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The diagnosis of HPV infection is generally carried out using immunological and molecular techniques based on high risk to probable high-risk HPV strains. The aim of this work is to generate a global representation of HPV strains for diagnosis and drug development. In this work, all the complete genomic DNA sequences of registered Human Papillomavirus (HPV) strains available in NCBI GenBank were used to obtain a consensus sequence of HPV using the Genetic Algorithm. The consensus DNA sequence was translated using the ExPASy software tool. In all, six longest amino acids frames were selected from the six translated frames. The amino acid sequence identity was carried out using the BLAST tool. The six amino acid sequences were identified as E1, E2, E6, E7, L1 and L2. The homology modeling method (Modeller Software Tool) was used to determine the secondary structure of these six identified primary amino acid sequence. The percentage of similarity ranged from 24% in L2 to 100% in E7 and L1. The functions of these structural domains were also determined from PDB databank, InterProScan and CATH. Hence the consensus sequence built using a genetic algorithm is representative of the HPV genome which can be used for diagnostics and drug development purposes.

We assessed carrageenan's potential to inhibit human papillomavirus (HPV) DNA extraction and amplification in vaginal swab samples collected in a trial, assessing the efficacy of a carrageenan-based gel against HPV infections. Experiment #1 consisted of adding gel (carrageenan-containing or placebo) to swabs and comparing HPV DNA detection by polymerase chain reaction (PCR) to unmanipulated samples collected from the same participants. For Experiments #2 and #3, we tested vaginal samples for inhibition by addition of an internal control and amplification by real-time PCR. Experiment #4 investigated carrageenan's interference with the extraction process by assessing HPV45 detectability in undiluted and diluted HPV45 positive samples (n = 3) with carrageenan versus no gel. In Experiment #1, there was a loss of HPV positivity with the addition of carrageenan (n = 9), but none with placebo gel (n = 5). In Experiments #2 and #3, the absence of the amplified product was observed in samples from the carrageenan arm: 3.3% (1/30) and 0.5% (1/199) of samples. In Experiment #4, HPV45 was not detected in undiluted carrageenan-containing samples, but after 1/50 dilution, the same HPV45 copy number was detected. Carrageenan does not affect the DNA extraction process, and inhibition of HPV DNA amplification by carrageenan occurs infrequently.

Background: The pathogenesis of penile intraepithelial neoplasia (PeIN) is unclear but human papillomavirus (HPV) infection and polymorphisms in human leucocyte antigen (HLA). Objectives: To examine the prevalence of HPV DNA and HLA in PeIN. Methods: Adult Caucasian men with a clinical and histological diagnosis of PeIN, that is, Bowenoid papulosis (BP), Bowen’s disease of penis (BDP) and erythroplasia of Queyrat (EQ) were selected and phenotyped from the clinical records. DNA was extracted from blood and paraffin-embedded sections for HLA and HPV typing, respectively. Human leucocyte antigen allele frequencies were compared with those derived from the UK–based Caucasian population. Results: Seventy-two cases of PeIN (20 BP, 34 BDP and 18 EQ) were studied. Human papillomavirus DNA was identified in 65/72 (90.2%) PeIN; Alphapapillomavirus types were detected in 62/72 (85%) followed by Betapapillomavirus types in 9/72 (12.5%) and cutaneous types in 7/72 (9.7%); HPV16 was the most prevalent genotype at 35/72 (48.6%) followed by HPV33 at 7/72 (9.7%); multiple infections were seen in 18/72 (25%) PeIN. HLA-C*15 (Bonferroni corrected p = 0.049) confers susceptibility to PeIN, whereas HLA-DQA1*01 (corrected p = 0.02) protects against PeIN. HPV16-associated PeIN cases showed no statistically significant association with HLA genotype after multiple corrections. Conclusion: Human papillomavirus is involved in the pathogenesis of PeIN. Immunogenotype may play a role in the pathogenesis of PeIN.

We report two cases of histologically documented pseudoepitheliomatous keratotic and micaceous balanitis in middle-aged male patients, which showed positivity for low-risk serotype human papillomavirus DNA. To our knowledge, only one other case has been documented. Further immunohistochemical proliferative markers were performed and compared to literature findings in penile epithelial proliferations. Evolution to invasive verrucous carcinoma has been associated with absence of HPV DNA. Thus, if confirmed by further studies, HPV testing should be included in pseudoepitheliomatous keratotic and micaceous balanitis assessment to address prognosis, and management.

Human papillomaviruses (HPVs) have been associated with a subset of head and neck squamous cell carcinomas (HNSCCs). The aim of this study was to determine the prevalence of HPV DNA in archived formalin-fixed paraffin-embedded tissue from patients with histologically confirmed HNSCCs in a South African cohort. A nested PCR was used for the detection of HPV DNA targeting the L1 gene. Positive samples were confirmed using an in-house hemi-nested PCR targeting the E6 gene and genotyped by sequence determination of amplicons. HPV DNA was detected in 57/780 (7.3%) samples, with the highest prevalence being in the sinonasal tract (16.0%) and oropharynx (10.8%). HPV16 was the most frequently detected type, being found in 26/57 (45.6%) positive samples. The prevalence of HPV DNA in HNSCCs found in this study was lower than that found in developed countries.

Low- and middle-income countries carry a high burden of preventable cervical cancer cases and deaths. Because human papillomavirus DNA-based testing is increasingly becoming the preferred method of screening for cervical cancer prevention, this commentary discusses next steps and key considerations for its expansion.

The relationship between infection with human papillomavirus (HPV) and tumorigenesis of salivary gland remains controversial.This study explored the relationship between HPV and salivary gland lesions as well as that of the HPV infection status and p16Tissue samples from 31 submandibular gland lesions were evaluated. p16Non-neoplastic lesion, benign tumors, and malignant tumors were noted in 9, 16, 6 cases, respectively. p16HPV-DNA positivity and p16

Summary Background Although previous meta-analyses have examined human papillomavirus (HPV) DNA prevalence in penile cancer, none, to our knowledge, have assessed pooled HPV DNA prevalence in penile intraepithelial neoplasia or p16INK4a percent positivity in penile cancer and penile intraepithelial neoplasia. Therefore, we aimed to examine the prevalence of HPV DNA and p16INK4a positivity in penile cancer and penile intraepithelial neoplasia worldwide. Methods In this systematic review and meta-analysis, we searched PubMed, Embase, and the Cochrane Library until July 24, 2017, for English-language articles published from Jan 1, 1986, onwards reporting the prevalence of HPV DNA and p16INK4a positivity, either alone or in combination, in at least five cases of penile cancer or penile intraepithelial neoplasia. Only studies that used PCR or hybrid capture for the detection of HPV DNA and immunohistochemical staining or methylation for the detection of p16INK4a were included. Data were extracted and subsequently crosschecked, and inconsistencies were discussed to reach consensus. Using random-effects models, we estimated the pooled prevalence and 95% CI of HPV DNA and p16INK4a positivity in penile cancer and penile intraepithelial neoplasia, stratifying by histological subtype and HPV DNA or p16INK4a detection method. Type-specific prevalence of HPV6, HPV11, HPV16, HPV18, HPV31, HPV33, and HPV45 in penile cancer was estimated. Findings Our searches identified 1836 non-duplicate records, of which 73 relevant papers (71 studies) were found to be eligible. The pooled HPV DNA prevalence in penile cancer (52 studies; n=4199) was 50·8% (95% CI 44·8–56·7; I2=92·6%, pheterogeneity Interpretation A large proportion of penile cancers and penile intraepithelial neoplasias are associated with infection with HPV DNA (predominantly HPV16), emphasising the possible benefits of HPV vaccination in men and boys. Funding None.

High-risk human papillomavirus (hrHPV) infection is a cause of cervical cancer development. The addition of hrHPV testing to cervical cancer screening and monitoring of cervical intraepithelial neoplasia treatment improves the efficacy of screening and treatment, respectively. Self-sampling for hrHPV testing seems a promising tool for increasing patient participation in cervical cancer screening. In this project, 1198 cervical swabs obtained by physicians and 176 cervicovaginal swabs obtained by self-sampling (not collected in parallel) were analyzed for the presence of 14 hrHPV genotypes using three commercially available assays in comparison. HPV DNA was detected in 21.2% of all samples (21% of cervical swabs and 22.7% of cervicovaginal swabs). The cobas 4800 HPV Test was the most sensitive (0.983) and specific (0.992) for hrHPV detection overall. The PapilloCheck HPV-Screening and LMNX Genotyping Kit HPV GP had comparable specificity with that of the cobas (0.989 and 0.955, respectively), but lesser sensitivity (0.897 and 0.909, respectively). In physician-obtained cervical swabs, the cobas showed the highest sensitivity and specificity (0.980 and 0.994, respectively) for hrHPV detection, whereas in cervicovaginal swabs, the cobas had the highest sensitivity (1.00), but the PapilloCheck had the highest specificity (0.993). In conclusion, all of the detection methods evaluated were highly sensitive and specific for hrHPV detection from both clinician-collected cervical swabs and self-sampled cervicovaginal swabs.

Purpose To determine whether plasma human papillomavirus (HPV) DNA predates clinical recurrence and compare its accuracy with 3-month fluorodeoxyglucose positron emission tomography (FDG-PET) in locally advanced cervical cancer. Methods This prospective multicenter study accrued 23 women with stage IB to IVA cervical cancer planned for definitive chemoradiation therapy (CRT). Plasma HPV DNA was measured serially by digital polymerase chain reaction, and FDG-PET was performed at 3 months post-CRT. Results Of the 19 women with HPV+ cervical cancer included in this analysis, 32% were stage IB, 58% IIB, and 10% IIIB/IVA. Median follow-up was 24 months (range, 18 to 30 months). All patients had detectable plasma HPV DNA before treatment. Six patients had detectable plasma HPV DNA at the end of CRT, and three of them developed metastases at 3 months. Of the 13 patients with undetectable plasma HPV DNA at end of CRT, to date, only one has developed recurrence. Six of those 13 patients had a positive 3-month FDG-PET with no definite residual disease on subsequent imaging or clinical examination to date, and four of these six had undetectable plasma HPV DNA at 3 months. Patients with undetectable plasma HPV DNA at end of CRT had significantly higher 18-month progression-free survival than those with detectable plasma HPV DNA (92% v 50%; P = .02). The area under the receiver operating characteristic curve (accuracy) of 3-month plasma HPV DNA and 3-month FDG-PET imaging for predicting recurrence at 18 months were 77% and 60%, respectively ( P = .008). Conclusion Detectable plasma HPV DNA at end of CRT predates the clinical diagnosis of metastases and is associated with inferior progression-free survival. Moreover, 3-month plasma HPV DNA level is more accurate than 3-month FDG-PET imaging in detecting residual disease. The clinical utility of plasma HPV DNA detection for guiding adjuvant/salvage therapy should be evaluated in future studies.

Efficient primary (vaccination) and secondary (screening) prevention strategies have the potential to eliminate cervical cancer worldwide. In this context, surveillance of HPV infections remains mandatory to assess the efficacy and the impact of screening and vaccination policies. Therefore there is a need to safely store cervical samples to conduct long-term studies in vaccinated and non-vaccinated subjects. Up-dated data on cervical specimen preservation on FTA® cards indicate that HPV DNA can be safely retrieved after 54 months of storage. A concordance of 97 % was achieved between HPV genotypes detected in initial cervical samples and on FTA® cards 4.5 years later. Even if a drop in HPV viral loads was observed in some cases at 4.5 years, using FTA® cards for safe and long-term storage of cervical samples represents an interesting option.

To investigate trends in prevalence of various human papillomavirus types in order to assess the unmet clinical needs for women affected by human papillomavirus-related disease. Data of consecutive 15 138 patients undergoing human papillomavirus DNA testing from 1998 to 2018 were retrospectively identified. Human papillomavirus types were classified at high-risk according to the classification of the International Agency for Research on Cancer. The International Agency for Research on Cancer included seven human papillomavirus types covered by nine-valent vaccine and five not yet covered by any available vaccines. Overall, 4159 (65.3%), 1500 (23.5%) and 714 (11.2%) women had human papillomavirus types covered by nine-valent vaccination, not covered by nine-valent vaccination and co-infections of human papillomavirus types of both groups. At least one high-risk human papillomavirus type(s) was detected in 1241 patients with genital dysplasia: 832 (67.1%), 291 (23.4%) and 118 (9.5%) women had human papillomavirus types covered by nine-valent vaccination, not covered by nine-valent vaccination and co-infections of human papillomavirus types of both groups. Over the twenty-year study period, the number of human papillomavirus types not covered by nine-valent vaccine increased dramatically (from 4 to 16%; P < 0.001, P for trend). Similarly, looking at patients with genital dysplasia, high-risk human papillomavirus types not covered by nine-valent vaccine increased from 3 to 13% (P < 0.001, P for trend). Our data highlight that human papillomavirus types covered by nine-valent vaccine represent are the main types associated with genital dysplasia. However, over the study period, we observed an increasing prevalence of confections and high-risk human papillomavirus types not covered by the nine-valent vaccine, thus suggesting the need of developing more complete vaccines against human papillomavirus.

Introduction: Nearly all cervical cancer cases are caused by one of several high-risk strains of the human papillomavirus (hr-HPV). Transmasculine (TM) individuals (persons who have a masculine spectrum gender identity, but were recorded female at birth) have low adherence to standard cervical cancer screening modalities. Introduction of self-collected vaginal swabs for hr-HPV DNA testing may promote initiation and adherence to cervical cancer screening among TM individuals to narrow screening disparities. The purpose of this study was to assess the rate of cervical cancer screening among TM individuals following the introduction of self-collected swabbing for hr-HPV DNA testing in comparison to clinician-administered cervical specimen collection. Methods: Rates of uptake and adherence to cervical cancer screening among TM individuals were assessed before and after the clinical introduction of self-collected swab testing in October 2017. Rates were compared with the rates of cervical cancer screening among cisgender women at a colocated Comprehensive Health Program during the time period of review. Results: Of the 121 TM patients seen for primary care in the 6-month baseline period before the October 2017 introduction of self-collected swabbing for hr-HPV DNA testing, 30 (25%) had cervical cancer screening documented in the electronic medical record. Following the implementation of self-swabbing, of 193 patients, 98 (51%) had a documented cervical cancer screening, a two-fold increase in the rates of adherence to cervical cancer screening (p

This study evaluated the presence of Human Papillomavirus (HPV) DNA in the cervix and peripheral blood of women with cervical intraepithelial neoplasia (CIN I, II, and III) and healthy individuals. Overall, 139 paired peripheral blood and cervix samples of healthy women and women with CIN I, II, and III (n = 68) were tested for HPV DNA by using standard procedures. Polymerase chain reaction (PCR) sequencing determined HPV types. Quantification of HPV16 E6 and E2 genes was performed to determine viral load and physical state. HPV DNA was detected in the cervix (21.1% in healthy individuals, 48.8–55.5% in CIN patients), blood (46.4% in healthy individuals, 44.1–77.7% in CIN patients) and paired peripheral blood and cervix samples (24% in healthy individuals, 32.5–44.4% in CIN patients). The most frequent types found in the cervix were HPV16, 18, 31, 33, 58, and 70, while HPV16, 18, 33, 58, and 66 were the most frequent types found in the blood. HPV DNA in the cervix was associated with previous sexually transmitted infections (STIs) (p = 0.023, OR: 2.978, CI:1.34–7.821), HPV DNA in the blood (p = 0.000, OR: 8.283, CI:3.700–18.540), and cervical lesions (CIN I/II or III) (p = 0.007). Binomial logistic regression showed that HPV DNA in the blood (p = 0.000, OR: 9.324, CI:3.612–24.072) and cervical lesions (p = 0.011, OR: 3.622, CI:1.338–9.806) were associated with HPV DNA in the cervix. However, we did not find an association between HPV DNA in the blood and cervical lesions (p = 0.385). Our results showed that only HPV DNA found in the cervix was associated with cervical lesions.

Human papillomavirus (HPV) was proven to play a significant role in cancer development in the oropharynx. However, its role in the development of laryngeal (LSCC) and hypopharyngeal squamous cell carcinoma (HPSCC) remains to be clarified. High-risk HPV (HR-HPV) viral proteins E6 and E7 are considered to be pertinent to HPV-related carcinogenesis. Hence, our aim was to estimate LSCC and HPSCC for HR-HPV DNA, p16, and E6/E7 oncoprotein status by using molecular virology and immunohistochemistry methods. The prevalence of HPV16 infection was 22/41 (53.7%) and 20/31 (64.5%) for LSCC and HPSCC, accordingly. The majority of HPV16+ tumor samples were stage III or IV. In most samples, the presence of either HPV16 E6 or HPV16 E7 viral protein in dysplastic or tumor cells was confirmed using immunohistochemistry. Our results suggest a high prevalence of HPV16 as a primary HR-HPV type in LSCC and HPSCC. The lack of HPV E6/E7 oncoproteins in some tumor samples may suggest either the absence of viral integration or the presence of other mechanisms of tumorigenesis. The utilization of p16 IHC as a surrogate marker of HR-HPV infection is impractical in LSCC and HPSCC.

6063 Background: HPV genomic DNA in plasma and saliva has been widely studied, however more recently, circulating tumor human papillomavirus DNA (ctHPVDNA) has emerged as a reliable biomarker for surveillance in HPV+ oropharyngeal squamous cell carcinoma (OPSCC). A commercial assay for this biomarker distinguishes tumor-derived viral DNA (tumor-tissue modified viral DNA or TTMV) from other non-cancer associated sources of HPV DNA. The use of this technology has been previously described in plasma, but its utility in saliva is currently unknown. Methods: A prospectively collected and banked biospecimen repository was used to identify 46 patients with HPV+ OPSCC with paired pre treatment plasma and saliva samples. All samples were assessed for DNA integrity and TTMV using a clinically validated ddPCR-based assay (NavDx™; Naveris Inc, Natick, MA) to measure TTMV for HPV-16, -18, -31, -33 and -35 from frozen plasma and saliva samples. Retrospective chart review was performed to collect clinical and pathological data. Graphpad was used for statistical analysis. Spearman’s r was used to correlate TTMV copies in saliva and plasma. Wilcoxon test was used to compare between sample types. Mann-Whitney test was used for categorical variables. Results: TTMV DNA was detectable in 43 of 46 plasma samples and in 44 of 46 saliva samples. One plasma sample failed quality control measures, one of each sample type had undetectable TTMV, and one of each type was indeterminate. Of 41 evaluable patients with paired samples, there were 38 (93%) males, 36 (88%) were stage I-II, 5 (12%) were stage III-IV (AJCC 8th, clinical staging), and 25 (61%) had a history of smoking with a median of 37.5 pack years. TTMV was significantly enriched in saliva compared to plasma (p10 pack-year history (p=0.011). Conclusions: This is the first study to demonstrate successful quantification of tumor-tissue modified HPV DNA in saliva. Compared to plasma, pre treatment saliva samples demonstrated significantly higher levels of TTMV. TTMV distinguishes ctHPVDNA from other sources of HPV. These data highlight the potential use of TTMV detection in saliva for early detection of HPV+ OPSCC as well as its potential role in local surveillance after treatment. More research is needed to elucidate the effects of smoking on TTMV levels.

The aim of this survey was to examine the knowledge, attitudes, and behavior of gynecologists in terms of human papillomavirus (HPV) DNA testing as a primary screening tool for cervical cancer. A national cross-sectional web survey was carried out through multistage sampling using an overall sample of 1000 gynecologists. Gynecologists were asked to fill in a self-administered questionnaire exploring their knowledge, attitudes, and practice toward cervical cancer screening and HPV-DNA testing. A total of 582 gynecologists completed the web questionnaire. Of these, 24.5% were uncertain on the higher sensitivity of HPV-DNA compared with the Pap test, whereas 19% were uncertain on the role of the HPV-DNA test as a primary test in women younger than 30 years old and only 44.9% knew that a negative HPV-DNA test allows for an extension of the test interval to 5 years. Most gynecologists showed a definite positive attitude on the role of screening for cervical cancer prevention and were prepared to accept new technologies. The HPV-DNA test was considered highly effective by 86.9%, whereas 94% recommend/perform HPV-DNA tests in women older than 30 years of age; 25.5% performed HPV-DNA as a primary test, followed by a Pap test in those cases that were positive. Only 56.3% recommended/performed HPV-DNA tests 1 year after a positive HPV-DNA test, followed by a negative Pap test, whereas 42.9% recommended colposcopy. Although the use of the HPV-DNA test is very widespread among Italian gynecologists performing cervical cancer screening, there is lack of standardization of practices according to current guidelines.

There is plenty of substantial evidence to support anti-tumor activity of viruses. Adeno-associated virus (AAV) may interact with human papillomavirus (HPV) to modify the risk of cervical neoplasia. The seroprevalence of AAV among women with cervical cancer has been reported to be lower than healthy ones. In spite of this finding, detection of AAV DNA in cervical biopsies does not entirely support the inverse association between AAV seropositivity and cervical cancer. This association is still controversial and requires more thorough evaluation in different countries. The aim of this case-control study was to find the prevalence of AAV and HPV DNA sequences in Iranian women with and without cervical cancer to assess the probable association of AAV infection and cervical cancer. In this study, paraffin-embedded tissue samples of 61 cervical cancer cases and 50 healthy controls (HCs) were investigated for AAV and HPV DNA by semi-nested and nested PCRs respectively. AAV DNA was detected in 7 cases (14%) of HCs and 9 specimens (14.8%) of case group. According to the branching in the phylogenetic tree, AAV2 was the only type detected in this study. Moreover, HPV DNA was detected in 8 cases (16%) of HCs and 44 specimens (72.13%) of case group. In conclusion, a low proportion of cervical biopsies from Iranian women contained AAV-2 genome. No significant difference in correlation between HPV and cervical cancer in presence or absence of AAV genome in cervix was found.

Objective To evaluate whether HPV DNA in urine has potential advantages as an alternative biomarker for HPV-based cervical cancer screening. Methods Among patients with Cobas HPV test results, a total of 67 HPV-positive (n = 42) and -negative (n = 25) women who agreed to participate in this study were willing to provide paired cervical and urine samples, and we observed concordance between sample types from each patient in identifying HPV genotypes using the nanowire assay. Results We detected high-risk strains of HPV DNA in unprocessed urine specimens using polyethyleneimine-conjugated nanowires (PEI-NWs). Concordance for high-risk HPV (hrHPV) between paired urine and cervical samples was 90.4% (κ = 0.90; 95% CI: 0.80–100.00). The virological sensitivity and specificity for detection of HPV DNA from a small urine sample (200 μL) were 81.3% (κ = 0.83; 95% CI: 62.1–100.0) and 98.0% (κ = 0.83; 95% CI: 94.2–100.0) for HPV16 group, 100.0% (κ = 0.65; 95% CI: 100.0–100.0) and 95.3% (κ = 0.65; 95% CI: 90.1–100.0) for HPV18 group, and 96.4% (κ = 0.97; 95% CI: 89.6–100.0) and 100.0% (κ = 0.97; 95% CI: 100.0–100.0) for other hrHPV group, respectively. Conclusions The nanowire assay demonstrated excellent ability to identify HPV DNA from urine specimens. We observed an excellent agreement in the detection of high-risk HPV between paired urine and cervical samples, even with small urine sample volume.

Background Testing for oral high-risk human papillomavirus (HPV) DNA may be useful for identifying individuals at increased risk for HPV-driven oropharyngeal cancer (OPC). However, positivity for HPV DNA provides no information on the transforming potential of the infection. In contrast, the detection of high-risk HPV E6/E7 messenger RNA (mRNA) may help to identify clinically significant infections because of the indispensable role of E6/E7 viral oncoproteins in the carcinogenic process. Methods Oral rinses were collected with a mouthwash from cancer-free individuals at increased risk for oral HPV infection. High-risk HPV DNA and mRNA were evaluated via the testing of the oral rinses with the Linear Array HPV genotyping test and the Aptima HPV assay, respectively. Results Overall, 310 subjects with no clinical evidence of lesions of the oral cavity and oropharynx were included in the study. Thirty-three (10.6%) harbored high-risk HPV DNA in their oral rinse. These cases, together with 10 random samples negative for high-risk HPV DNA, were tested with the Aptima assay. A valid result was obtained for 41 of the 43 specimens (95.3%). Among the 31 cases that were positive for high-risk HPV DNA and had a valid Aptima result, 4 (12.9%) were positive for HPV mRNA. HPV mRNA was not detected in any of the samples negative for high-risk HPV DNA. Conclusions HPV mRNA is detectable in oral rinses of cancer-free subjects. Oral HPV mRNA testing may be useful in the screening and/or early detection of HPV-driven OPC by possibly identifying active and transforming oral infections. The testing of individuals at increased risk for HPV-related OPC via simply and noninvasively collected oral specimens is an attractive option for future screening strategies.

Introduction: Oral Lichen Planus (OLP) is a chronic inflammatory disease with cell mediated immunopathologic response. Even though OLP has been studied widely for decades, the aetiology of the disease is yet not hypothesised exactly. Autoimmunity, viruses like Hepatitis C Virus (HCV), Human Papillomavirus (HPV), Human Immunodeficiency Virus (HIV), Herpes Simplex Virus (HSV), Candida albicans and stress have been hypothesised in recent articles. The fact that there is paucity of studies on the aetiopathogenesis of oral lichen planus pivoted the study to analyse the presence of HPV in OLP. Aim: The aim of this study is to detect the presence of HPV DNA in the biopsy samples of OLP and also to determine the role played by the virus in aetiopathogenesis of OLP. Materials and Methods: Biopsy samples comprising 15 Oral Lichen Planus tissues and 15 normal buccal mucosa tissues embedded in paraffin blocks were collected from different dental colleges located in Telangana erstwhile Andhra Pradesh. Parameters like age, sex, site of lesions, type of lesion were also compared. The DNA was extracted from the fixed tissue by using modified protocol and Polymerase Chain Reaction (PCR) was performed by using general primers to amplify HPV L1 gene. On comparison of sequences of 10 patients by using Basic Local Alignment Search Tool (BLAST), the sequences were found similar to HPV-18 L1 gene. Results: Thirteen out of 15 (86.6%) lichen planus cases and none out of 15 controls were HPV 18 positive. Compared between males and females and controls significant relation was observed between sex and OLP too. However, no significant relation was observed based on the site of the lesion and based on type of lesion. Conclusion: The molecular studies confirm presence of HPV18 in OLP tissues of 13 patients with 8 Single Nucleotide Polymorphism (SNPs) indicating probably new variant of HPV18 in Indian patients.

Epidemiological and molecular biological data suggest that the beta human papillomavirus (HPV) can cause the development of a number of epithelial skin tumors, but this interconnection is currently not fully understood. Besides detection of beta HPV, it is necessary to measure and quantify the viral load, it is a new approach in the diagnosis of HPV infection. 52 patients were examined to identify the association of epithelial neoplasias beta human papilloma virus in immunocompromised and immunocompetent patients in the department of chronic hemodialysis and kidney transplantation of Moscow Regional Research and Clinical Institute. HPV detection was performed by polymerase chain reaction (PCR) with hybridization-fluorescence detection in real time (Real-Time). Amplification and detection were performed on the tool Rotor-Gene 3000 (“Corbett Research”, Australia). For quantification of beta HPV recombinant plasmid positive controls, as well as control plasmidfragment offi-globin human gene (FSIS CRIE Rospotrebnadzor) were used. In fibroepithelial polyp and in apparently healthy skin of immunosuppressive patients - renal transplant recipients (RPT) with a high frequency was detected beta HPV DNA - 64 and 54%, respectively, compared to healthy skin (47% of cases). In fibroepithelial polyp in 57% of immunosuppressive RPT mixed infection was revealed. Viral load in the fibroepithelial polyp was higher than that as in apparently normal skin and in donors skin. The high detection ofHPVDNA in RTP, as in the fibroepithelial polyp and in apparently healthy skin was found. Immunocompetent patients had high HPV DNA detection only in fibroepithelial polyp compared with apparently healthy skin.

Cervical cancer, being the fourth leading cause of cancer death in women worldwide, predominantly originates from a persistent infection with a high-risk human papillomavirus (HPV). Detection of DNA sequences from these high-risk strains, mostly HPV-16 and HPV-18, represents promising strategy for early screening, which would help to identify women with higher risk of cervical cancer. In developing countries, inadequate screening options lead to disproportionately high mortality rates, making a fast and inexpensive detection schemes highly important. Electrochemical sensors and assays offer an alternative to current methods of detection. We developed an electrochemical-chip based assay, in which target HPV DNA is captured via magnetic bead-modified DNA probes, followed by an antidigoxigenin-peroxidase detection system at screen-printed carbon electrode chips, enabling parallel measurements of eight samples simultaneously. We show sensitive detection in attomoles of HPV DNA, selective discrimination between HPV-16 and HPV-18 and good reproducibility. Most importantly, we show application of the assay into both cancer cell lines and cervical smears from patients. The electrochemical results correlated well with standard methods, making this assay potentially applicable in clinical practice.

Context All Food and Drug Administration–approved methods in the United States for human papillomavirus testing including the Hybrid Capture 2 human papillomavirus assay and the Roche cobas human papillomavirus test are approved for cytology specimens collected into ThinPrep media but not for specimens collected into SurePath solution. Objective To compare the performance of the Roche cobas and Hybrid Capture 2 tests for the detection of high-risk human papillomavirus using both ThinPrep and SurePath preparations as part of a validation study. Design One thousand three hundred seventy-one liquid-based cytology samples, including 1122 SurePath and 249 ThinPrep specimens, were tested for high-risk human papillomavirus DNA using the Roche cobas human papillomavirus test and the Hybrid Capture 2 human papillomavirus assay. For cases with discrepant results, confirmatory testing was performed using Linear Array human papillomavirus testing. Results One hundred and fifty-six (11.38%) and 184 (13.42%) of the 1371 specimens tested positive for high-risk human papillomavirus DNA using the Hybrid Capture 2 human papillomavirus assay and Roche cobas human papillomavirus assay, respectively. In addition, 1289 (94.0%) of 1371 specimens demonstrated concordant high-risk human papillomavirus results with a κ value of 0.72 (95% confidence interval, 065–0.78). There was no statistically significant difference in the percentage of positive high-risk human papillomavirus results between the 2 liquid-based preparations with either assay. Discordant results between the 2 assays were noted in 82 of 1371 cases (6%). Twenty-seven of 82 cases (32.9%) were Hybrid Capture 2 positive/Roche cobas negative and 55 of 82 cases (67.1%) were Roche cobas positive/Hybrid Capture 2 negative. Two of 20 Hybrid Capture 2–positive/Roche cobas–negative cases (10%) and 26 of 37 Roche cobas–positive/Hybrid Capture 2–negative cases (70%) tested positive for high-risk human papillomavirus by Linear Array. Conclusions Both assays showed good agreement and excellent specificity with either ThinPrep or SurePath preparations. The number of discordant results was relatively small. The performance of both assays was similar for ThinPrep specimens, but the Roche cobas test demonstrated higher sensitivity with SurePath specimens.

e12582 Background: It has been demonstrated an increased risk of breast cancer (BC) incidence in patients with previous cervical dysplasia, suggesting a role of human papillomavirus (HPV) of cervical lesions in the development of BC. Although, the origin of HPV spreading to the mammary gland and its mechanism of dissemination is not clear. Methods: Seven serum samples from healthy donors and 58 from early BC patients collected pre-surgery were analyzed for the presence of HPV DNA. For 49/58 patients HPV DNA was analyzed also on the primary tumor tissue. 17 patients had luminal A BC (4 relapsed, 13 non relapsed), 16 had luminal B BC (5 relapsed, 11 non relapsed), 13 had triple-negative BC (6 relapsed, 7 non relapsed), 12 had HER2-positive BC (4 relapsed, 8 non relapsed). Circulating DNA was extracted from 500 μl of serum by Qiamp DNA Mini kit (Qiagen, Milan, Italy) and tumor DNA was extracted from at least four 10-micron sections by QIAamp DNA FFPE Tissue Kit (Qiagen, Milan, Italy). Circulating HPV DNA was amplified by a multiplex PCR with HPV E6 or E7 gene-specific primers and the sequence was assessed by a high-throughput MALDI-TOF mass spectrometry-based method. Results: HPV DNA was detected in only 5 serum of BC patients and in none of the healthy controls. 4/5 BC cases had high-risk HPV DNA (type 39,45,52,59) and 1 had type 73 low-risk HPV DNA. 4/5 HPV DNA-positive patients had previously low-grade cervical intraepithelial neoplasia (CIN I) detected by Pap smear. These 5 patients with circulating HPV DNA did not show HPV positivity in the BC tissue. 2 out of 49 cases were positive for universal HPV DNA sequence in tissue and only 1 case showed HPV type 51. No relation was found between HPV infection and tumor subtype or prognosis, neither for HPV DNA positivity between serum and tissue. Conclusions: Our data support the feasibility of HPV DNA detection by liquid biopsy in BC. The presence of circulating HPV could be due to a viral spread from other organs. More data are needed to establish the role of circulating HPV DNA and its potential association with HPV infection of the breast and/or of the cervix. [Table: see text]

Background Accurate and internationally comparable human papillomavirus (HPV) DNA detection and typing services are essential for HPV vaccine research and surveillance. Objectives This study assessed the proficiency of different HPV typing services offered routinely in laboratories worldwide. Study design The HPV Laboratory Network (LabNet) has designed international proficiency panels that can be regularly issued. The HPV genotyping proficiency panels of 2013 and 2014 contained 43 and 41 coded samples, respectively, composed of purified plasmids of sixteen HPV types (HPV 6, 11, 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68a and 68b) and 3 extraction controls. Proficient typing was defined as detection in both single and multiple infections of 50 International Units of HPV 16 and HPV 18 and 500 genome equivalents for the other 14 HPV types, with at least 97% specificity. Results Ninety-six laboratories submitted 136 datasets in 2013 and 121 laboratories submitted 148 datasets in 2014. Thirty-four different HPV genotyping assays were used, notably Linear Array, HPV Direct Flow-chip, GenoFlow HPV array, Anyplex HPV 28, Inno-LiPa, and PGMY-CHUV assays. A trend towards increased sensitivity and specificity was observed. In 2013, 59 data sets (44%) were 100% proficient compared to 86 data sets (59%) in 2014. This is a definite improvement compared to the first proficiency panel, issued in 2008, when only 19 data sets (26%) were fully proficient. Conclusion The regularly issued global proficiency program has documented an ongoing worldwide improvement in comparability and reliability of HPV genotyping services.

Background Squamous papillomas are exophytic proliferations of surface oral epithelium. Human papillomavirus (HPV) infection is widely accepted as the etiology of squamous papillomas however the virus cannot be detected in a significant percentage of lesions. Material and methods Using polymerase chain reaction (PCR), we tested 35 formalin-fixed paraffin-embedded (FFPE) squamous papillomas for the presence of HPV DNA. Results Six papillomas (17%) tested positive for HPV DNA; four contained HPV-6 and two contained HPV-11. Given that β-globin DNA was only identified in half of the samples, DNA degradation appears to have significantly impacted the results. Conclusions The results likely represent an underestimation of the true number of HPV-positive specimens in our study. Potential explanations for HPV-negative squamous papillomas include transient HPV infection, failure of the experiment to detect HPV if present, or the possibility that some lesions may not result from HPV infection. Key words:HPV, PCR, FFPE, papilloma, oral.

It is crucial to understand the epidemiology and natural history of human papillomavirus (HPV) infection in both men and women, to prevent the increasing HPV-related disease burden in men. Data on HPV prevalence among men in the general population are limited. In this cross-sectional population-based study, we aimed to estimate genital HPV infection prevalence in Danish men using 2 different test methods.Penile swab samples from 2460 male employees and conscripts at military barracks in Denmark were tested for HPV DNA with the hybrid capture 2 (HC2) method, and a polymerase chain reaction (PCR) assay, Inno-LiPA. The overall and age- and type-specific prevalence of HPV infection with 95% confidence intervals (CIs) were estimated, and the correlation between the 2 assays was assessed.The overall HPV prevalence was 22.2% (95% CI, 20.6-23.9) in the HC2 test and 41.8% (95% CI, 39.9-43.8) with PCR. Of the PCR-positive samples, 50.9% were negative in the HC2 test. Of 183 PCR-positive samples that could not be genotyped (HPVX), 88.0% (95% CI, 83.2-92.7) were HC2 negative. The most prevalent types were HPV-51, HPV-16, HPV-66, HPV-53, and HPV-6. The prevalence of high-risk and low-risk HPV peaked among men aged 20 to 29 years, whereas the HPVX prevalence increased with age.Human papillomavirus is highly prevalent in the general male population of Denmark, with HPV-16 and HPV-51 being the most prevalent. Polymerase chain reaction detects twice as many positive samples as HC2 but includes HPVX, possibly representing cutaneous HPV types found on normal genital skin.

Cytological screening has substantially decreased the cervical cancer incidence, but even better protection may be achieved by primary high-risk human papillomavirus (hrHPV) screening. In the Netherlands, five-yearly cytological screening for women aged 30-60 years will be replaced by primary hrHPV screening in 2016. The new screening guidelines involve an extension of the screening interval from 5 to 10 years for hrHPV-negative women aged 40 or 50 years. We investigated the impact of this program change on the lifetime cancer risks in women without an hrHPV infection at age 30, 35, 40, 45 or 50 years. The time to cancer was estimated using 14-year follow-up data from a population-based screening intervention trial and the nationwide database of histopathology reports. The new screening guidelines are expected to lead to a reduced cervical cancer risk for all age groups. The average risk reduction was 34% and was smallest (25%) among women aged 35 years. The impact of hrHPV screening on the cancer risk was sensitive to the duration from cervical intraepithelial neoplasia grade 2/3 (CIN2/3) to cancer; a small increase in the cancer risk was estimated for women aged 35 or 40 years in case a substantial proportion of CIN2/3 showed fast progression to cancer. Our results indicate that primary hrHPV screening with a ten-yearly interval for hrHPV-negative women of age 40 and beyond will lead to a further reduction in lifetime cancer risk compared to five-yearly cytology, provided that precancerous lesions progress slowly to cancer.

ObjectiveTo compare the clinical characteristics of four human papillomavirus (HPV) assays: hybrid capture 2 (HC2), cobas, CLART, and APTIMA in Danish women with abnormal cytology.MethodsSurePath samples from 367 consecutive women from Copenhagen, with atypical squamous cells of undetermined significance or worse, were tested with the four assays. These women were routinely recommended for repeated testing or were referred for colposcopy. Their worst histological diagnosis in 29months from baseline was retrieved from the Danish National Pathology Data Bank.ResultsOf the 367 women, 16 (4%) had no follow-up, 232 (63%) had

Background: Cervical cancer is a major cause of cancer mortality in women and more than a quarter of its global burden is contributed by developing countries. In India, in spite of alarmingly high figures, there is no nationwide government-sponsored screening program. This study was conducted to assess the burden of cervical cancer in India and review the performance characteristics of available cervical cancer screening tools, so as to provide evidence-based recommendations for application of most practically suited screening test to be used in resource-poor field settings. Materials and Methods: MEDLINE and Web of Science electronic database were searched from January 1990 to December 2015, using the keywords such as “cervical cancer”, “screening”, “early detection”, “cervical cytology” and “visual inspection”, and their corresponding MeSH terms in combination with Boolean operators “OR, AND.” Two authors independently selected studies that are published in English and conducted in India. A total of 11 studies were found to be relevant and eligible to be included in the present study. Results: In India, cervical cancer contributes to approximately 6–29% of all cancers in women. The age-adjusted incidence rate of cervical cancer varies widely among registries; highest is 23.07/100,000 in Mizoram state and the lowest is 4.91/100,000 in Dibrugarh district. The pooled estimates of sensitivity and specificity of visual inspection with acetic acid (VIA), magnified VIA, visual inspection with Lugol's iodine (VILI), cytology (Pap smear), and human papillomavirus DNA were found to be 67.65% and 84.32%, 65.36% and 85.76%, 78.27% and 87.10%, 62.11% and 93.51%, and 77.81% and 91.54%, respectively. Conclusions: In developing countries because of lack of necessary infrastructure and quality control, high-quality cytology screening may not be feasible for wide-scale implementation. Hence, cervical cancer screening program based on visual screening test such as VIA/VILI should be adopted as an integral part of primary health-care setup in resource-poor countries like India.

Hybrid Capture 2 (HC2) has been demonstrated to be a feasible screening method for cervical cancer. Based upon HC2 technology, careHPV is a simple, rapid, accurate, and inexpensive screening test for women in low-resource settings. This study aims to characterize both the careHPV test and HC2 test, and to compare careHPV results of specimens stored in careHPV test collection medium (TCM) to HC2 results from partner specimens stored in Qiagen specimen transport medium and TCM. The positive rates of high-risk HPV in careHPV, HC2, and HC2 (TCM) were 13.2% (108/818), 13.2% (108/818), and 13.6% (111/818), respectively. The agreement rates of pairwise tests were 95.8% (95% CI: 94.5-97.2%), 96.7% (95% CI: 95.5-97.9%), and 97.2% (95% CI: 96.1-98.3%), respectively. The Kappa values of the pairwise tests were 0.82 (95% CI: 0.76-0.88), 0.86 (95% CI: 0.81-0.91), and 0.88 (95% CI: 0.83-0.93), respectively. Based on these findings, although careHPV is demonstrated to be a viable alternative to the HC2 test, improvements on the careHPV test are still required prior to its implementation as a suitable screening method for women in low-resource settings. Further studies on the significance and applicability of the careHPV test must be performed.

To investigate the utility of urine sampling for detecting human papillomavirus (HPV) DNA among pregnant women and to compare HPV DNA detection in urine with detection in vaginal samples.In a cross-sectional study, urine and vaginal samples were self-collected from pregnant women attending prenatal care at Hospital Divina Providencia, Frederico Westphalen, Brazil, between October 2006 and August 2007. Part of the L1 region of the HPV genome was amplified via GP5(+)/bioGP6(+) primers. Positive urine was genotyped for high-risk HPV genotypes (HPV16, HPV18, HPV31, HPV33, HPV39, HPV45, and HPV59).During the study period, urine samples were obtained from 133 pregnant women, 63 of whom also self-collected vaginal samples. HPV DNA was detected in 54.0% (34/63) and 61.9% (39/63) of urine and vaginal samples, respectively. HPV infection was significantly associated with first intercourse at younger than 20 years of age (P=0.008). There was substantial agreement in HPV DNA test results between the urine and vaginal samples (κ value, 77.3%; P0.0001). HPV31 and HPV16 accounted for 80.7% of the oncogenic types identified.Detection of HPV DNA in urine showed good agreement with detection in self-collected vaginal samples, indicating that urine might be a reliable sample for HPV testing among pregnant women.

OBJECTIVE Kaiser Permanente Northern California (KPNC) introduced 3-year Pap and human papillomavirus DNA cotesting for cervical cancer screening in women 30 years or older in 2003 to 2004. Patient and provider willingness to extend screening intervals and the impact on annual cervical cancer incidence after interval extension are evaluated. MATERIALS AND METHODS Age-adjusted cervical cancer rates and screening intervals were calculated from KPNC Regional Laboratory databases and Northern California Cancer Registry from 2000 to 2009. RESULTS The median screening interval between negative cotests was 36 months compared to the 16 months after a negative Pap test alone before the implementation of cotesting. The age-adjusted invasive cancer rate was 6.5 per 100,000 women in 2000 and 6.3 in 2009; there was no difference in the rates of cervical cancer in women 30 years or older from 2000 to 2009 (p(trend) = .7). CONCLUSIONS Patients and providers were compliant with the extension of screening intervals with cotesting. Cervical cancer rates remained constant during the 10-year study period despite extending screening intervals after a negative cotest.

Background: Variable detection of human papillomavirus (HPV) DNA can result in misclassification of infection status, but the extent of misclassification has not been quantitatively evaluated. Methods: In 2005–2007, 33 women of ages 22 to 53 years self-collected vaginal swabs twice per week for 16 consecutive weeks. Each of the 955 swabs collected was tested for 37 HPV types/subtypes. Assuming that a woman's underlying infection status did not change over the short study period, biases in prevalence estimates obtained from single versus multiple swabs were calculated. Using event history analysis methods, time to recurrent gain and loss of at least one HPV type was determined, separately. Baseline any-type and high risk–type HPV prevalence was 60.6% and 24.2%, respectively. Cumulative any-HPV and high-risk HPV prevalence over the 16-week period was 84.8% and 60.6%, separately. Results: Overall, there were 319 events of detection and 313 events of loss of detection. Median times to a recurrent detection and loss of detection were 11 and seven days, respectively. Neither vaginal sex nor condom use during follow-up was associated with recurrent viral detection or loss of detection. Assuming the cumulative 16-week prevalence reflects the true prevalence of infection, the baseline any-HPV prevalence underestimated infection status by 24.2%, with a bootstrapped mean of 20.2% [95% confidence interval (CI), 8.9%–29.6%]. Conclusions: These findings suggest that a substantial proportion of HPV-infected women are misclassified as being uninfected when using a single-time DNA measurement. Impact: Short-term variation in detectable HPV DNA needs to be considered while interpreting the natural history of infections using single samples collected at long intervals. Cancer Epidemiol Biomarkers Prev; 23(1); 200–8. ©2013 AACR.

ABSTRACT: Papillomavirus-associated cervical cancer remains a major disease burden for young women. Therefore, it plays an important role in the pathological diagnosis. Besides cytological and classical pathological methods, molecular biology tests are used for the analysis of HPV status. Here we compare three PCR assays for the detection of human papillomavirus DNA from 50 gynecological samples, namely the HPV Genotyping Kit HPVsign® (Qiagen, Germany), a novel HPV-genotyping kit based on pyrosequencing, the LCD array HPV Type 3.5 (Chipron, Germany) and an in-house PCR of the National Reference Center for Papilloma- and Polyomaviruses at the Institute for Virology of the University Cologne (Germany). The HPVsign assay mainly detected the assumed leading HPV type, while both the in-house PCR and the Chipron HPV Assay 3.5 reproducibly detected single and multiple infections.

Purpose: Head and neck squamous cell carcinoma of an unknown primary site (HNSCCUP) is a heterogeneous group of tumors that includes the human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma. To investigate the relationship between HNSCCUP and HPV, we reviewed p16 overexpression and HPV DNA in lymph node metastases and examined their correlation with the primary site and clinical features. Materials and Methods: Thirty-three patients with HNSCCUP were retrospectively studied. Dissected neck metastases were analyzed for p16 overexpression by immunohistochemistry, and the presence of HPV DNA was investigated by in situ hybridization. Results: Of the 33 patients, 8 (24%) exhibited p16 overexpression. p16-positive lymph node metastases contained significantly more HPV DNA and were most frequently associated with occult primary lesions in the oropharynx and a favorable prognosis. Patients with a lower alcohol consumption, only level II/III metastasis, and cystic lymph node metastasis tended to have p16 overexpression. Conclusions: This is the first report on the relationship of HNSCCUP with p16 and HPV DNA status in Asian patients. In total, 24% of the HNSCCUP patients were p16 positive. p16 overexpression in neck metastasis was predictive of both an occult primary lesion in the oropharynx and an association with HPV infection. Alcohol consumption, location, and features of neck metastasis were correlated with p16 expression.

Pathology departments routinely process and store formalin-fixed, paraffin-embedded (FFPE) tissue samples for clinical diagnosis. These collections often contain decades' worth of samples and represent a treasure trove of specimens that can be analyzed for retrospective epidemiological studies, diagnostics, and pathogen discovery. Accurate amplification and sequencing of DNA from these samples is critical for the usability of these FFPE samples. Here we present a collection of protocols that describe extraction of DNA from FFPE tissues, PCR amplification of human papillomavirus DNA, and subsequent genotyping of the infecting virus. © 2016 by John Wiley & Sons, Inc.

To test the hypothesis that self-collected urine could be used to detect high-risk human papillomavirus (HPV) DNA with sensitivity and specificity comparable to those of standard cervical testing.Women attending a gynecology clinic for evaluation of abnormal cytology were recruited. Fifty-two participants (21-60years of age) collected urine samples, and clinicians collected cervical brush samples. When appropriate, cervical biopsies were obtained during colposcopy. HPV detection and typing were performed on DNA extracts from each sample, using commercial reagents for L1 consensus polymerase chain reaction (PCR) and type-specific hybridization. HPV 16 viral load was determined by quantitative PCR in HPV 16-positive samples. A diagnostic test analysis was conducted for urine samples.Fifty paired samples were analyzed, with 76% agreement between samples. The 12 discrepant pairs were all urine negative/cervix positive. The most common HPV types detected were 16, 51, 53, and 62. The urine test correctly identified 100% of the uninfected and 65% of the infected patients.The results indicate that HPV DNA detection using urine is less sensitive than cervical sampling in a population with abnormal cytology. Further exploration is warranted to determine clinical utility when other options are unavailable.

The novel HPV Direct Flow CHIP commercial system for Human Papillomavirus (HPV) genotyping is based on rapid PCR and automatic reverse dot blot hybridization to genotype-specific probes, allowing the detection of 36 HPV genotypes. This study examined the performance of HPV Direct Flow CHIP in formalin-fixed paraffin-embedded (FFPE) samples (n= 99). Each sample was analyzed both by Direct PCR, using crude cell extracts without DNA purification, and by conventional PCR, using purified DNA. Pair-wise analysis of the results demonstrated strong concordance between the results obtained with the two protocols, although a slightly higher rate of multiple infections was detected by conventional PCR. In summary, HPV Direct Flow CHIP achieves effective HPV detection from FFPE samples with both Direct PCR and Conventional PCR protocols.

We evaluated the effect of storage at 2 to 8°C on the stability of human genomic and human papillomavirus (HPV) DNA stored in BD SurePath and Hologic PreservCyt liquid-based cytology media. DNA retained the ability to be extracted and PCR amplified for more than 2.5 years in both medium types. Prior inability to detect DNA in archived specimens may have been due to failure of the extraction method to isolate DNA from fixed cells.

Aim: To detect the prevalence and type distribution of human papillomavirus (HPV) in different cytological diagnostic categories. Materials and methods: Between 2007 and 2010, a total of 1014 liquid-based thin preparations of cervical smears were selected and classified according to cytology results. HPV DNA polymerase chain reaction (PCR) was also performed using these samples. HPV DNA-positive samples were genotyped by DNA sequencing. Results: Of those enrolled in the study, 45.3\% were negative cytologically, 36.4\% had atypical squamous cells of undetermined significance, 0.3\% had atypical squamous cells preventing the exclusion of a high-grade squamous intraepithelial lesion, 16.8\% showed a low-grade squamous intraepithelial lesion, and 1.3\% had an high-grade squamous intraepithelial lesion. Conclusion: PCR assays showed HPV positivity in 63.0\% of cytologically negative and 90.8\% of cytologically positive samples. The most common types of HPV detected were 16, 6, 18, 31, 66, 56, 53, 81, 45, and 62. Of HPV DNA-positive samples, 47.7\% were high, 4.7\% were intermediate, and 17.9\% were low risk. The high-risk types of HPV detected were 16, 18, 31, 56, 53, 45, 62, 58, 59, 67, 51, 35, 73, 52, 33, 39, 68, and 82.

Background: There is a need to simplify cervical cancer screening to reach more women. Tampon-collected specimens can be tested using molecular methods, but this type of self-screening has not been properly evaluated as a screening method in South Africa before. The objective of this study was to evaluate human papillomavirus (HPV) DNA testing of self-collected tampons as a screening method in an urban and peri-urban population in Gauteng by comparing the results with the current standard of conventional cytology. In addition, HPV prevalence, type, distribution and incidence of cytological abnormalities in this population are described. Method: Seven hundred and twenty women attending public healthcare facilities in and around Tshwane, Gauteng province, were invited to participate. The women collected a tampon sample for molecular testing, and were then screened by healthcare workers collecting a conventional cervical cytology smear. HPV testing was undertaken using the Linear Array® HPV Genotyping Test (Roche Molecular Systems). Results: Data for analysis were available for 631 women. Three hundred and fifty-four (58%) were positive for highrisk HPV, while (15.4%) had an abnormal cytology result. Women aged 30-39 years had the highest prevalence of both high-risk HPV (75%) and abnormal cytology (22%). Infection with multiple types was common. Higherrisk viruses were not over-represented in, and no dramatic decrease in HPV prevalence was observed in, older women. Cytological abnormalities were detected in only 3.74% of women who tested negative for high-risk HPV, but were found in 24.2% of high-risk HPV positive women. Conclusion: HPV testing on self-collected tampon samples was feasible, highly sensitive and demonstrated a high negative predictive value for current cytological abnormalities in this population.

The discovery of the necessary cause of cervical cancer, human papillomavirus (HPV), has led to important technological advances, including the development of molecular tests for HPV to identify women with cervical precancerous lesions. HPV testing has proven to be more sensitive and more reliable, albeit less specific, for detection of cervical precancer than cytologic methods of detection. As the result, HPV testing can reduce the incidence of cervical cancer within 4-5 years and reduce the mortality due to cervical cancer within 8 years compared to cytology. Additionally, a negative HPV test provides greater reassurance against cervical cancer than a negative Pap test. HPV testing, because of its attributes, is useful for screening out low-risk women who do not need further intervention for 5 or more years. Thus, HPV testing can shift the emphasis of the use of Pap testing or any other more specific diagnostic test from frequent use in the entire population to the ~10% subset of women who tested positive for the causal factor, HPV. Here, we highlight the current and future status of the introduction of HPV testing into routine cervical cancer screening. This article forms part of a special supplement entitled "Comprehensive Control of HPV Infections and Related Diseases" Vaccine Volume 30, Supplement 5, 2012.

Increasing evidences support the role of Merkel cell polyomavirus (MCPyV) and human papillomavirus (HPV) in non-cutaneous and cutaneous tumours. Porocarcinoma is a rare malignant neoplasm that arises from the intraepidermal ductal portion of the eccrine sweat glands. The aetiology of porocarcinoma is largely unknown and no systematic studies have been done to investigate the implication of infectious agents in the pathogenesis of this tumour.To investigate the possible association between MCPyV and/or HPV infection and porocarcinoma.Forty-four formalin-fixed paraffin-embedded (FFPE) porocarcinomas (40 primary and 4 metastatic) and 10 healthy skin specimens (controls), were analysed for the presence of MCPyV and HPV DNA using molecular detection methods.MCPyV DNA was found in 27/40 (68%) primary porocarcinomas and in 3/10 (30%) controls (Fisher exact test: p0.04). No significant difference in viral load was observed between tumours and healthy skin. Moreover, 2/40 primary porocarcinomas tested positive for high-risk HPV16. Cutaneous beta-HPV infection was detected in 16/40 (40%) porocarcinomas and in 6/10 (60%) controls. No particular beta-HPV types were significantly associated with tumour or with healthy skin. Two out of 4 metastatic biopsies were MCPyV DNA positive. All metastatic samples had mixed infections with cutaneous HPV types.This study demonstrated a significantly high prevalence of MCPyV and the presence of a broad spectrum of HPV types in porocarcinoma and provided the first available data about viral infections in this tumour. To understand the role, if any, of viral infections in the pathogenesis of porocarcinoma further studies are needed.