High grade HPV infections and persistence are the strongest risk factors for cervical cancer. Nevertheless other genital microorganisms may be involved in the progression of HPV associated lesions.Chlamydia trachomatis was identified by strand displacement amplification (SDA) and the other microorganisms were detected by conventional methods.Cervical samples were collected to search for human Papillomavirus (HPV), bacteria and yeast infections in gynaecologic outpatients. HPV typing was carried out by PCR and sequencing on cervical brush specimens. C. trachomatis and Ureaplasma urealyticum (at high density) detection, whereas no correlation was found between HPV infection and bacterial vaginosis, Streptococcus agalactiae, yeasts, Trichomonas vaginalis and U. urealyticum. Mycoplasma hominis was isolated only in a few cases both in HPV positive and negative women and no patient was infected with Neisseria gonorrhoeae.In this cross-sectional study on 857 enrolled outpatients, statistical analyses revealed a significant association of HPV with C. trachomatis was found and interestingly also with U. urealyticum but only at a high colonization rate. These data suggest that it may be important to screen for the simultaneous presence of different microorganisms which may have synergistic pathological effects.Although bacterial vaginosis was not significantly associated with HPV, it was more common among the HPV positive women. A significant association between HPV and  Chlamydia trachomatis [Infection with high grade Human papillomavirus (HPV) plays a central role in cervical carcinogenesis. Epidemiological and molecular investigations have shown unequivocally that high grade HPV infection is the main causal factor in initiating the progressive transformation that leads to cervical intraepithelial neoplasia and to cervical cancer ,2. Many chomatis  and Herpchomatis ,12, althChlamydia trachomatis, Neisseria gonorrhoeae, Streptococcus agalactiae, Mycoplasma spp, Trichomonas vaginalis. Yeasts and bacterial vaginosis were also investigated.The aim of this cross-sectional study was to evaluate the prevalence of co-infection HPV and other genital microorganisms: Women attending clinics for routine gynaecologic care in the Department of Gynaecology, Perinatology and Child Health of "Sapienza" University in Rome, between 2000 and 2006 were included in this study. Informed consent was obtained from all women and this study was approved by the Ethics Committees and/or Institutional Review Boards of the participating institution. Cervical samples were taken by cytobrush from the ectocervix and from the endocervix for cytological analysis, detection of HPV DNA and bacteriological tests. Cytology was classified according to the Bethesda system, using standard forms and classified as negative, atypical squamous cell of undetermined significance (ASCUS), atypical glandular cell of undetermined significance (AGUS), low-grade intraepithelial lesion (LSIL) or high-grade (HSIL) categories . All theE6 gene and part of the E7 gene [Cervical brush samples collected in Phosphate Saline Buffer (PBS) were centrifuged at low speed and the cell pellets underwent DNA extraction using QIAampTissue kit . Human Leukocyte Antigen (HLA)-specific primers were used in polymerase chain reaction (PCR) analysis to assess the suitability of the target DNA (9). A 450 bp fragment from the L1 region of HPV-DNA was amplified using the consensus primer MY09/11 as described elsewhere . Four pa E7 gene , thus alHPV-positive reactions were purified using a QIAquick PCR purification kit (QIAGEN), and an automatic DNA sequencer  was used for sequencing utilising the PCR forward primer(s). Sequence homology was determined using BLAST  and Clustal W  programs.Streptococcus agalactiae (group B) strains were identified from clinical samples by being weakly beta-haemolytic on Columbia CNA Agar A (Oxoid). The suspected colonies were serologically confirmed (Lancefield's classification) [ication) ,17.C. trachomatis using the BD ProbeTec ET Assays  according to the manufacturer's instructions. This method amplifies DNA from C. trachomatis in separate wells and optionally can monitor inhibition of amplification for each specimen using strand displacement amplification (SDA) and detection by fluorescent energy transfer (FRET) probes, producing a method-other-than-acceleration (MOTA) score for each specimen. Specimen processing and BDPT performance followed the manufacturer's instructions. The original algorithm involved retesting specimens with MOTA scores between 2000 and 9999. A negative repeat result (MOTA score <2000) was considered indeterminate.endocervical swabs were tested for the presence of N. gonorrhoeae was carried out by growth on media selective for pathogenic Neisseria species (Oxoid) incubated for up to 48 hours in 5–10% CO2 at 35–37°C. Colonies obtained were identified by API NH (bioMerieux) according to manufacturer's instructions.presumptive identification of Mycoplasma hominis and Ureaplasma urealyticum according to the manufacturer's instructions. The diagnostic kit provided information regarding the presence or absence of M. hominis and U. urealyticum and an estimate of the density of each organism (cut-off 104-colour-changing units- CCU/ml) together with its antimicrobial susceptibilities to doxycycline, josamycin, ofloxacin, erythromycin, tetracycline, pristinamycin, azithromycin, clarythromycin, ciprofloxacin.Mycoplasma IST 2 (bioMérieux) was used for the isolation of C. albicans or C. dubliniensis), microscopic morphology, and growth on Oxoid Chromogenic Candida Agar (Oxoid) [cervical-vaginal samples were cultured on Sabouraud dextrose agar (Oxoid) aerobically at 37°C for 24–48 h ,17. The  (Oxoid) .Non-Candida albicans yeast isolates were further speciated with API 20C AUX or API 32C.the vaginal swab was placed in 0.2 ml of sterile physiologic saline for wet mount evaluation and examined microscopically (400×) within 15 min for motile trichomonads.T. vaginalis and incubated aerobically a 37°C. The broth was examined microscopically for motile trichomonads on days 1, 3 and 5 [The culture was carried out on Oxoid's Trichomonas Medium n.2 for the recovery of  3 and 5 ,17.et al [the presence of bacterial vaginosis was evaluated microscopically on samples collected from the posterior vaginal fornix. Bacterial vaginosis was diagnosed by the presence of three of the clinical and microscopic findings standardized by Amsel et al : vaginal2 test was used in the statistical analysis, while the Odd Ratio was used to measure the strength of the association with HPV. Statistical tests were considered significant if the p value was 0.05 or less. Logistic regression analysis was used to assess the simultaneous effect of more than one variable on the risk of HPV infection and to identify possible confounding factors.The χOut of 857 enrolled outpatients, two groups of patients were identified on the basis of HPV DNA test positivity. The demographic and behavioural characteristics of the study population (266 HPV positive and 591 HPV negative women) are presented in table et al. (2006) [HPV genotypes found in samples from HPV positive women were shown in table . (2006) , the aggC. trachomatis was revealed in 8.05% of the samples, U. urealyticum was present in 28.35% and S. agalactiae in 8.86%; T. vaginalis was found only in 1.16%, bacterial vaginosis in 6.30%, and yeast in 13.06%.In the total population, M. hominis was isolated only in a few cases whereas N. gonorrhoeae was never found.S. agalactiae was found in 9.4% of samples, yeasts in 13.5%, bacterial vaginosis in 5.2%, C. trachomatis in 5.4% and U. urealyticum in 24.8%. Generally U. urealyticum is a common commensal of the female lower genital tract and it seems to be an important opportunistic pathogen during pregnancy and in other genital diseases such as cervicitis. Its pathogenic potential seems to be related to the high density in the analysed samples . In HPV negative patients the percentage of U. urealyticum > 10,000 CCU/ml was 5.24% .The number of the specimens in which S. agalactiae, yeasts, T. vaginalis and U. urealyticum < 10,000 CCU/ml detection.Statistical analyses did not reveal any association between HPV presence and Although the bacterial vaginosis was diagnosed in a higher percentage among HPV positive women (8.6% vs. 5.2% control group), the difference in the prevalence of bacterial vaginosis was not statistically significant between the 2 groups.C. trachomatis (p < 0.001) was observed  (table Moreover another significant association was found between the HPV positivity and the presence of 1) table .Logistic regression analyses did not substantially change these results.Our findings confirm that an increased risk of HPV infection is strongly associated with a young age at first intercourse, and with an elevated number of sexual partners as would be expected for a sexually transmitted viral infection.C. trachomatis has been established [C. trachomatis infection may increase susceptibility to HPV causing microabrasions or alterations of epithelial cells thus facilitating the entry of virions. Some reports have suggested that concurrent C. trachomatis infection can reduce host ability to resolve HPV infection: chronic cervical inflammation seems to influence the HPV persistence through a raised production of free radicals and a reduction of host cell-mediated immunity [C. trachomatis infection induces a shift in the immune response and the unresolved infections have been associated with a humoral (T helper cell type 2) immune response, whereas cellular (T helper cell type 1) immune response is important for the clearance of HPV lesions. Therefore, the modulation of cervical immune response by C trachomatis may influence the clearance of HPV lesions [Epidemiological studies have demonstrated the role of other sexually transmitted infectious agents in the pathogenesis of cervical cancer, but knowledge about the specific role of these pathogens in the natural history of HPV infection is limited. There are several mechanisms by which these co-infections may act, such as direct genotoxicity, but probably the most likely biologic mechanism is the induction of cervical inflammation leading genotoxic damage through oxidative metabolites . An assoablished . In factimmunity ,10. C. tC. trachomatis infection rate; this result may suggest more accurate screening for C. trachomatis in HPV positive patients with an atypical cytology. Since both pathogens are co-variables related to sexual behaviour, probably they synergize in inducing cervical epithelium alterations [Our data showed, in HPV positive patients, particularly in HR HPV positive women, an increased erations ,23.U. urealyticum in HPV positive group was also observed, but a significant association was found only with high density cervical presence. The biological role of U. urealyticum infection in the HPV outcome was not previously clarified, but high level of U. urealyticum seems to be a cofactor in the development of HPV related cervical dysplasia [U. urealyticum may play a role both in initiating cellular anomalies and in viral persistence. It was reported that Mycoplasma infections cause in vitro chromosomal changes and cell transformation throughout gradual progressive chromosomal loss and translocations [An increased infection rate of ysplasia . It is tocations . Robertsocations .Previous researches have produced conflicting results regarding the role of bacterial vaginosis as cofactor in the development of cervical intraepithelial neoplasia . Thus, aC. trachomatis transmission.This study evidenced also the lack of protective precautions against sexually transmitted infections, both in HPV positive and HPV negative women. Despite the use of condom is not reported to be highly effective in reducing HPV transmission ,31, counC. trachomatis was found and interestingly also with U. urealyticum at a high colonization rate. Although bacterial vaginosis was not significantly associated with HPV, it was more common among HPV positive women. These data confirm that screening for genital infections may be important to reveal the simultaneous presence of different sexually transmitted microorganisms. These results suggest and emphasize the value of the screening for genital infections in HPV positive patients in order to decrease the presence of other microorganisms and to reduce the probable synergistic effects of co-infections. Prevention is important not only to avoid other STDs and their sequelae, but also to reduce the influence of concomitant microorganisms on HPV infection.In this study a statistically significant association between HPV and The authors declare that they have no competing interests.RV, AP, EC and AMD conceived the study, collected and interpreted the data and wrote the manuscript; EM, EC and RV collected the samples and analysed the clinical data; JO performed the statistical analyses and drafted the manuscript; RN and FC carried out the bacterial analyses and contribute to draft the manuscript; MB performed the HPV DNA tests and participated in the sequence analysis with AP and AMD; GA participated to the design of the study and was involved in drafting and revising critically 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/1471-2334/9/16/prepub


To provide an accurate evaluation of the association of human papillomavirus (HPV) with lung cancer, 36 cases of lung cancer were analysed for HPV DNAs by polymerase chain reaction (PCR) with dot-blot and Southern blot analyses, and for the transcripts from the E6-E7 transforming region by in situ hybridisation (ISH). HPV-18 DNA was detected in three (8%) of 36 specimens; histologically, in one (10%) of 10 squamous cell carcinomas and two (9%) of 22 adenocarcinomas. Neither HPV-16 nor -33 DNA was detected in any cases examined. Expression of E6-E7 mRNA was confirmed in the cases which contained, HPV-18 DNA. HPV-18 may play an important role in the development and progression of cancer in some cases of both squamous cell carcinoma and adenocarcinoma of the lung.


A number of reports associate human papillomavirus (HPV) with cervical cancer and cancer cell lines derived from this tumour type. Considerably fewer reports have focused on the role of HPV in carcinomas from other sites of female anogenital squamous epithelia. In this study we have tested for the presence of HPV in eight low-passage vulvar carcinoma cell lines and one extensively passaged cell line, A431. One cell line from a primary vaginal carcinoma was included. The presence of the HPV was evaluated by the polymerase chain reaction (PCR), by Southern blot analysis and by two-dimensional gel electrophoresis. General primer-mediated PCR was applied by using primers from the L1 region, E1 region and HPV 16 E7 region. Southern blot hybridisation was performed under low-stringency conditions (Tm = -35 degrees C) using a whole genomic HPV 6/16/18 probe mixture and under high stringency conditions (Tm = -18 degrees C) with the whole genomic probes of HPV 16 and 33. HPV 16 E6-E7 mRNA was assessed by ribonuclease protection assay (RPA). HPV was found in only one vulvar carcinoma cell line, UM-SCV-6. The identified type, HPV 16, was integrated in the cell genome and could be amplified with all primers used. Also E6-E7 transcripts were found in these cells. Five original tumour biopsies were available from the HPV-negative cell lines for in situ hybridisation. All these were HPV negative with both the HPV 6/16/18 screening probe mixture under low stringency and the HPV 16 probe under high stringency. The results indicate that vulvar carcinoma cell lines contain HPV less frequently than cervical carcinoma cell lines and suggest that a significant proportion of vulvar carcinomas may evolve by an HPV-independent mechanism.


Epidermodysplasia verruciformis HPV types were found in 86% of SCC cases and 36% of control subjects (Odds ratio=12.0), suggesting a role of HPVs in the aetiology of SCC.A total of 21 squamous-cell carcinoma of the conjunctiva (SCC) and 22 control subjects had conjunctival samples tested for human papillomavirus (HPV) types using PCR-based assays.  Squamous-cell carcinoma of the eye conjunctiva (SCC) is a rare tumour associated with heavy sun exposure, which has greatly increased in some Sub-Saharan African countries, coincident with the spread of HIV . An incrEpidermodysplasia verruciformis (EV)  . A subgrmis (EV) . Low-rismis (EV) . SquamouBetween March 2000 and January 2001, we conducted a pilot case–control study on 21 SCC cases  and 22 age frequency-matched control patients with benign conjunctival lesions  in New Mulago Eye Department, Kampala, Uganda, where SCC has become the most common ocular malignancy. This aimed to assess the feasibility of a larger case–control study in that location, and to test methods.All patients signed an informed consent and the study was approved by the local Ethical Committee. In total, 10 controls had pterygium, seven pingueculum, four solar keratosis and one pigmented naevus. Tissue specimens from SCC cases and control patients were collected, histologically confirmed, and deep-frozen before shipment to the International Agency for Research on Cancer (IARC), Lyon, France, for HPV testing. To minimise the risk of contamination, the collection of the specimens was performed at different days using disposable material. DNA was extracted from each sample in an IARC laboratory that is exclusively used for this purpose. To monitor the possible occurrence of cross contamination between the different specimens during the extraction of DNA, tubes containing distilled water (one every 10 specimens) were included.HPV detection was performed by means of polymerase chain reaction (PCR) assays using different sets of primers  in sterile conditions, while the PCR and analysis of products were performed in different laboratories. As in the DNA extraction protocol, negative controls were included in the different PCRs. Products obtained with all type-specific primers were also tested by Southern blotting. All analyses, including cellular DNA extraction, PCR, and Southern blotting, were performed in duplicate using consecutive sections of frozen specimens.We first determined the presence of the mucosal HPV types using the general primers (GP) 5+/6+, GP1/GP2 and specific primers for HPV16, HPV18 and HPV45. No mucosal types were found in either cases or controls, but in one SCC case (patient number 21) we detected, by means of GP1/GP2 primers, a novel HPV type closely related to EV HPV type RTRX7  in addition to several undetermined EV HPV types related to HPV38. HPV38 displays The extreme heterogeneity of the cutaneous HPV family and the lack of standard GP for their detection greatly complicate the study of the relationship between the cutaneous HPV types and cancers like SCC. However, the strong association that we found between EV HPV types and SCC points to a possible aetiological role of these viruses.


Sera from 1483 female subjects in England aged 10–29 years were tested. The age-standardised seroprevalence was 10.7%  for human papillomavirus (HPV) 6, 2.7% (1.8–3.6) for HPV 11, 11.9% (10.2–13.6) for HPV 16, 4.7% (3.5–5.8) for HPV 18, and 20.7% (18.6–22.7) for any of the four types. Infection with human papillomavirus (HPV) types identified as ‘high-risk’ is a pre-requisite for developing cervical cancer  utilising virus-like particles have been used successfully for seroprevalence studies in several countries including the USA  as previously described  and positivity for other HPV types.P<0.01). Being seropositive for one type was significantly associated with being seropositive for another (P<0.05), except for the case of HPV 18. HPV 18 seropositivity was only significantly associated with HPV 16 seropositivity (P<0.01) and not seropositivity for HPV 6 or 11. There was no consistent, significant risk of HPV seropositivity associated with sample origin from the North or South of England.Based on our sample, less than 5% of girls under the age of 14 years were seropositive for any HPV type. From age 14 years onwards, the seroprevalence increased sharply until the early 20s, and then stabilised or declined. The proportion seropositive varied by type, being highest for HPV 16 and lowest for HPV 11. Seropositivity for each of types 6, 11 and 18 was significantly associated with seropositivity for type 16, which is responsible for the majority of cervical cancers.Quantitative titres from the study are not comparable to those from studies which use different assays, because currently no international HPV standard reference serum exists, and different laboratories therefore use ‘in-house’ standard sera making direct comparisons nonviable (There are a number of issues to consider before using these data to estimate the incidence of HPV infection. Firstly, seroprevalence is likely to underestimate the proportion of women who have had an HPV infection, since other studies have suggested that only 65–90% of HPV DNA-positive female subjects seroconvert, with differences in estimates depending on factors including the testing systems used, how long HPV DNA persists and whether or not there is progression to disease (The population tested in the survey is not randomly selected but consists of individuals accessing health care in England and having blood samples taken for diagnostic or screening investigations. Previous studies using the same collection have, however, found a consistency between seroprevalence for other vaccine preventable infections and population vaccine coverage data (The apparent decline in seroprevalence observed around the age of 25 years suggests that seroprevalence is not a straightforward marker of all past HPV infection. Such a marker would be expected to continue to rise in older women, since DNA studies show evidence of incident HPV infections in women of all ages, albeit at a lower rate in older women (Despite these complications, these data offer an additional viewpoint on the epidemiology of HPV in England, and a rare view of the age-dependent risk of HPV infection in female subjects who are not selected for their known sexual activity and who are below the age eligible for routine cervical screening and in the age range likely to be targeted with HPV vaccination. They suggest that there is already a substantial risk of HPV infection in girls in England by the age of 14 years, which has implications for the age at which vaccination should be delivered. These data and further analyses should help to determine the most effective strategies for routine and catch-up HPV vaccination.


In Norway, women with negative or low-grade cervical biopsies  are followed up after six months in order to decide on further follow-up or recall for screening at three-year intervals. A high specificity and positive predictive value (PPV) of the triage test is important to avoid unnecessary diagnostic and therapeutic procedures whereas a low risk of high-grade disease among triage negative women assures safety.At the University Hospital of North Norway, cytology and the HPV mRNA test PreTect HPV-Proofer, detecting E6/E7 mRNA from HPV types 16, 18, 31, 33 and 45, are used in post-colposcopy follow-up of women with negative or low-grade biopsy. In this study, women with negative biopsy after high grade cytology (ASC-H/HSIL) and/or positive HPV mRNA test in the period 2005–2009 were included (n = 520). Histologically confirmed cervical intraepithelial neoplasia of grade 2 or worse (CIN2+) was used as study endpoint.Of 520 women with negative or low-grade biopsy, 124 women (23.8%) had CIN2+ in follow-up biopsy. The sensitivity and specificity of the HPV mRNA test were 89.1%  and 92.5% , respectively. The ratios of sensitivity, specificity and PPV of HPV mRNA testing compared to repeat cytology for finding CIN2+ was 1.05 (95% CI: 0.92–1.21), 1.21 (95% CI: 1.12–1.32), and 1.49 (95% CI: 1.20–1.86), respectively. The PPV of mRNA was 77.3%  in women aged 40 or older.Women with negative cervical biopsy require follow-up before resumption of routine screening. Post-colposcopy HPV mRNA testing was as sensitive but more specific than post-colposcopy cytology. In addition, the HPV mRNA test showed higher PPV. A positive mRNA test post-colposcopy could justify treatment in women above 40 years. Cervical cancer is the third most common cancer affecting women worldwide Unfortunately, colposcopy does not have optimal sensitivity for CIN2+. The National Health Service Cervical Screening Programme (NHSCSP) Guidelines for Colposcopy and Programme Management, which guides British practice, ask for evidence of a colposcopic accuracy of 65% Several HPV tests for use in triage and follow-up are available and different hospitals in Norway use different HPV tests. One of these tests is the PreTect HPV-Proofer assay, detecting HPV E6/E7 mRNA from the five most prevalent subtypes causing cervical cancer. The University Hospital of North Norway (UNN) has five years of experience with the use of this test In three previous papers The Regional Committee for Research Ethics (REK Sør-Øst C) has approved the study. Written consent from the patients for their information to be stored in the hospital database and used for research was not needed because the data were obtained and analyzed anonymously. The ethics committee specifically waived the need for consent.In the routine diagnostic practice at the University Hospital of North Norway (UNN), the HPV E6/E7 mRNA test PreTect HPV-Proofer  is used in the triage of women with a Pap-smear showing ASC-US or LSIL and in post-colposcopy follow-up after negative biopsy. The Department of Clinical Pathology receives cervical smears from the population of Troms and Finnmark County. Approximately 23 000 cervical smears are analyzed each year. In 2005–2009, smears from 63 740 women and cervical biopsies from 6 027 women aged 25–69 years were processed. A total of 1 484 women had the cytological diagnoses ASC-H, HSIL or ASC-US/LSIL with a positive HPV mRNA result.The following patients were excluded from the study: women without a biopsy (n = 199); women with CIN2+ on the first biopsy who were referred to treatment (n = 587); and women with one or several abnormal Pap-smears and/or a positive HPV-test at post-colposcopy follow-up but without re-biopsy n = 178, ,Considering the criteria above, 520 women with negative or low-grade biopsy were included in the study. Among these, a liquid based cytology (LBC) follow-up sample was taken for 192 women (36.9%), allowing for additional HPV mRNA testing (HPV group). From the other 328 women (63.1%), a conventional Pap-smear was received, not allowing for HPV analysis  for cytological examination. Of the remnant liquid with cervical cells, DNA/RNA was isolated from 5 ml sample, eluted in 50 µl elution buffer and analyzed with PreTect HPV-Proofer. The mRNA testing was performed according to the manufacturer's instructions (NorChip AS) and in accordance with national guidelines for HPV testing INK4a immunostaining  in order to detect occult CIN lesions.Cytological and histological diagnoses were obtained from the diagnostic database (SymPathy) at the Department of Clinical Pathology, UNN. Biopsies were evaluated by experienced pathologists and histological results were reported using CIN terminology Outcome assessment was based on the histological result of biopsies, where CIN2+ was considered as the target disease and CIN1 and CIN0 (no CIN) were considered as absence of disease. Moreover, women with a complete negative follow-up (two negative post-colposcopy Pap-smears or double negative liquid based cytology (LBC) and HPV mRNA result) were assumed to be free of disease. The clinical sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were calculated in 2×2 tables for post-colposcopy cytology , for the HPV-test alone and HPV mRNA test in combination with cytology (with cut-off ASC-US+ and ASC-H+).http://www.r-project.org/. P-value<0.05 was considered statistically significant.Pearson's Chi square was used to assess associations between test results and final disease status. To assess differences in accuracy, ratios of the sensitivity, specificity and PPV of mRNA testing versus repeat cytology  were computed. The McNemar Chi square test was used for comparisons in sensitivity and specificity in the second follow-up group where both the RNA test and repeat cytology were applied. The Anova test was used to assess differences in average age of the women in the cytology only group and in the HPV-test group. The statistical computations were performed with the software R version 2.9.0 (2009-04-17), In Of the 328 women in the post-colposcopy cytology only group  and 3, 7nd group) compared to repeat cytology (in the 1st group) for finding CIN2+ was 1.05 (95% CI: 0.92–1.21), 1.21 (95% CI: 1.12–1.32), and 1.49 (95% CI: 1.20–1.86), respectively. These findings were consistent with the intra-group comparison of mRNA testing versus repeat cytology at ASC-US+ cutoff: equal sensitivity  and superior specificity of mRNA testing . The detection rate of CIN2+ in follow-up of women with negative biopsy by HPV mRNA positive/negative versus cytology abnormal/normal is illustrated in Of the 192 women with post-colposcopy HPV mRNA test , and 6, HPV mRNA test in combination with cytology with cut-off ASC-US+ had a sensitivity of 97.8%, specificity of 63.0%, PPV of 45.5% and NPV of 98,9% . The HPVOf the 520 women with negative biopsy, 124 women (23.8%) had CIN2+, 72 women (13.8%) had CIN3+ and seven women (1.3%) had cervical cancer in follow-up . Five ofOf the 192 women with post-colposcopy HPV mRNA test, one woman  had cervical cancer. The woman was HPV mRNA positive for HPV type 33. Of the 328 women in the post-colposcopy cytology only group, six women  had cervical cancer. Five of the six women had an abnormal cytology. One woman had normal cytology , but her cervical cancer was detected by hysterectomy with indication uterine leiomyoma (data not shown).Colposcopy and colposcopically-guided biopsies do not have optimal sensitivity for detection of CIN2+. Of the 520 women with negative biopsy, seven women actually had cervical cancer. For these women, the first biopsy or histological slide may not have been representative for the underlying disease. Alternatively, a microlesion difficult to detect by colposcopy, colposcopy-directed biopsies or by evaluation of histology may have been present. Nevertheless, this emphasizes the need for additional methods in order to increase the overall sensitivity of the screening algorithm in the prevention of cervical cancer.The current poor performance of colposcopic impression, colposcopically-guided biopsies and histological diagnosis limits the potential benefit of a very sensitive screening test The main cause of invasive cervical cancer is the deregulated and persistent production of HPV E6 and E7 oncoproteins When summarizing results from all women included in the study , 124 of It is also important to have in mind that in clinical practice, the cytological diagnosis is commonly influenced by the HPV result. We see a tendency that a preliminary diagnosis of for example normal or ASC-US is upgraded when a positive HPV mRNA result is acknowledged. As a result, the clinical sensitivity of cytology as such will be over-estimated. In order to get two groups for comparison, a study with two separate arms was conducted. One arm with post-colposcopy repeat cytology only and one arm with both repeat cytology and HPV test. National guidelines recommend that a woman with negative biopsy should be followed up with repeat cytology and an HPV test after 6–12 months. The HPV-test however is not always performed due to use of conventional Pap-smears instead of LBC, a method not allowing for HPV testing. This made it possible to get a separate arm with repeat cytology only. The HPV test result is objective and not dependent on the cytological diagnosis. Women referred to histology based on the cytological result alone  made it possible to calculate clinical properties also including HPV negative results.Until 2006, only conventional Pap-smears were used at our hospital. When HPV-testing was introduced, liquid based cytology was recommended, but still almost half of the samples for repeat cytology are conventional Pap-smears. A limitation of the study is the absence of formal randomization, which can result in imbalanced groups. Still, one argument about comparability is the similar age and similar prevalences of CIN2+. In the group with cytology only, follow-up and in the HPV group, the average age was 41.1 and 42.4 years, respectively (p = 0.235). The median age was 38 and 39 years, respectively. The prevalence of CIN2+ was similar in the two groups .Referral to biopsy in the HPV-category is based on both the cytology and the HPV result and therefore does not represent the true number of women referred to biopsy based on HPV testing alone. For example, women with HSIL and ASC-H were always referred to biopsy independent of the HPV result. Women with ASC-US and LSIL were followed with new cytology after 12 months if the HPV mRNA test was negative.With regard to follow-up cytology, statistical parameters were calculated both at cut-off ASC-US+ and ASC-H+. The reason for this is different follow-up in the two groups. Women with ASC-H+ are referred to colposcopy and biopsy. Women with ASC-US and LSIL are followed up with cytology and HPV-testing. Post-colposcopy cytology with cut-off ASC-US+ has a high sensitivity (84.6%) for CIN2+, but only a moderate specificity (76.4%). In contrast, repeat cytology with cut-off ASC-H+ has a low sensitivity (53.8%), but a high specificity (96.4%). The combination of HPV mRNA testing and cytology with cut-off ASC-H+ has both high sensitivity (93.5%) and high specificity (89.0%).The HPV mRNA data for women above and under 40 years of age were analyzed separately. This was done both because older age is associated with lower rates of regression of lesions, a higher degree of recurrence after treatment of precancer and the risk of obstetrical adverse effects is of less importance due to the lower rate of planned children For the 97 women under 40 years of age with HPV mRNA test, the estimates of sensitivity, specificity, PPV and NPV were 88.9%, 91.4%, 80.0% and 95.5%, respectively. For the 95 women above 40 years of age with HPV mRNA test, the estimates of sensitivity, specificity, PPV and NPV were 89.5%, 93.4%, 77.3% and 97.3%, respectively .In the present study, the HPV mRNA test shows a specificity of 92.5%. Previous data on HPV mRNA testing in the triage of ASC-US and LSIL Women with negative biopsy require follow-up before resumption of routine screening. In the present study, the HPV mRNA test is as sensitive and significantly more specific than repeat cytology at cut-off ASC-US+ in follow-up of women with negative biopsy. Thus, the study findings provide evidence that supports our hypothesis that HPV E6/E7 mRNA testing is more specific than follow-up cytology without loss in sensitivity in postdiagnostic management of women who have a negative biopsy. In addition, the HPV mRNA test has a significantly higher PPV than cytology alone. These data indicate that the HPV mRNA test is a better follow-up test than cytology for women with negative biopsy. When the follow-up test is HPV mRNA positive, our data suggest direct treatment for women above 40 years of age contributing to a better clinical safety for these patients. The use of HPV mRNA test can reduce the time from a false negative biopsy to diagnosis and treatment for CIN2+. Further studies are needed in order to reveal whether post-colposcopy HPV mRNA testing and E6/E7 detection may replace cytology in follow-up of women with negative biopsy.


The Human Papillomavirus (HPV) is a sexually transmitted virus that causes cervical cancer. Since 2008 a vaccination program targeting 12-year-old girls has been initiated in Italy, backing up the cervical screening program already active since 1996. We propose a mathematical model of HPV transmission dynamics with the aim of evaluating the impact of these prevention strategies. The model considers heterosexual transmission of HPV types 16 and 18, structured by sex, age and sexual activity level, where transition to sexual activity is explicitly modeled from recent survey data. The epidemiological structure is a hybrid SIS/SIR, where a fraction of individuals recovering from infection develops permanent immunity against reinfection. Infections may progress to cervical lesions and cancer and heal spontaneously or upon treatment. Women undergoing hysterectomy (either after treatment of HPV lesions or by other causes) also transmit HPV infection. The model fits well both the age-specific prevalence of HPV infections and the incidence of cervical cancers in Italy, and accurately reproduces the decreasing trend in cancer incidence due to the introduction of the screening program. The model predicts that if the screening coverage is maintained at current levels, even in the absence of vaccination, such trend will continue in the next few decades, eventually plateauing at 25% below the current level. The additional initiation of routine vaccination targeting 12-year-old girls will further reduce cervical cancer incidence by two thirds at equilibrium, under realistic assumptions of 70% coverage and a duration of protective immunity of 50 years. If catch-up immunization of 25-year-old women at first cervical screening is also introduced, about 3,000 cervical cancer cases overall can be averted, corresponding to 9.6% of all cases expected in the scenario without catch-up. We conclude that HPV vaccination in addition to cervical screening will significantly reduce the burden of cervical cancer in Italy. Many types of human papillomavirus (HPV) are known. Of these, about 50 have a high tropism for the ano-genital mucosa and are sexually transmitted. According to the most recent classification of the International Agency for Research on Cancer (IARC) 12 genotypes are defined as high-risk (HR) oncogenic, 13 are classified as “probable” and “possible” oncogenic risk and the others are classified as “non-carcinogenic” Two different vaccines are available for the prevention of HPV-related cancers, both targeting HPV 16 and 18. The vaccines have shown an efficacy close to 100% in preventing precancerous lesions in HPV-naïve individuals The widespread diffusion of HPV infection puts all female sexually active population at risk of cancer. For this reason in the past decades, many countries worldwide have initiated cervical screening programs, aimed at the timely detection and treatment of precancerous lesions In this work, a mathematical model is used to evaluate the effect of adding an immunization program for females aged 12 years against HPV 16 and 18 only (i.e. without considering the possible effects of cross-immunization versus other HR HPV genotypes) to the existing screening protocol. Vaccination schedules combining pre-adolescents immunization with various catch-up options at different ages are also evaluated with respect to the predicted incidence of pre-cancerous lesions and cervical cancers and to the number of needed treatments.Compared to previous modeling studies on HPV in Italy Screening coverages for 1996–2008 were provided by the National Observatory on Screening A mathematical model described by ordinary differential equations was built to reproduce the heterosexual transmission dynamics of HPV infections caused by types 16 and 18, and progression from infection through various stages of disease. The population is assumed to be at demographic equilibrium with a fixed inflow of births per year and a realistic but time-invariant life table, and is structured by sex, chronological age, and sexual activity level, as standard for HPV models A flow-diagram of the model for a specific age class and sexual activity level is reported in z of developing permanent immunity. Lesions of grade CIN-2 or higher have a severity-specific probability of being diagnosed  and treated. The rates of diagnosis and treatment for cervical screening were calculated in such a way to mimic the Italian screening protocol Susceptible individuals of both sexes acquire HPV infection at a time-varying rate  specific for sex, age and sexual activity level A severity-specific proportion of treated lesions Full technical details on the model are given in HPV vaccination is included through a flexible schedule considering both routine immunization for females aged 12 years and catch-up immunization. Vaccination is assumed to be effective only on individuals not currently infected with HPV, and its protective effect can wane in time. We first consider three hypothetical reference scenarios: i) “no-intervention”, where neither screening nor vaccination are considered; ii) “screening only”, where the screening program implemented during 1996–2008 is continued thereafter at the 2008 level (coverage at about 60%); iii) interrupted “actual vaccination”, considering, in addition to the continuation of current screening program, vaccination of females aged 12 years only during 2008–2012, at vaccine uptake observed in Italian regions since 2008 As most available models of HPV transmission dynamics Model predictions of age-specific prevalence of infection  and incidence of cervical cancer were calculated by running the model until equilibrium and then introducing the screening program  with a time-varying coverage following official estimates R:Parameter estimates were computed by exploring the whole parameter space by Latin Hypercube Sampling (LHS) 1 is the number of data points for HPV prevalence and D▪1 and P▪1 are the vector of observations and model predictions, respectively. Similarly, N2 is the number of data points for cervical cancer incidence and D▪2 and P▪2 are the vector of observations and model predictions. This error allows goodness of fit evaluations for heterogeneous data having different order of magnitudes, as is the case for HPV infection prevalence and cancer incidence R Click here for additional data file.


The primary aim was to assess the distribution of HPV types using this new assay in atypical squamous cells of undeterminate significance (ASCUS). The secondary aim was to correlate the HPV types with the severity of the disease. The study population comprised 376 ASCUS women. The women were all Hybrid Capture II (HCII) positive and were admitted in three European referral gynecology clinics between 2007 and 2010. A colposcopy with histological examination was performed in all these patients. HPV 16 was typed in 40 % of patients, HPV 18 in 7 %, and HPV 31 in 17 %, and 18 % of patients had mixed genotypes. Patients aged over 30 more often had the HPV 16 genotype than patients aged under 30 . The risk of cervical intra-epithelial neoplasia of grade 2 or more (CIN2 +) when HPV 18 positive is lower than the probability associated with HPV 16 or HPV 31: 28 % vs. 58 % and 52 %, respectively . The Digene LQ®, a new sequence-specific hybrid capture sample preparation, is fast and efficient and allows high-throughput genotyping of 18 HR HPV types by PCR compared to traditional non-sequence-specific sample preparation methods.A new genotyping-based DNA assay (Digene LQ High-risk types of human papillomavirus (HPV) are causative agents for cervical cancer. The effectiveness of the cervical screening program could be improved by testing for the DNA of high-risk types of HPV as a primary screening tool –3. More In view of the increasing importance of HPV genotyping, it is important to develop robust, high-throughput assays. Recently, HPV genotyping tests have been presented as relevant for management of screened women, in order to identify which HPV-positive women have persistent oncogenic HPV infection , 11. Stu®) for the identification of 18 HR HPV types on GP5+/6+-PCR products was developed and compared analytically to the established Reverse Line Blot (RLB) genotyping assay [A novel, commercial system (Digene LQng assay . Godinezng assay  recentlyThe primary aim of this study was to assess the distribution of HPV types using this new assay. The secondary aim was to correlate the HPV types to the severity of the disease (cytology and histology) with calculation of probabilities of CIN2+ by HPV type.The study population comprised 376 atypical squamous cells of undetermined significance (ASCUS) women who were admitted to three European referral gynecology clinics between 2007 and 2010: 158 patients from Madrid (Spain), 123 from Marseille (France), and 95 from Milan . Criteria for eligibility were age between 18 and 60 years, abnormal cervical smears and HC-II-positive results, and referral for colposcopy with histology. Before colposcopy, a cervical sample was obtained using the ThinPrep method . The cervical scrapes were collected with the PreservCyt transport medium . All of these tests were performed on the samples collected in PreservCyt liquid medium for liquid-based cytology (ThinPrep). Informed consent was obtained from each participant according to the ethics committee guidelines. This study was approved by the CPP Sud-Med I (Comité de Protection des Personnes Sud Méditerranée I) under reference number 07 22.The following assays were carried out and scored in strict accordance with the manufacturer’s instructions.(HCII) (Digene): this assay detects 13 HR HPV genotypes . The HCII test was performed using the automated HCII assay system as described previously (13). For each specimen, results were expressed in relative light unit/cutoff (RLU/CO), corresponding to the ratio of the specimen luminescence relative to the luminescence of the 1.0 pg/ml HPV16 standard provided with the kit. Samples with an RLU/CO value ≥1 were considered HCII-HR positive. Samples with an RLU/CO value <1.0 were considered HCII-HR negative.The Digene LQ test utilizes probes for 18 HR HPV types  that are the same as the respective RLB probes, with minor modifications, and are immobilized on color-coded beads. The Digene LQ test detection was performed in the Luminex 100 IS System (Luminex Corporation). In brief, 3B buffer was added to the HR HPV beads to minimize the background in the final Luminex read-out. Subsequently, GP5+/6+-PCR products were added. Next, heat-denaturation, hybridization under stringent conditions, and incubation with streptavidin-conjugated R-phycoerythrin detection conjugate were followed by read-out according to the specified instrument settings, resulting in MFI levels per HPV type for each specimen.Two-sided P-values were calculated by Chi-square or Fisher exact tests and placed on 2 × 2 contingency tables; Cochran-Armitage test for trend was used for testing trend binomial age proportions across levels of cervical intra-epithelial neoplasia. All P-values < 0.05 were considered statistically significant. Calculations were performed using SAS software .Table Figure Figure Figure Figure Table ® assay . The distribution of genotypes using this assay indicated, as expected, that HPV types 16, 31/33, 18 and mixed genotypes are the most prevalent, in accordance with previously reported results [This multicenter study evaluated for the first time the clinical utility of HPV genotyping using the recently developed Digene LQ results .The recent ASCCP guidelines recommend the use of HPV genotyping for patient management, with a direct referral to colposcopy based on HPV16- and/or HPV18-positive results . MoreoveThis new sequence-specific Digene LQ test is fast and efficient and allows direct HPV genotyping of 18 HR types by PCR, compared to traditional non-sequence-specific test methods. The utility of this method was evaluated on cervical samples positive for HR HPV by the HCII screening assay from patients referred for colposcopy.®, the probability of being CIN2+ when having HPV16 is 58 %, similar to the probability associated with HPV31, 52 %, and the probability is 28 % if HPV18 positive. The probability of being CIN3+ when having HPV16 is 23 % . The relative probability of CIN2+ when being HPV16+/HPV18- is 58 % (the probability of CIN3+ is 24 %). Similarly, the probability of CIN2+ when being HPV16+/HPV31 is 59 % (the probability of CIN3+ is 25 %), and the probability of CIN2+ when being HPV31+/HPV18- is 54 % (the probability of CIN3+ is 13 %).Among women with ASCUS cytology, the relative probability of pre-cancer is higher in HR HPV-positive women than HR HPV-negative women . InteresThese data are similar to those reported in the recently published ATHENA study: HPV16/HPV18+ women had a greater probability of CIN 2 or worse compared with pooled HR-HPV+ and HR-HPV- women  .Among HR-HPV-positive women, risk of disease likely increases over time, as has been reported for genotypes 16 and 18 by Khan et al. . Up to nWhile all women with ASCUS cytology testing positive for HR HPV should be referred for colposcopy, the results from this study demonstrate that those women with ASCUS cytology testing positive with HPV16 and/or 18 and 31 are at a particularly high risk for ≥ CIN3 disease, reinforcing the need for immediate colposcopy and more intense follow-up, particularly in the case of a negative colposcopy. These findings corroborate those of other studies , 28.This study points out the greater probability of CIN2 and CIN3 for patients testing positive for HPV31. HPV31 is a more prevalent type than HPV18 in the ASCUS population in European countries , 29. NotAnother prevalent genotype in our population was HPV56, found in the same proportion as HPV18.Regarding the laboratory technical characteristics, this assay uses multiplex, bead-based xMAP technology and an automated, high-throughput read-out by either the LiquiChip 200 workstation  or Luminex 100 IS System . The test was developed for identification of the 18 HR HPV genotypes associated with cervical cancer using GP5+/6+-PCR products. The Digene LQ Test and the RLB assay have been reported to have a high level of agreement in detection and genotyping of 18 HR HPV types in HCII-positive specimens .®, a new sequence-specific HC sample preparation is fast and efficient and allows high-throughput genotyping of 18 HR HPV types by PCR compared to traditional non-sequence-specific sample preparation methods. This study showed that HPV types 16, 31, 33, 18 and mixed genotypes are the most prevalent genotypes and that ASCUS women who are HPV16 positive have the highest probability of CIN2+ and CIN3+.The Digene LQ


A cross sectional study to investigate HPV prevalence according to age and cytology.Women presenting to a gynaecological outpatient clinic for a Pap smear test were included in the study (n=3177). All women had cervical cytology and HPV testing.Overall prevalence of any 24 HPV type analysed was 33.1% (95% CI 31.5% to 34.7%) and HPV 16 and HPV 42 were the most frequent (6.7% (95% CI 5.8% to 7.6%), 6.8% (95% CI 5.9% to 7.6%)), in total samples. Multiple HPV infection rate was 12.9% (95% CI 11.8% to 14.1%). High risk HPV (hrHPV) types were present in 27.4% (95% CI 25.8% to 28.9%) of the samples.HPV prevalence was highest among 14 to 19 y.o (46.6% (95% CI 40.7%-52.4%)) and second highest among 30–34 y.o. . HPV 16 was highest among 20–24 (9.0% (95% CI 6.4%–11.6%)) and second highest among 50 to 54 y.o. (6.3% (95% CI 2.9% to 9.8%).In Low-grade Squamous Intraepithelial Lesions (LgSIL) cytology samples, the most frequently detected hrHPV types were: 16 (14.5% (95% CI 12.1% to 16.9%)), 51 (13.0% (95% CI 10.7% to 15.3%)) and 53 (9.1% (95% CI 7.2% to 11.1%)) and in High-grade Squamous Intraepithelial Lesions (HgSIL) were: HPV 16 (37.2% (95% CI 26.5% to 47.9%)), HPV 51 (17.9% (95% CI 9.4% to 26.5%)) and HPV 18 (12.8% (95% CI 5.4% to 20.2%)).In the population studied, HPV 16 and 51 were the most frequent detected hrHPV types. HPV positivity, hrHPV and multiple HPV types infections were higher in young women, while HPV prevalence declined with increasing age and presented two peaks a higher (14–19 y.o.) and a lower one (30–34 y.o.) These results may contribute to the creation of a national screening programme. Overall and age specific distribution of high risk HPV (hrHPV) and low risk HPV (lrHPV) types data across geographical regions is crucial for the optimization of prevention strategies in each country -4. Over HPV 16 and HPV 18 are the most common oncogenic types associated with cancer, and are targeted by recently developed vaccines [In order to improve the current cervical screening process, it is essential to combine cytology and HPV testing. Large scale randomized controlled studies need to be performed to establish the most efficient and cost effective cervical cancer screening programme. In Greece there are limited epidemiological data on HPV prevalence -12.The aim of this study is to describe the prevalence and age distribution of different HPV types among women presenting for a Pap smear in an outpatient clinic in Greece.st Department of Obstetrics and Gynaecology, University of Athens, ‘Alexandra’ Hospital (participation rate 33%), between January 2007 and November 2010 in order to have their annual test Pap.The study group consisted of a consecutive sample of 3177 Caucasian women between 14 and 70 years old who proceeded to the Outpatient Gynaecological Clinic of Regional Anticancer Oncology Hospital of Athens “St. Savvas” (participation rate 67%) and 1The patients were also offered HPV test with the knowledge that it is not part of the screening. Women were eligible if they were no pregnant and had no history of HPV cervical disease. All patients included in the study have been only checked once throughout these years. Adequate specimens had been obtained from all of them, except 7, which were excluded. All patients included in the study gave their written informed consent after discussing with gyneacologists that there would be no implications to their health. Sexually active adolescents attended either an adolescent health clinic or an adolescent gynecology clinic in order to have gyneacological tests, such as conventional genital cultures, and were offered cervical screening and HPV typing as part of a sexual health prevention protocol. All adolescents and their guardians gave their written informed consent prior to being examined after discussing with gynaecologists that it would have no impact on their well being. Ethical approval was granted by the ethics committee of Regional Anticancer Oncology Hospital of Athens “St. Savvas”.The studied population was divided into three age groups according to the biological alterations in women’s reproductive system , in order to facilitate the recognition of specific differences in the biological and clinical behaviour of HPV infection across the years. Cervical cell scrapings were collected by a gynaecologist with a cytobrush  from the ecto- and endo-cervix of the uterus. A slide was prepared for conventional cytology and the cytobrush was then placed in specimen transport medium  and stored at 4°C until prepared for HPV molecular analysis. Cytological findings were classified in line with the 2001 Bethesda classification system. The smears were submitted to two cytopathologists who worked separately and the results were discussed. In rare cases of discrepancy the smear was submitted to the Head of the Cytopathology department. The Cytopathology department has established an internal and also external quality control based on international external quality assurance tests  5 ml of Thin Prep samples were used for DNA isolation. They were centrifuged, diluted into lysis buffer  and finally subjected to the NucliSENS® easyMAG®platform (bioMérieux Hellas S.A) for automated extraction, according to manufacturer’s instructions. Nucleic acids were eluted in 55 μl of elution buffer. DNA quality test was carried out using Human Globin, Beta, Primer set kit  according to manufacturer’s instructions. Aliquots were stored appropriately for further processing.The PapilloCheck® HPV-Screening  was used. This technology is based on a DNA chip for the type-specific identification of 24 HPV types  [Data were analysed using SAS v9.0. Absolute and relative frequencies were used to present the HPV types distribution according to age and cytology. The HPV prevalence and 95% confidence intervals were calculated using binomial methods and stratified by cytology report and HPV category. Chi-squared tests were performed to assess statistical significance of any differences in prevalence. 2 × 2 contingency tables Fisher’s exact test was performed along with Odds Ratio and 95% confidence intervals calculation. Pearson’s Chi Square test was used in cases of contingency tables bigger than 2 × 2. Cochran-Armitage test for trend was used to investigate trend in distribution of certain types and multiple HPV infection according to escalation of age. 2-independent samples t-test was used, along with relevant descriptive statistics  to compare average age among patients depending on presence of specific types and multiple infections. 5% level of statistical significance and 95% confidence intervals were used.The response rate was 62% 3177/5124) and the final sample consisted of 3170 women with complete cytological and HPV-DNA testing results. The majority of them were Greek  and a 5.6% (177/3170) were Caucasian immigrants living in Greece. Women were 14–70 years old (mean age 34.2) and were divided according to their age in three groups: 14-25 , 26–46 y.o.  and 47-70 y.o . Their cytology results and distribution by age group are shown in Table 77/5124 aOverall, 1049 of the 3170 (33.1% (95% CI 31.5% to 34.7%)) samples were found to harbour an HPV infection. We examined HPV prevalence in 5-year periods in order to assess age trends in relation to HPV infection in more detail. HPV prevalence exhibited a peak of 46.6% (95% CI 40.7%–52.4%) at 14–19 y.o. and a second peak at the age group 30-34 y.o.  which decreased thereafter , 34% , 74.4%  and 92.3% , respectively . Overall, HPV 16 was the most common high risk HPV type present with a prevalence of 6.7% (95% CI 5.8% to 7.6%), followed by HPV 51 , HPV 53 , HPV 56  and HPV 31  , 51  and 53 . Among ASCUS cytology samples, only hrHPV types were detected; HPV 16 was once more the most frequent type  followed by HPV 51  and HPV 45 . The three most frequently detected hr HPV types were 16 , 51  and 53  in LSIL. In HSIL the most common hrHPV types were 16 , 51  and 18  , followed by HPV 44  and HPV 6 . These three lrHPV types were detected in this order in women with normal cytology (3.6% (95% CI 2.8% to 4.4%), 0.9% (95% CI 5.0% to 13.0%) and 0.6% (95% CI 3.0% to 10.0%) respectively). HPV 42 was the most frequent lrHPV type detected in LSIL  followed by HPV 6  and HPV 44   and HPV 16 prevalence remained relatively high until the age of 35-39 y.o. and decreased sharply thereafter. A second peak of 6.3% (95% CI 2.9%–9.8%) was present at women aged 50–54 y.o and the prevalence dropped in older women  of all samples and 39.1%  of the HPV infected samples ) than in ASCUS , Fisher’s exact test p value p<0.001) or normal , Fisher’s exact test p value p<0.001).Among women with multiple infections, 66.8% (95% CI 62.3% to 71.4%) were infected with two HPV types; the most common combination was of 16 with 51, 56 with 66 and 6 with 42. There were 22.6% (95% CI 18.6% to 26.7%) of infected women with three HPV types, the most common combinations of which were 11 with 33 and 45 and the combination of 16 with 42 and 45. There were also cases with 4 HPV types , 5 HPV types  and 6 HPV types .We present a large study of HPV prevalence in women of different ages in the area of Athens, from which interesting conclusions can be drawn. As HPV DNA typing is gradually incorporated in national screening programmes knowledge of HPV type variations in different geographical regions is useful information. Similarly, as HPV vaccination is introduced as a means of primary cervical cancer prevention, better prediction models of its efficacy specific for each area can be made .In our study and in concordance with what is seen in the general population, normal cytology dominated all age groups. The highest LgSIL rates appeared among women aged 14–25 years. Only a minority (1.5%) of young girls had a high grade lesion and this is again consistent with what is seen in the literature. It also justifies the current American and British screening programme that suggest smear testing after the age of 21 and 25 respectively .In our study, the HPV prevalence in normal samples was 15.7%. For Eastern Europe, HPV prevalence in a meta-analysis of 4053 samples tested with normal cytology was 21.4% ,12 whichIn an attempt to compare our results with other countries’ epidemiological data, we presented an age-related prevalence curve of HPV infection. This curve showed a peak at the 14–19 age group, a second one at 30–34 y.o. and then dropped. Although the study by Coupe et al conducted in the Netherlands demonstrated an association between age and HPV prevalence , others As expected, HPV prevalence, increased as lesions progressed to higher grade ones. The same trend was observed in hrHPV positivity in relation to cytological status. Investigating the results per age group, we observed that hrHPV infection dominated in 14-25 year old women irrespective of cytology. However young women are less likely to present cancerous lesions as in the majority of cases the lesion regresses after 2–3 years ,22.Previously, in a 1636 women cohort study conducted by this group, where the prevalence of subtypes 6, 11, 16, 18, 31 and 33 was tested, HPV 11 was found to be the most frequent . In the HrHPV distribution in Mediterranean countries like Italy, Portugal, Spain -26 is siMultiple infections were found in 12.9% of Greek population. Multiple HPV positivity was more common in younger women. In addition, women with cervical lesions had a higher rate of multiple infections compared to those who had normal cytology. Multiple infections might be a risk factor for development of cytological abnormalities. The majority of multi-HPV infected women harboured at least one hrHPV type and the majority of those carried two types, which is consistent with other epidemiological studies [Although this study presented here included a large number of women of a broad age range, it has some limitations. Our sample is representative of women presenting for smear testing. However, as smear taking in Greece is done on an ad hoc basis rather through an organised screening system, our sample cannot be considered representative of the Athenian population. These data may give us important information regarding regional HPV prevalence but large epidemiological studies from different regions of our country are needed.HPV 16 and 51 were the most commonly identified hrHPV types, present in 6.7% and 5.7% overall and 37.2% and 17.9% in HgSIL cytology samples, respectively. Multi infection was present in 39.1% of infected cytology samples. Our results were comparable with what has been found in other Mediterranean countries.HPV positivity, hrHPV and multiple HPV types infections were higher in young women. HPV prevalence declined with increasing age and presented two peaks a higher (14–19 y.o.) and a lower one (30–34 y.o.). HPV 16 prevalence remained high until 35-39 y.o. and presented a second peak in women aged 50–54 y.o. These results may contribute to the creation of a national screening programme.The authors declare that there are no competing interests.EA carried out the molecular tests, participated in the statistical analysis and drafted the manuscript. SP and ET carried out the molecular tests, participated in the statistical analysis and helped to draft the manuscript. LM and EM performed gynaecologic examinations and helped to draft the manuscript. IP participated in the statistical analysis and helped to draft the manuscript. DD performed the cytologic analysis. IT and GM performed gynaecologic examinations. EP designed and coordinated the study 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/1471-2334/13/53/prepubHPV type distribution according to the cytology subgroups.Click here for fileHPV type distribution according to the age groups.Click here for file


Circadian oscillation provides selection advantages through synchronization to the daylight cycle. However, a reliable clock must be designed through two conflicting properties: entrainability to synchronize internal time with periodic stimuli such as sunlight, and regularity to oscillate with a precise period. These two aspects do not easily coexist, because better entrainability favours higher sensitivity which may sacrifice regularity. To investigate conditions for satisfying the two properties, we analytically calculated the optimal phase–response curve with a variational method. Our results indicate an existence of a dead zone, i.e. a time period during which input stimuli neither advance nor delay the clock. A dead zone appears only when input stimuli obey the time course of actual solar radiation, but a simple sine curve cannot yield a dead zone. Our calculation demonstrates that every circadian clock with a dead zone is optimally adapted to the daylight cycle. Circadian clocks are acquired through evolution independently in bacteria, fungi, plants and animals  obeys a simple one-dimensional diffusion equation, its solution is represented byWith sufficiently weak noise, Kuramoto  for furtT isEquation (2.11) shows that the variance of the phase after one period ϕ starting from 0 to 2π, and the period variance is the variance of the first passage time. Because direct calculation of the period variance is difficult, we approximate the period variance et al.  = 0 yields the optimal iPRCψM and ψm themselves depend on iU(ϕ), they have to satisfy a self-consistent condition, i.e. equation (2.18) is maximal with ψM and ψm. Consequently, we maximize the following function:Δ = ψM – ψm and δ = ψm. The optimal iPRC can be obtained by first finding the maximum solution of Ψ with respect to Δ and δ, and then substituting the obtained solution ψm = δ and ψM = δ + Δ into equations (2.20) and (2.21).The variational equation to be optimized isa et al. , which o2.5.0I = I cos ϑ and I = 0 when the sun is above the horizon (0 ≤ ϑ < π) and below the horizon (π ≤ ϑ < 2π), respectively, where ϑ is the zenith angle and I0 is the maximum irradiance [x) is the ramp function defined by ramp(x) = x for x ≥ 0 and ramp(x) = 0 for x < 0. We call equation (2.24) the solar radiation input, whose plot is shown in figure 4a (the shaded region represents night). In order to show the validity of the solar radiation modelling, we compare equation (2.24) with observed irradiance data from Vick & Moss [figure 4b. In figure 4b, equation (2.24) is plotted by the solid line (left axis) and the observed data by the dashed line (right axis). The solar radiation input of equation (2.24) is shifted horizontally, so that equation (2.24) becomes a good fit to the data. From figure 4b, the solar radiation input is in good agreement with the observed data, which verifies the validity of equation (2.24) as a solar radiation model.Optimal PRCs depend on input signals, as seen in equations (2.20) and (2.21). The most common synchronizer in circadian oscillators is sunlight, for which the strength is determined by 24 h-periodic solar irradiance. The solar irradiance is calculated by radiance . It can k & Moss , which aFor comparison, we also use a sinusoidal input, which is common in nonlinear sciences:p(ωt) = B + sin(ωt), where B is an arbitrary constant, also yields the same optimal PRCs as equation (2.25), because a constant B in the signal is offset in equations (2.20)–(2.23). Although a constant B does not play any roles in formation of the optimal PRCs, different B result in different Arnold tongues in general. For calculating the optimal PRCs, we use equations (2.24) and (2.25).Note that 3.3.1.jth molecular species includes a parameter ρ asρ, and kx is the concentration of the kth molecular species. Here, k∈{1,2, … ,N} can take any value, regardless of j . For example, let figure 4c be a gene regulatory circuit of a hypothetical circadian clock, where symbols → and ⊣ represent positive and negative regulations and ix are molecular species  with j = 2, k = 1 and ρ being the translation rate. In equation (3.1), although we can also consider an alternative case Light stimuli generally affect the oscillatory dynamics multiplicatively, i.e. they act on the rate constants or transcriptional efficiency of the gene regulatory circuits ,40. We a & Tyson  for typixLC, and hence the points on the limit cycle can be uniquely determined by the phase ϕ. Consequently, under the phase reduction, kx is replaced by xkLC,(ϕ) in equation (3.1), where xkLC,(ϕ) is the kth coordinate of xLC  constitutes a core clock component and is generally a smooth 2π-periodic function, we approximate it with a sinusoidal function:u is the initial phase and α denotes the amplitude of the oscillation (d). To ensure α ≤ 1, and α = 0 recovers the additive case. Because the initial phase u does not play any role (u is offset by δ in equation (2.23)) when the white Gaussian noise is additive (i.e. iQ(x) ∝ 1), we also set u = 0. The parametric approximation of equation (3.2) enables an almost closed form for the overall calculations. Although we assumed in equation (3.1) that effects of ρ only depend on kx, we can generalize equation (3.1) to K(x) is a nonlinear function and is assumed to be well approximated by 1 – αsin(ϕ + u). By this generalization, our theory can be applied to other possible light entrainment mechanisms such as the intercellular coupling [iU(ϕ) of molecular species not having light input entry points. Consequently, for a noise term iQ(x), we assume that the white Gaussian noise is additive and is present only in the jth coordinate equation (q is the noise intensity and iQ(ϕ) = 0 for i ≠ j).When using phase reduction, the dynamics of the limit cycle are considered on the unperturbed limit-cycle trajectories illation d. To enscoupling . Our moda–c shows the landscape of Ψ as functions of Δ and δ, and figure 5d–f expresses the optimal iPRCs jU(ϕ) and pPRCs Z(ϕ) for the solar radiation input  is given in appendix A). The optimal PRC shape does not depend on the model parameters such as the period T, its variance q. These three parameters only act on the magnitude of the PRCs . Consequently, we normalized T = 1, q = 1, as shown in α, Ψ is plotted for three cases: α = 0, (figure 5a), α = 0.5 (figure 5b) and α = 1.0 (figure 5c), where the maximal points  yield the optimal PRCs using equations (2.20) and (2.21). The maximal parameters Δ and δ are calculated numerically. Figure 5d–f describes the optimal iPRCs (solid line) and pPRCs (dashed line) for α = 0, 0.5 and 1.0, respectively. When α = 0, i.e. the input signal is additive, Ψ achieves a maximum for Δ = π and arbitrary δ, yielding sinusoidal PRCs as the optimal solution (figure 5d). Although the input signal p(ϕ) is not sinusoidal, the optimal PRCs obtained using the variational method become sinusoidal. In other words, considering optimality, resonator-type oscillators have an advantage over integrator-type oscillators. For α > 0, the input signal p(ϕ) depends on the concentration of the kth molecular species. From figure 5b, the optimal parameters for α = 0.5 are  =  and , which are different from Δ = π . Figure 5e shows the optimal iPRCs jU(ϕ) and pPRCs Z(ϕ) for α = 0.5. Interestingly, the optimal iPRCs and pPRCs for α = 0.5 have a dead zone (region of e) in which the input signal neither advances nor delays the clock. From equations (2.20)–(2.21) and the solar radiation input of equation (2.24), the optimal PRCs inevitably include a dead zone if the optimal Δ is not π. For α = 1.0, there are four sets of parameters  that give optimal PRCs: , ,  and  . Consequently, the optimal PRCs shown in figure 5f have a dead zone as in the case of α = 0.5.Figure 53.2.α > 0. We next studied the length of the dead zone as a function of α (a) and improvements in the entrainability induced by the dead zone (b) for the solar radiation input. Because the dead zone, which is a null interval in PRCs, emerges when the optimal parameter is Δ ≠ π, we can naturally define its length asΔ is the maximum value of Ψ. As seen in figure 6a, a dead zone clearly exists when α > 0, and the length increases with increasing α for α < 0.8. Even for α = 0.1, when the oscillation amplitude of xkLC,(ϕ)  is very small, we observe a dead zone with a length of L = 0.475, which corresponds to about 3 h within 24 h, indicating the universality of having a dead zone in order to attain optimality. The improvement in the entrainability that is induced by a dead zone is calculated by comparing the entrainability of the optimal PRCs with that of typical sinusoidal PRCs. We consider sinusoidal functions for both the iPRC jU(ϕ) and pPRC Z(ϕ) by settingc is the parameter to be optimized so that entrainability is maximized for each α (see appendix B for the explicit expressions). Equations (3.4) and (3.5) are scaled, so that they satisfy the constraints on the period variance (equation (2.12)). We calculated the ratiosUR and ZR quantify the improvement rate of the optimal PRCs over the sinusoidal iPRC (UR) and pPRC (ZR). In figure 6b, the solid and dashed lines show UR and ZR, respectively, as a function of α. Both ratios monotonically increase as α increases, which shows that the optimal PRC with a dead zone exhibits better entrainability when the oscillation of xkLC,(ϕ) has a larger amplitude. When the concentration of xkLC,(ϕ) is low, the effects of the input signal on the circadian oscillators are smaller. This is because pPRC Z(ϕ), which quantifies the extent of the phase shift owing to the stimulation of the parameter, depends on the concentration xkLC,(ϕ) (see equation (2.14)). However, even within the range ϕ where xkLC,(ϕ) has smaller values, the iPRC jU(ϕ) contributes to an increase in the variance of the period, regardless of the concentration. From this, we see that having an iPRC with a smaller magnitude when the concentration of xkLC,(ϕ) is smaller results in a smaller variance, which results in a larger entrainability for a constant variance of the period. Although this qualitatively explains the benefit of a dead zone, for some input values, the optimal PRCs may not contain a dead zone for any value of α. This will be shown in the following.From the results discussed above, the optimal PRCs have a dead zone when ion of α a and impead zone b for the3.3.Ψ is calculated in a closed form  is given in appendix A), which is plotted as functions of Δ and δ in figure 7a–c for three cases: α = 0 (figure 7a), α = 0.5 (figure 7b) and α = 1.0 (figure 7c). As can been seen from figure 7a–c, Ψ yields the maximal value for  =  for 0 < α ≤ 1, where n is an integer and when α = 0, δ can take any value. Figure 7d–f expresses the optimal iPRCs jU(ϕ) and pPRCs Z(ϕ) for the sinusoidal input. For α = 0, the optimal PRC is sinusoidal (figure 7d) and for α = 0.5, the optimal PRC is still close to a sinusoidal function (figure 7e). When increasing α to α = 1.0, the PRC diverges from the sinusoidal function and exhibits almost positive values (figure 7f). We see that the optimal PRCs owing to equations (2.20) and (2.21) do not exhibit a dead zone for any α-values (figure 7d–f) when the input signal is a simple sinusoidal function.Because the optimal PRCs depend on input signals (equations (2.20) and (2.21)), we next consider a typical periodic input signal, a sinusoidal function (equation (2.25)). In this case, 4.α = 0 (additive stimulation). Along with the fact that T, σT and q affect only the scaling of the optimal PRCs, when the input signal affects the dynamics multiplicatively (i.e. α > 0), the existence of a dead zone always provides a synchronization advantage. This is supported by many experimental studies of various species that report the existence of a dead zone in the PRC [c,d). Our general result suggests that circadian oscillators have fully adapted to solar radiation to improve synchronization. Indeed, many experimental findings imply that circadian oscillators have adapted to actual solar radiation [α = 0, optimal inputs and optimal PRCs do not have one-to-one correspondence. Thus, the optimal inputs are not trivial and this problem should be pursued in our future studies.The existence of a dead zone optimizes both entrainability and regularity. It is rather obvious that optimization of regularity alone leads to a dead zone , because the PRC   is low (0 ≤ ϕ ≤ π in figure 4d), and that to achieve optimality, the concentration of xkLC,(ϕ) should be maximal in the region where the PRCs exhibit a large phase shift. In Drosophila, the timeless (tim) gene is regarded as the molecular implementation of xkLC,(ϕ). It is experimentally known that light enhances the degradation of the gene product (the TIM protein) [Figure 2c shows observations of the PRC of Drosophila against light pulses as a function time from Hall & Rosbash [Our result also agrees with other experimental observations. Our theory implies that a dead zone should be located where the concentration protein) ,49, and  Rosbash ; circlesIn summary, we have constructed a model that regards circadian oscillators as a global optimization of entrainability and regularity. We have shown that our model is consistent with much experimental evidence as mentioned above. The extension and improvement of our method are possible and they are left as an area of future study.


In contrast to consistent epidemiologic evidence of the role of sexual transmission of human papillomavirus (HPV) in adults, various routes may be related to HPV infection in infants. We have assessed the extent of HPV infection during the perinatal period, and the relationship between mode of delivery and vertical transmission.A total of 291 pregnant women over 36 weeks of gestation were enrolled with informed consent. Exfoliative cells were collected from maternal cervix and neonatal buccal mucosa. HPV infection and genotypes were determined with an HPV DNA chip, which can recognise 24 types. The HPV-positive neonates were re-evaluated 6 months after birth to identify the presence of persistent infection. HPV DNA was detected in 18.9 % (55/291) of pregnant women and 3.4 % (10/291) of neonates. Maternal infection was associated with abnormal cytology (p = 0.007) and primiparity (p = 0.015). The infected neonates were all born to HPV-positive mothers. The rate of vertical transmission was estimated at 18.2 % (10/55) which was positively correlated with maternal multiple HPV infection (p = 0.003) and vaginal delivery (p = 0.050), but not with labour duration and premature rupture of membranes. The rate of concordance of genotype was 100 % in mother-neonate pairs with vertical transmission. The neonatal HPV DNAs found at birth were all cleared at 6 months after delivery.Vertical transmission of HPV DNA from HPV infected mother to the neonate increased when the infant was delivered through an infected cervix. However, the absence of persistent infection in infants at 6 months after delivery may suggest temporary inoculation rather than true vertical infection. Approximately 200 different genotypes of human papillomavirus (HPV) have been identified and >40 types are associated with anogenital diseases . While tIn contrast to the consistent epidemiologic reports, including the role of sexual transmission of HPV in adults , potentiThe extent and risk factors of HPV infections in infants have been controversial. For pregnant women and their babies, the only meta-analysis to date show that vertical transmission developed in one-third of neonates born to infected mothers mainly through vaginal deliveries . HoweverHPV DNA chip and PCR-based oligonucleotides microarray have been useful in many screening programs for the early diagnosis of cervical dysplasia and cancer . This teIn this study, we have determined the prevalence of HPV infection in pregnant women and the rate of vertical transmission during the perinatal period. The risk factors associated with viral transmission were also explored, including HPV genotypes.A total of 300 term pregnant women were enrolled over the study period. Analyses were restricted to 291 women who gave birth in the institutes through vaginal delivery  and caesarean section .The mean age of pregnant women was 32.8 years and the prevalence of HPV infection was 18.9 % (55/291) in maternal cervical swabs at enrolment determined by means of HPV DNA chips. In the cytology performed at the first trimester, 283 women were negative for intraepithelial lesions and 8 had atypical squamous cells of undetermined significance. Women with abnormal cytology were referred for colposcopic examination and the results indicated no dysplasia in the cervix. Prevalence of HPV infection in the newborns was 3.4 % (10/291) from swabs of oral mucosa at delivery.Details of the maternal characteristics regarding age, gestational history, bacterial genital infection, gestational diabetes mellitus, and abnormal cytology are given in Table Among the types specified by HPV DNA chip, HPV-16 was the most frequent type in the maternal cervix (n = 55), comprising 8 cases (14.5 %) and other specified HPV types were as follows: HPV-6, -56 each in 5 cases (9.1 %); -53, -58, -70 each in 4 cases (7.3 %); -39, -40 each in 3 cases (5.5 %); and −44, -51, -31 each in 2 cases (3.6 %). Each of HPV-11, -18, -33, -35, -45, and −66 was noted only in one participant. HPV-other type, which was positive in PCR but negative in HPV DNA chip, was noted in 29.1 % (16/55). Genotyping showed that 58.2 % (32/55) were infected with at least one type of high-risk HPVs.Co-infection with multiple genotypes was noted in 8 mothers; 3 mothers were infected with 3 types, and 5 with 2. HPV-53 (n = 4) was the most frequent type found in multiple infection. HPV-40 was detected in 3 cases and HPV-44 and −56 were detected in 2 cases. HPV-11, -16, -33, -35, -58, -66 and −68 were detected once.At birth, HPV DNA was detected in 10  neonates who were born to HPV-positive mothers. There was no infection among infants born to HPV-negative mothers. Therefore the overall frequency of HPV transmission from HPV-positive mothers to their infants was 18.2 % (10/55). Genotypes were concordant in all of the mother-neonate pairs  with HPV DNA chip, which can identify genotypes even in the cases with multiple infections through automated and efficient procedures. This technique is appropriate for large epidemiological studies and its efficacy for genotyping of HPV infection has been demonstrated in cervical cancer screening programs /55 with . Our resWe could add some data to the current consensus that HPV in a neonate was probably acquired during the passage through an infected cervix . The genHowever, many studies also presented biological evidence suggesting other possible routes of viral transmission. HPV DNAs have been detected in sperm, amniotic fluid, placenta, cord and maternal blood -26. FurtIn pregnant women, a few studies have assessed the extent of multiple infections, which is considered uncommon. With the introduction of laboratory techniques that can simultaneously identify multiple types of HPV, some investigators have reported that co-infection was detected in 17-66 % of the infected women ,22,23. SIt has been argued whether or not the detection of HPV DNA after birth reflects a true infection or contamination. Because HPV infection has a long latent period, the repeated detection of HPV in the same area is considered as persistent infection. Some early studies used PCR and serially detected HPV DNA in the buccal and genital swabs at birth and at weeks to months of postpartum. Cason et al.  demonstrAs commented in previous literature reviews, the estimates should be interpreted with caution in respect of geographic and demographic characteristics, research design, and detection methods of HPV ,5,12. BeIn conclusion, the infection and transmission of HPV was prevalent in pregnant women and their neonates. We suggest that the perinatal transmission of HPV takes place through an infected birth canal based on the concordance of HPV genotypes and the predominance of vaginal delivery. However, the clearance of HPV in all infants up to 6 months suggests that HPV DNAs of newborns do not indicate persistent infection in infants.This prospective study was approved by the institution’s ethics committee. Near term pregnant women anticipating deliveries at the department of obstetrics in a university affiliated hospital were recruited between April 2010 and January 2011. Participants fully understood the protocol of this study and signed written consent forms.Pregnant women over 36 weeks gestation were included. Exclusion criteria were as follows: severe intra-uterine growth restriction, severe oligohydroamnios or preterm premature rupture of membranes, severe hypertension during pregnancy, history of malignancies within 5 years, liver cirrhosis, renal failure, chronic diseases affecting maternofetal immunity and a history of psychological disease and alcoholism. On the day of admission for delivery, newborns of the participants were enrolled after maternal re-consent.At enrolment, cervical swabs were taken of the pregnant participants. Exfoliative cervical cells were collected using cytobrush for HPV DNA chip analysis. Immediately after delivery, the oral mucosa of newborns was scraped for HPV DNA chip analysis. The test was repeated at 6 month postpartum for HPV-positive infants to identify persistent infection.HPV detection and genotyping was performed using the HPV DNA Chip, PCR-based DNA microarray system, purchased from MyGene Company . The HPV DNA Chip contains 24 type specific probes; 15 types from the high-risk group  and 9 types from the low-risk groups . Briefly, DNA was isolated from swab samples using a DNA isolation kit , and target L1 regions of HPV DNA were amplified using consensus GPd5+/GP6day + primers. Ten μlof the HPV-amplified product was denatured for 5 min at 95°C. The samples were mixed with a hybridization solution then applied onto the DNA Chip.Hybridized HPV DNA was visualized using a DNA Chip scanner . HPV amplicons can be hybridized with the corresponding type-specific oligonucleotide probe and visualized on HPV DNA Chip slides as double-positive spots when HPV DNA is present in amplified PCR product.The samples that showed a positive band of 150 bp on the gel electrophoresis, but were negative on the HPV DNA Chip slide, were designated as HPV-other. None of the negative controls (without DNA) revealed HPV positivity.t-test and discrete with chi-square test or Fisher’s exact test. The Mantel-Haenszel test was used to control for the confounding from age to some factors which were significantly associated with maternal HPV status under univariate analyses. P < 0.05 was considered significant with the use of a 2-sided test.To identify the factors associated with HPV infection in pregnant women and neonates, participants were assigned into 2 groups according to the presence or absence of HPV. Continuous variables were compared with Student’s The authors declare that they have no competing interests.HP and SWL participated in the study design and the statistical analysis, and drafted the manuscript. IHL and HMR contributed to acquisition and interpretation of the data. ARC and YSK carried out laboratory procedures and helped to draft the manuscript. SRH carried out cytological classification of specimens and participated in HPV genotyping. SSK and SJS contributed to the conception of the study and participated in the statistical analysis. SMS participated in the conception and design of the study and acquired the data of infants. TJK conceived of and designed the study, and co-ordinated the acquisition of the data and draft of the manuscript. All authors read and approved the final manuscript.