed to face to face interviews (FTFI). It is currently unknown how these survey modes differ, and in particular whether they attract similar demographics and therefore lead to similar overall findings.A comprehensive survey of HPV awareness and knowledge, including sexual behavior, was conducted among 3,045 Singaporean men and women, half of whom participated via CATI, the other half via FTFI.Overall levels of awareness and knowledge of HPV differed between CATI and FTFI, attributable in part to demographic variations between these survey modes. Although disclosure of sexual behavior was greater when using CATI, few differences between survey modes were found in the actual information disclosed.Although CATI is a cheaper, faster alternative to FTFI and people appear more willing to provide information about sexual behavior when surveyed using CATI, thorough assessments of HPV awareness and knowledge depend on multiple survey modes. Healthy People 2010 in the U.S = 18.28, p < .01).For men, educational attainment was the only demographic variable found to significantly vary between the surveys. In particular, more lower educated men were in the FTFI group and more higher educated men were in the CATI group . Although more men in the CATI group (17.9%) reported an awareness of HPV than in the FTFI group (14.2%), this difference was not significant.Respondents were asked whether they had heard of HPV, to which they answered "yes', "no", or "don't know". Significantly more women in the FTFI group (21.3%) reported an awareness of HPV than in the CATI group (18.6%)  and occupation  were the only two variables significantly associated with HPV awareness. Awareness tended to increase with greater education . Meanwhile, women who were self-employed (29.0%), students (23.1%), or working in white collar occupations (23.8%) were more likely to have heard of HPV than those performing home duties full-time (17.3%) or working in blue collar occupations (11.4%). However, when differences in the demographic profiles between the survey modes were held constant, women in the FTFI group still displayed significantly greater HPV awareness , therefore suggesting additional reasons for these differences in awareness.Respondents who had heard of HPV were asked 13 questions to assess their overall knowledge, to which they answered "yes", "no", or "don't know". Table Because the demographic profiles varied between the survey modes, multiple regression was used to determine the demographic variables most associated with men's knowledge scores. Language spoken at home was the only variable significantly associated with knowledge scores , with English speakers (M = 7.46) displaying higher scores than Mandarin (M = 6.37), Malay (M = 6.33), Tamil (M = 6.38), and non-Mandarin Chinese (M = 6.30) speakers. However, even when differences in the demographic profiles between the survey modes were held constant, men in the CATI group still displayed better overall knowledge than those in the FTFI group , therefore suggesting additional reasons for survey mode differences in men's knowledge scores.Participants were asked five questions about their sexual behavior, including total number of sexual partners, number of sexual partners over the past 12 months, number of current regular sexual partners, whether they used contraception during first sexual intercourse, and their age when they first had sex. We examined whether their willingness to disclose information to these questions varied according to survey mode.2  = 30.01, p < .001) and men .For all remaining sexual behavior questions, both men and women in the FTFI group were more likely to refuse disclosure than those in the CATI group, although this tendency was greater for women than for men  awarded funds to Suzanne Garland to support this research in the interests of informing their commercial decisions regarding HPV vaccination. GSK, however, had no involvement with the design or data analysis of this study. All other authors declare that they have no competing interests.AS, MP, SC, SG, MLW, and EHT designed the study. AS, MP, and SC supervised the project. AS, AL, MP, SC, and RR analysed and interpreted the data. AL wrote the manuscript and all authors contributed to the final draft. All authors approved this manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2458/9/429/prepub


Around 65% of women with cervical carcinoma in Sweden have not attended an organised screening. We therefore investigated the value of using self-sampling at home in combination with a test for high-risk human papilloma virus (HPV) to increase participation.A total of 2829 women 30–58 years old, who had not attended the organised screening for ⩾6 years, were recruited. They were offered self-sampling at home (Qvintip) and recommended to send the collected vaginal fluid to a laboratory for analysis of the presence of high-risk HPV (Hybrid Capture 2 method).A total of 39.1% of the women accepted home sampling. These women disclosed a relatively high prevalence of high-risk HPV, which decreased with age, from 11.1% in women 30–39 years old to 2.9% in women ⩾50 years . Follow-up disclosed histological cervical intraepithelial neoplasm (CIN) 2–3 lesions in 43.2% of the women with a persistent HPV infection, corresponding to 2.0% of the total number of participating women. The sensitivity of a single smear to detect the histological CIN 2–3 lesions were only 52.6%, even if all abnormal smears –CIN 3)) were included.The use of self-sampling at home in combination with testing for high-risk HPV increases the participation rate of the organised screening and detects almost twice as many women with pre-malignant cell alterations (CIN 2–3) in comparison those with a single cytological smear. A recent nationwide investigation in Sweden disclosed that around 65% of all cervical carcinomas occur in women not attending the organised gynaecological screening and that around 25% are found in women participating in the screening, but with a series of normal cytological specimens preceding the tumour diagnosis. Thus, non-participation and the relatively low sensitivity of cytology are the two major disadvantages associated with the organised screening  is the major factor for tumour transformation and that DNA of oncogenic (high-risk) HPV types is present in both invasive carcinoma and the pre-malignant mucosal progenitors. As HPV DNA tests are more sensitive than cytology to detect pre-malignant alterations in the cervix, this method is now considered as an adjunct or an alternative to cytology as a preventative screening method . From this data base 3000 women, who were 30–58 years old and had not attended the organised screening for ⩾6 years, were identified. Among the 3000 women selected, 171 had to be excluded because of an incorrect address or a previously performed hysterectomy.The remaining 2829 women received an information letter and a form by which they could order a device  to be used for self-sampling of the vaginal fluid at home. The collected material was to be returned in a prepaid envelope to the laboratory, where a test for high-risk HPV was performed. After 3 weeks the women received a letter reminding them to order the self-sampling device. Women who had ordered the device received a letter after about 2 months reminding them to perform the self-sampling of the vaginal fluid. A few women who sent in a sample for analysis more than 3 months after the launch of the study were not included in the investigation.All participating women received a return letter with the results of the HPV analysis. Women who turned out to be HPV positive were informed that they were going to be called to the Department of Gynaecology for further examination within 3–6 months.μ thin sections, which were stained with haematoxylin–eosin stain. The sections were examined in a light microscope and grouped into normal (including cervicitis and metaplasia), cervical intraepithelial neoplasm (CIN) 1 (including condylomas), CIN 2 and CIN 3.At the gynaecological surgery, women were examined by colposcopy and material was collected for histology, cytology and a repeated HPV test. The histological material was fixed in 10% buffered formalin, embedded in paraffin, sectioned in about 4-The cytological smears were fixed in 95% ethanol and stained with Pap stain before examination. The smears were grouped into normal, atypical squamous cells of unknown significance (ASCUS), CIN 1, CIN 2 and CIN 3.–1, which is proportional to the light emission of the positive control and corresponds to 5000 HPV genomes per specimen in the well.The test for high-risk HPV  identifies 13 high-risk HPV types . The Digene hc2 technique can detect HPV DNA concentrations over 1 pg mlThe self-sampling device was ordered by 1609 women (56.9%) and 1107 (39.1%) performed sampling of the vaginal fluid and sent the material to the laboratory for testing for high-risk HPV. An HPV-positive reaction was obtained in 6.7% (74 out of 1107) of the samples. There was no significant age difference with regard to participation rate. However, the HPV prevalence decreased with age. It was 11.1% in women 30–39 years old and 2.9% in women ⩾50 years .Most women decided to order the self-sampling device within a few weeks and after 8 weeks 1600 women had ordered the device. However, after obtaining the device the decision to collect a vaginal fluid sample was slower, 970 samples were obtained within 12 weeks, and even after that time occasional samples were sent to the laboratory .The 74 women with a high-risk HPV-positive reaction were admitted to a gynaecological surgery for further examination. However, at that time seven women had moved out of the county and four had chosen to visit a midwife surgery. A repeated test for high-risk HPV, a cervical biopsy and a cytological smear were obtained in 60 of the 63 remaining women. A persistent high-risk HPV infection was seen in 73% (44 out of 60) of the women, 43.2% of which (19 out of 44) showed CIN 2–3 alterations in the cervical biopsies . The preA biopsy was obtained in 63 of the 74 HPV-positive women and 22 biopsies showed histological CIN 2–3 lesions, corresponding to 2.0% (22 out of 1107) of the total number of women performing self-sampling of vaginal smear. The prevalence of CIN 2–3 was 2.9% among participating women under 40 years and 1.1% among women over 40 years.The cytological smears taken concomitant with the cervical biopsies at the gynaecological examination showed a normal picture in 75% (45 out of 60) of the cases and various kinds of cell alterations (ASCUS–CIN 3) in 25% (15 out of 60). Among the 19 women with CIN 2–3 in the biopsies, the cytological smear was normal in 47.4% (9 out of 19) of the cases. In only four (21%) of the cytological slides CIN 2–3 cell alterations were recorded .In countries with an organised gynaecological screening, the majority of cervical cancer cases occur among women who have not attended midwife surgeries for collection of a cervical smear . The parIn addition to the increased coverage obtained by the self-sampling method, its combination with a test for high-risk HPV makes it considerably more sensitive in comparison with a cytological examination. This means that in addition to an increase in coverage, the women are offered an analytical method that is more relevant to identify women with an increased risk to develop cervical cancer as a result of a persistent high-risk HPV infection. That a single cytological examination has a low sensitivity is obvious from this investigation, in which 53% of the women with a biopsy-verified CIN 2–3 lesions on the cervix showed normal cytology. This also explains why around 25% of all cases of cervical cancer occur among women regularly participating in the organised screening . This means that around twice as many CIN 2–3 lesions are detected with self-sampling combined with testing for high-risk HPV compared with cytological screening.This study shows that 43% of the women with a persistent HPV infection had histological CIN 2–3 lesions on the portio. There was no obvious age difference, although the number of women with CIN 2–3 was slightly higher in women <50 years. In total, 2.0% of all the participating women in the study showed histological CIN 2–3 lesions. In comparison, 0.9% of all cytological smears in the organised gynaecological screening in Sweden show CIN 2–3 and the number of histologically verified CIN 2–3 lesions is about the same (In summary, self-sampling at home in combination with a test for high-risk HPV will increase the number of women participating in the organised screening for cervical cancer, and also identify around twice as many CIN 2–3 lesions for treatment with conisation compared with cytological screening.


Human papillomavirus diversity in high-grade lesions and SCC on cytology suggests that HPV 16- and 18-based vaccines may not be adequately polyvalent to induce protective immunity in this population.We screened 145 HIV-infected non-pregnant women at a tertiary care centre in Lusaka, Zambia. Liquid-based cytology and human papillomavirus (HPV) genotyping with PGMY09/11 biotinylated primers (Roche Linear Array® HPV genotyping test) maximised sensitivity of cytology and HPV assessments. Among high-risk (HR) types, HPV 52 (37.2%), 58 (24.1%) and 53 (20.7%) were more common overall than HPV 16 (17.2%) and 18 (13.1%) in women with high-grade squamous intraepithelial lesions or squamous cell carcinoma (SCC) on cytology. High-risk HPV types were more likely to be present in women with CD4 P=0.02). We were able to study HPV genotypes on stored samples from these women.Increasing numbers of HIV-infected women in sub-Saharan African settings like Zambia are now accessing life-prolonging antiretroviral therapy (ART)  at room temperature (37°C) for <2 weeks before being batch transported for cytological analysis.Slides prepared from the PreservCyt® vials were analysed by the liquid-based ThinPrep® cytological imaging system . All slides were initially screened by a senior cytotechnologist and all abnormal results and 10% of negatives were subsequently reviewed by a board-certified cytopathologist. The cytology results were classified and reported as no abnormality detected , atypical squamous cells of undetermined significance (ASCUS), low-grade squamous intraepithelial lesions (LSIL), HSIL and suspicious for squamous cell carcinoma (SCC).We performed HPV typing by polymerase chain reaction (PCR)-based amplification of target DNA using the Linear Array® HPV genotyping test (LA-HPV) , an enhanced and commercialised version of the PGMY line blot assay (PGMY-LB)  during the same visit unless a result was available within 3 weeks of the visit date from our rigorously certified laboratory .All participants underwent visual inspection of the cervix with acetic acid and diagnostic colposcopy during the same clinic visit. A CD4+ cell counts <200 μ l−1 against those of women with CD4+ cell counts ⩾200 μl−1, the threshold to make clinical decisions for initiating ART in persons without symptoms in Zambia. Similarly, we calculated OR and 95% CIs for comparing the type-specific HPV prevalence in women with HSIL or SCC on cervical cytology and those with lower grade  or no cytologic abnormalities. Statistical analysis was carried out using SPSS 14.0 for Windows™ . We restricted our analysis to 145 participants with complete data both on CD4+ cell counts and individual HPV genotypes.Using unconditional logistic regression to calculate prevalence OR and 95% CIs, we compared the type-specific HPV prevalence in HIV-infected women with CD4The mean and median ages of these 145 participants were 36.2 years (s.d.: ±6.3) and 36 years (IQR: 31–41), respectively. About a third  were married and cohabiting with their husband, just under a half were educated beyond high school  and a majority  reported a family income of less than 500 000 Zambian kwacha (∼US$110) per month. Little over a third  reported their age of first sexual intercourse as less than 18 years, and 116 out of 145 (80%) reported between one and five lifetime sexual partners.Nine (6.2%) women had cytologic results within normal limits. Atypical squamous cells of undetermined significance was reported in 25 (17.2%), LSIL in 34 (23.5%), HSIL in 49 (33.8%), and SCC in 28 (19.3%) participants. Thus 93.8% of these HIV-infected Zambian women seeking medical care had abnormal Pap smears, 76.6% had SIL or SCC, and 53.1% had high-grade SIL or SCC.+ lymphocyte counts were 208 μ l−1 (s.d.:±177.5) and 165 μ l−1 (IQR: 85–299) respectively. Almost two thirds of the participants  had a CD4+ count <200 μ l−1, 43 (29.6%) had CD4+ counts between 200–499 μ l−1 and 11 (7.5%) had CD4+ cell counts ⩾500 μ l−1. Validation of results was confirmed with appropriate negative and positive controls for PCR amplification and β-globin gene detection. Human papillomavirus DNA was detected in 141 out of 145 (97.2%) participants, while four (2.8%) women had no evidence of cervicovaginal HPV DNA. High-risk HPV was detected in 131 (90.3%) women. We found a single HPV genotype in 14 (9.7%) women and multiple (⩾2) HPV genotypes in 127 (87.5%) women. The median number of HPV types per woman was 4 (IQR: 2–6), while the mean was 4.4 (s.d.: ±2.8). Of the 37 HPV types identifiable by the LA-HPV test, all except type 64 were detected in the 145 samples. The test identified 656 distinct HPV infections in 145 women, of which 350 (53.3%) were HR-HPV while 306 (46.6%) were LR or UR-HPV types. None of the women carried a HR-HPV type exclusively . Among the HR types, HPV 52 was found to be the most prevalent (37.2%). However, the probe for detection of HPV-52 amplicons in the LA-HPV test is a cross-reactive probe that also hybridises with types 33, 35, and 58. It was observed that at least one of these three HR-HPV types was present in 54 (37.2%) samples along with HPV 52, while the latter was present exclusively in 11 (7.6%) samples. Human papillomavirus 58 was present in 35 (24.1%) and HPV 53 in 30 (20.7%) samples while types 16, 35 and 45 were equally common (17.2%). Human papillomavirus type 18 (13.1%) was rank-ordered 10th among high risk types.Mean and median CD4Among the 28 women with SCC on cervical cytology, at least one HR-HPV type was present in all women while multiple (⩾2) genotypes were present in 22 (78.6%) women. Human papillomavirus type 52 (mixed probe as described above) was the most prevalent in 13 (46.4%), while types 58 and 16 were present in 10 (35.7%). The other HR types in decreasing order of their prevalence in women with SCC on cytology were HPV types 35 (28.6%), 53 (25%), 31 and 51 (21.4% each), 18 and 45 (17.9% each), 33, 56, 59, and 68 (14.3% each), 39, 36, and 66 (10.7% each), and 73 (3.6%). High-risk type 82 was not detected with SCC. Among LR or UR HPV types, types 62, 61, 70, and 84 had a prevalence of greater than 15% in cases with SCC on cytology but none of these types were present exclusively without the concurrent presence of a HR-HPV type.+ counts <200 μ l−1 than with higher counts . Barring HPV 16 and 39, all HR types showed greater prevalence in women with CD4+ counts <200 μ l−1 but only type 51 achieved statistical significance (P=0.05), given our sample size of 145. Among HPV-types of UR, HPV 71 was more likely to be present in women with CD4+ counts <200 μ l−1 . High-risk human papillomavirus 58 and 31 were significantly more likely in women with HSIL or SCC (P=0.01 and 0.02 respectively). No other high, low, or UR type was significantly higher in prevalence in women with HSIL or SCC than in other women  is one of the highest reported among an HIV-infected (or any other) population of women worldwide. Also, HPV types 16 and 18 were not the most prevalent HR-HPV types and a relatively high frequency of HPV types 52, 58, 35, 53, 31, 51, and 45 was observed in women who had HSIL lesions or SCC on cytology. This finding has implications in the eventual implementation of prophylactic HPV vaccines based on HR-HPV types 16 and 18 . Our res+ cell counts in our participants, nor any association between LR/UR HPV types and presence of HSIL or severe abnormalities.We noted that the majority (87.3%) of participants carried multiple HPV genotypes and that the mean number of HR types increased with increasing immunosuppression. Comparable studies using PCR-based detection of HPV in HIV-infected women have reported 12–79% of study participants with multiple HPV types (The PGMY09/11 primers used in the LA-HPV test have been validated for reproducibility and accuracy in diverse populations (The results reinforce the importance of ensuring adequate cervical cancer screening services for HIV-infected women in resource-limited settings like Zambia. They are important given the recent availability of prophylactic HPV vaccines. Considering the diversity of HPV types found, HPV vaccine constructs with polyvalency would be needed for primary prevention of cervical cancer among HR women in developing country settings.