Those living in low- andmiddle-income countries and/or low-resource settings remain at increased risk fordeveloping and dying as a result of cervical cancer, largely a result of a lack ofaccess to routine screening and follow-up monitoring and treatment of dysplasticlesions.3 The Papanicolaou test/cytology, which remains the goldstandard for cervical cancer prevention and early detection, requires extensiveresources, including highly trained physicians and complex laboratoryequipment.2 Places without suchinfrastructure have historically been vulnerable to cervical cancer disparity.Widespread adoption of screening has led to dramatic decreases in the global burden ofcervical cancer.6 Many of these have found that point-of-care HPV testing,especially when paired with self-sampling, is both highly sensitive and cost effectivefor primary cervical cancer screening. However, few previous studies have examinedoptimal methods for the triage of women with abnormal primary screenings. The article byPoli et al7 fills that gap. It presentsan innovative study comparing VIA, cytology, and colposcopy for triage among a largesample of underserved women who screened positive by point-of-care HPV testing inHyderabad, India. The primary aim of this inquiry was to determine the optimal triagestrategy within this and other comparable, low-resource settings.Several previous studies have examined the relative feasibility, acceptability, andefficacy of various primary screening strategies, including Papanicolaou test, humanpapillomavirus (HPV) self-sampling, and visual inspection with acetic acid (VIA), inimproving cervical cancer screening uptake and dissemination within resource-limitedsettings.7 concluded that VIA was the optimal triage technique forthe study setting, given that primary screening with point-of-care HPV testing, VIAtriage, and treatment could all be conducted within the same clinic visit. Moreover,point-of-care HPV testing with VIA triage was identified as the most cost effectiveamong each screening algorithm considered.The researchers considered a multitude of factors when evaluating each triage strategy,including test sensitivity and logistic barriers  to implementation. Although they found that VIA was indeed less sensitivethan cytology for identifying cervical dysplasia, Poli et alStudy findings highlight the importance of considering contextual factors in determiningthe optimal screening algorithms in resource-limited settings. Although cytologyperformed substantially better than VIA in terms of sensitivity, both the cost andfeasibility of cytology for triage of women with HPV-positive disease were prohibitivewithin this low-resource context. Ultimately, although point-of-care HPV testing islikely to usurp VIA as the best practice for primary screening in many low-resourcesettings, the current study demonstrates how VIA can be used to effectively triage womenfor colposcopy and treatment when access to cytology is challenged.Another notable aspect of the study was the use of community health workers and mobilescreening strategies to promote the uptake of cervical cancer screening among the localpopulation. HPV self-sampling has been shown to have high acceptability across a varietyof marginalized populations worldwide, and when delivered by culturally competent healthworkers, has been shown to circumvent structural and cultural barriers to cervicalcancer prevention, such as beliefs regarding modesty and discomfort with the speculumexamination. Moreover, the establishment of mobile screening protocols will likelyincrease the effectiveness and reach of the point-of-care HPV testing/VIA triageapproach. Notably, neither of these screenings require a clinic setting to besuccessfully conducted.Ultimately, these findings lend themselves to several areas of future inquiry. Thelarge-scale effectiveness of the point-of-care HPV testing/VIA triage method has yet tobe established. Moreover, this triage method may also be advantageous within otherresource-limited settings. For example, although VIA has been successfully used in ruralHaiti as part of a screen-and-treat approach, the addition of point-of-care HPV testingas a primary screening strategy has not yet been examined. It is likely that thisscreening algorithm will have a substantial impact on improving the sensitivity andeffectiveness of low-resource screening programs worldwide. Finally, although thecurrent point-of-care HPV test, careHPV, has been shown to be effective in a variety ofcontexts, the test requires 2.5 hours to be completed. This is a vast improvement overtraditional HPV testing, although it may still prohibit same-day triage and treatment incertain contexts. Thus, future work is needed to optimize point-of-care testing andreduce the amount of time necessary to receive test results.


Extra II (INNO-LiPA) within the VALGENT-3 framework. VALGENT is designed to assess the analytical and clinical performance of HPV tests with genotyping capacity. The VALGENT-3 panel comprised 1300 consecutive cervical cell specimens enriched with 300 samples with abnormal cytology obtained from women attending the Slovenian cervical cancer screening programme. The INNO-LiPA allows type-specific detection of 32 HPV types; however, for the clinical accuracy assessment, we considered it as high-risk (hr)HPV positive when at least one of the following HPV types was present: HPV16, HPV18, HPV31, HPV33, HPV35, HPV39, HPV45, HPV51, HPV52, HPV56, HPV58, HPV59, and HPV68. Clinical accuracy for detection of cervical intraepithelial neoplasia grade 2 or worse (CIN2+) was compared between INNO-LiPA and Hybrid Capture 2 (HC2), which is a standard comparator test for HPV tests used in cervical cancer screening. In addition, hrHPV and type-specific detection HPV types were compared between INNO-LiPA and Linear Array HPV Genotyping Test (Linear Array). The prevalence of hrHPV determined by INNO-LiPA was 17.1%  in the screening population. HrHPV testing with INNO-LiPA had a sensitivity for CIN2+ of 96.9%  which was non-inferior to HC2  and a specificity for ≤CIN1 of 85.3%  which was inferior to HC2 . Genotyping agreement between INNO-LiPA and Linear Array was excellent for hrHPV, HPV16, HPV18, HPV35, HPV45, HPV58 and HPV59, but good or fair for other HPV types. To conclude, INNO-LiPA demonstrated non-inferior clinical sensitivity but lower specificity compared to HC2 in addition to excellent concordance compared to Linear Array for hrHPV and some genotypes.In this diagnostic test validation study, we assessed the clinical accuracy and HPV genotyping performance of the INNO-LiPA HPV Genotyping  Over 200 human papillomavirus (HPV) types have been identified and classified based on their nucleotide sequences, with new HPV types being characterized at an increasing rate . Among tMany HPV tests are available on the market but only few have been clinically validated for use in primary screening settings . The HybExtra II assay  evaluated in the current study is the most recent assay launched by the company in 2015, targeting 32 types, four types more than the previous version. This new version contains genotype specific probes for more decisive genotyping results, an upgraded SPF10 primer set resulting in improved sensitivity , improved human DNA control primers and provides a ready-to-use amplification reagent [INNO-LiPA HPV genotyping assay, based on the principle of reverse hybridization after highly sensitive PCR amplification with SPF10 primers, have been used for HPV genotyping over two decades ,21. Duri reagent .In the present study, the VALidation of HPV GENotyping Tests (VALGENT) framework was used to evaluate the clinical accuracy of INNO-LiPA in comparison with HC2. For the first time, it was verified whether hrHPV testing with INNO-LiPA fulfills the minimal requirements for use in primary cervical cancer screening . In addiThe characteristics of the VALGENT-3 study population, including demographics, cytological and histological results have been described previously ,24. Of tThe accuracy data for the INNO-LiPA and HC2 for the outcomes CIN2+, CIN3+ and ≤CIN1 are shown in pmcn = 0.6547; pn.inf = 0.0002) for CIN2+ and 1.01  for CIN3+. The relative specificity of INNO-LiPA for ≤CIN1 was 0.95 . Similar results were obtained if the analysis was restricted to women ≥30 years  lower than the comparator test.INNO-LiPA is a SPF10 PCR that targets a short highly conserved region in the L1 gene ,26. The INNO-LiPA provides for each HPV type a qualitative output, which is translated into a positive/negative result. The appreciation of presence or absence of blue lines is not quantifiable. Therefore, adaptation of the cut-off, which may allow a more optimal balance between clinical sensitivity and specificity, is in case of INNO-LiPA not possible.pmcn < 0.001) in the total study population. The significant difference of positivity rate for hrHPV with these two genotyping methods is consistent with the tests’ clinical performances with regard to relative specificity compared to HC2.Excellent analytical agreement between INNO-LiPA and Linear Array was observed for 13 hrHPV types overall, HPV16, HPV18, HPV35, HPV45, HPV58 and HPV59. In addition, INNO-LiPA detected more positive cases than Linear Array for all individual HPV types that are common to both assays, with a positivity rate of hrHPV determined by INNO-LiPA statistically significantly higher than that determined by Linear Array  following the manufacturer’s instructions. Bound DNA was finally eluted with 50 µL of ATE buffer and stored at −70 °C prior to further testing. According to the manufacturer’s instructions, we have used 10 µL of extracted DNA for INNO-LiPA testing. Similarly, from the second aliquot of original ThinPrep DNA was extracted for Linear Array testing, where 50 µL of extracted DNA was used for further testing.The sample collection, aliquoting procedure and storage details have been described in detail in previous VALGENT-3 manuscripts ,24. BrieThe INNO-LiPA is a line probe assay based on the principle of reverse hybridization for qualitative detection and identification of 32 different HPV types, including 13 hrHPV , 6 possible hrHPV , 9 low-risk HPV  plus 4 other HPV genotypes . INNO-LiPA uses the biotinylated consensus primers (SPF10) to amplify a 65-bp region within the L1 region of multiple alpha HPV types. The resulting biotinylated amplicons are then denatured and hybridized with specific oligonucleotide probes. A primer set for the amplification of the human HLA-DPB1 gene is included to monitor sample quality and extraction. The INNO-LiPA assay  was performed fully automated using the AutoBlot 3000H . Interpretation of the developed strips was done by scanning and automated interpretation using with the LiRAS for LiPA HPV software . The test was performed in accordance with the manufacturer’s instructions.HC2 detects 13 hrHPV types  and is accepted as a standard comparator test for the clinical validation of hrHPV DNA assays that may be used for primary cervical cancer screening . For theThe Linear Array is an HPV test with full genotyping capacity, which detects 37 high- and low-risk HPV types  that is frequently used in virological and epidemiological research. In the present study, Linear Array is used as a comparator test to evaluate the analytical genotyping accuracy of the INNO-LiPA.As described in previous VALGENT-3 reports ,24, cytoWomen with histologically confirmed CIN2+ results were considered as diseased subjects. Due to the fact that women with normal cytological results of negative for intraepithelial lesion or malignancy (NILM) were not referred to colposcopy verification in our study, we considered them as subjects without disease only if they had two or more consecutive NILM cytological results (at enrolment and at subsequent screening between 12 to 48 months later). We used this group of women to compute the clinical specificity for ≤CIN1.The clinical sensitivity and specificity of the INNO-LiPA for CIN2+ and CIN3+ were calculated. We compared the clinical accuracy of INNO-LiPA to HC2 for CIN2+ and CIN3+, using non-inferiority statistics with a relative sensitivity threshold of 90% and a relative specificity threshold of 98% . The McNp-value of <0.05 indicates significant discordance between genotyping results determined by the two assays.Separate and consensus genotyping agreement for the types common to INNO-LiPA and Linear Array was assessed using κ  and McNe


Atypical squamous cell of undetermined significance (ASCUS) is a common cervical cytological diagnosis. At present, HPV DNA assay is used to triage these patients, but its lower specificity brings a series of problems. The purpose of this study was to evaluated the value of p16/Ki67 immunostaining, HPV E6/E7 mRNA testing in triaging women with ASCUS by comparing HPV DNA assay.Liquid based cytology specimens were collected from 300 patients. P16/Ki67 immunocytochemistry using the CINtec® Plus Kit and HPV E6/E7 mRNA testing by QuantiVirus®HPV E6/E7 mRNA assay used the same cytology sample. Detection rates of each test were evaluated against histopathology.All assays yielded a high sensitivity for the detection of CIN3+ (100% (86.7–100) for HPV DNA assay, 88.0% (70.0–95.8) for HPV E6/E7 mRNA testing and 100% (86.7–100) for p16/Ki67 immunocytochemistry) and CIN2+ (98.2% (90.2–99.7) for HPV DNA assay, 87.0% (75.6–93.6) for HPV E6/E7 mRNA testing, 98.2% (90.2–99.7) for p16/Ki67 immunocytochemistry). The specificity to detect high grade dysplasia was highest for p16/Ki67 immunocytochemistry (74.2% (68.7–79.0) in CIN3+ and 82.5% (77.3–86.8) in CIN2+), followed by HPV E6/E7 mRNA testing (39.6% (34.0–45.5) in CIN3+ and 42.7% (36.7–48.9) in CIN2+) and HPV DNA assay (16.0% (12.1–20.8) in CIN3+ and 17.5% (13.2–22.7) in CIN2+).p16/Ki67 immunostaining and HPV E6/E7 mRNA testing, especially the former, may be promising tools in triage of ASCUS. Cervical cancer screening based on the Papanicolaou (Pap) smear is widely accepted as the most important public health strategy and have reduced cervical cancer incidence and mortality during the last decades in many countries . The goaINK4a (p16) is a tumor suppressor protein playing a crucial role in cell-cycle regulation. p16-overexpression is considered as a surrogate marker for deregulated E7 expression and hence for transforming HPV infections [Recently, many disease-specific molecular markers of cervical cancer have been recognized based on our recognition of HPV-related carcinogenesis. Only the persistent infection and malignant transformation in cervix can cause the occurrence and development of cervical malignant transformation . The mosfections , 11. Ki6fections , 13. Undfections . Many difections .HPV E6/E7 mRNA test has been proposed as a biomarker for HPV oncogene expression. Previous studies showed HPV E6/E7 mRNA test has a very good clinical sensitivity and higher specificity compared to HPV DNA detection –18. p16/The study population consisted of women who attended outpatient gynecological screening at the gynecology department of The Third Affiliated Hospital of Zhengzhou University between 2015 and 2017. Women meeting these conditions are enrolled in our study:(1) Cytological diagnosis for ASCUS; (2) non-pregnancy; (3) No immunodeficiency disease; (4) No cervical surgery or chemo and/or radiotherapy for cervical malignant disease. A total of 300 women were consecutively included and met those conditions in this study. All women underwent Pap test, colposcopy biopsy and histopathological examination. After the Pap test was done, the residual LBC specimen was used for assays evaluated in this study, HPV E6/E7 mRNA and p16/Ki67 immunocytochemistry. The Ethics Committee of The Third Affiliated Hospital of Zhengzhou University reviewed and approved our study. All enrolled women were aware of the research purposes and signed the consent forms. The median age was 36 years (interquartile range 18–70 years).LBC technology was used to perform Cytological detection. We used Thin Prep 2000 Processor  to make Thinlayer slides according to the manufacturer’s instructions. Two experienced cytopathologists independently diagnosed cytological specimens at the same time without knowing the results of the other assays. If the two cytopathologists gave different diagnosis, cervical samples were reviewed again. Using the 2001 Bethesda Reporting System Criteria to report the diagnosis of cytological specimens . The anaCytology slide for p16/Ki67 immunocytochemistry was prepared from the residual LBC specimen using a T2000 slide processor . We purchased CINtec® Plus Kit  to carry immunocytochemistry of p16/Ki67 according to the manufacturer’s instructions. An experienced cytotechnologist reviewed all cervical cytology slides for detecting the staining performance of two markers. If ≥1 cervical epithelial cell stained both with a brown cytoplasmic stain (p16) and a red nuclear (Ki67) no matter how abnormal the morphology of the cells were, this case was considered as positive of p16/Ki67 immunocytochemistry. Slides without brown cytoplasmic stain and /or a red nuclear stain were called negative .Residual Pap specimens were also used to detect the 14 types of HR-HPV E6/E7 mRNA by QuantiVirus®HPV E6/E7 mRNA assay  according to the manufacturer’s instructions. The assay used branched DNA (b-DNA) technology , which don’t need the step of RNA purification or RT-PCR to quantitatively detect HPV mRNA. In each sample, the result of mRNA was marked as light unit. Special calculation software can convert light unit to copy number under the fact of light emission related directly to the amount of HPV mRNA. If the copy number was equal or greater than 1.0, the result of E6/E7 mRNA was positive. If less than 1.0, negative is the result .Hybrid Capture 2 assay  was used to detect 13 types of HR-HPV DNA according to the manufacturer’s instructions. If the number of RLU/CO equal or greater than 1.0, the result of HPV DNA was positive.INK4a-histology kit . For CIN2, it diagnosed clinically as CIN2 when p16-histology positive, diagnosed clinically as CIN1 when p16-histology negative.Women with ASCUS underwent colposcopy biopsy within 4 weeks after cervical exfoliative cytology test. In this study, some patients with ASCUS but a negative HR-HPV DNA result required to do colposcopy biopsy because of fear of suffering from cervical malignant disease, thus we underwent colposcopy biopsy for them. Two experienced pathologists independently diagnosed histological slides without knowing the results of the other assays. If the two pathologists gave different diagnosis, histological slides were diagnosed again for a consensus result. The diagnosis result of histological slides meets the standard of current World Health Organization classification. The diagnoses of cervical inflammation and mild cervical intraepithelial neoplasia  are referred as CIN1-. cervical inflammation, CIN1 and Moderate cervical intraepithelial neoplasia  are referred as CIN2-. CIN2, severe cervical intraepithelial neoplasia  and carcinoma are referred as CIN2 +. CIN3 and carcinoma are referred as CIN3+. In this study, all histological specimens were subjected to p16-immunohistochemistry. The staining was performed on 2 μm sections from formalin fixed, paraffin embedded tissues according to the manufacturer’s instructions using the CINtec p16p value < 0.05 was considered as statistical significance for all analysis.SPSS 21.0 software was used to perform statistical analysis. Wilcoxon rank sum test of two independent samples was used to compare the differences of the expression levels of HPV E6/E7 mRNA between different groups. Chi-square tests were used to compare the differences of percentage between different groups. The gold standard for cervical disease was histologically confirmed CIN2+ and CIN3+. Diagnostic sensitivity and specificity for the detection of CIN2+ and CIN3+ was calculated for all assays based on 2 × 2 tables and results are given with 95% confidence intervals. Youden’s index was calculated as sensitivity% + specificity% - 100. Receiver operating characteristic curve (ROC) was used to compare the diagnostic performance of different tests and find the best diagnostic threshold for the E6/E7 mRNA expression testing. A total of 300 women with ASCUS underwent cervical histopathology diagnosis. 46.0% (138/300) of ASCUS had cervical inflammation, 36.0% (108/300) had CIN 1, 9.7% (29/300) had CIN2 and 7.3% (22/300) had CIN3. Three cases of cervical squamous cell carcinoma were diagnosed in this study (1.0%). Among them, 54 cases were diagnosed as CIN2+ (18.0%) and 25 cases as CIN3+ (8.3%).We calculated the positive rate of different methods in different levels of cervical tissue. The overall test positivity was 85.3% (256/300) in HPV DNA assay, 62.7% 188/300) in HPV E6/E7 mRNA testing and 32.0% (96/300) in p16/Ki67 immunocytochemistry. HPV E6/E7 mRNA testing and p16/Ki67 immunocytochemistry were less frequently positive in ASCUS than HPV DNA test. The p16/Ki67 immunocytochemistry positivity was 32.0%, indicating that if we use p16/Ki67 immunocytochemistry instead of HPV DNA assay to triage ASCUS, we will reduce the rate of colposcopy referrals by 53.3% . This trend was also seen in HPV E6/E7mRNA testing and p16/Ki67 immunocytochemistry, and also seen between CIN2- and CIN3+ (Table HPV DNA positive rate was 82.5% (203/246) in CIN1-, 98.1% (53/54) in CIN2+, and the differences between the lesions were statistically significant (P < 0.05).The expression level of HPV E6/E7 mRNA testing in different cervical lesions is shown in Fig. χ2 = 3.375, P = 0.066 (HPV DNA assay vs. HPV E6/E7 mRNA testing), χ2 = 3.375, P = 0.066 (HPV E6/E7 mRNA testing vs. p16/Ki67 immunocytochemistry), χ2 = 0.000 (HPV DNA assay vs. p16/Ki67 immunocytochemistry)), but the specificity is statistical difference , χ2 = 83.377, P < 0.001 (HPV E6/E7 mRNA testing vs. p16/Ki67 immunocytochemistry), χ2 = 208.13, P < 0.001 (HPV DNA assay vs. p16/Ki67 immunocytochemistry)). ROC curve was used to determine an optimal cut-off value and further demonstrate the diagnostic performance of HPV E6/E7 mRNA testing for detecting CIN2+ , but a higher specificity(P = 0.0001). Increasing the p16/Ki67 threshold to two or more dual stain-positive cells led to a substantial reduction in test positivity compared with cytology and the dual stain assay at the usual cutoff. At a cutoff of five or more dual stain-positive cells, the sensitivity for detection of CIN3+ was statistically significantly lower compared with both cytology and the dual stain assay at the usual cutoff, while the specificity and the PPV were statistically significantly increased [2 = 208.13, P < 0.001). These finding support that p16/Ki67 can be a viable option for ASCUS triage. In a review that compared HPV E6/E7 assay versus HPV DNA test (Hybrid Capture 2 method) in triage of women with ASCUS or LSIL cervical cytology, the pooled sensitivity and specificity of HPV E6/E7 assay to triage ASCUS to detect underlying CIN3 or worse was 96.2% (95% CI: 91.7–98.3%) and 54.9% (95% CI: 43.5–65.9%), respectively. HPV E6/E7 assay and HPV DNA test showed similar pooled sensitivity; however, the specificity of the former was significantly higher  [2 = 37.147, P < 0.001), while the sensitivity is similar (98.2% vs. 87.0%). HPV assays for detecting the mRNA of 5 hrHPV types may reduce the over-diagnosis of women who have minor cytologic abnormalities. However, given the lower sensitivity, women with negative mRNA test results cannot be considered free of CIN2+ and require further surveillance [The present study also evaluates the performance of HPV E6/E7 mRNA testing, p16/Ki67 immunocytochemistry and HPV DNA assay in triaging women with ASCUS in a cross-sectional clinical setting. HPV E6/E7 mRNA testing and p16/Ki67 immunocytochemistry were identified in recent studies to increase specificity for the detection of high grade cervical disease compared to HPV DNA detection . In womencreased . In our P = 0., but a heillance . It is ieillance . The dateillance . These rFrom our study, we can see that both p16/Ki67 immunocytochemistry and HPV E6/E7 mRNA testing have higher specificities than HPV DNA test, when we group analysis by 30 years old, the accuracy of each test is higher in the group over 30 years old. However, we didn’t find a study that solve such a problem: which is better in triaging ASCUS for p16/Ki67 immunocytochemistry and HPV E6/E7 mRNA testing. Our present data suggested that p16/Ki67 immunocytochemistry has a higher sensitivity than HPV E6/E7 mRNA testing . ROC curve was used to further demonstrate the diagnostic performance of p16/Ki67 immunocytochemistry, HPV E6/E7 mRNA testing and HPV DNA test for detecting CIN2+, we can see that the AUC of p16/Ki67 immunocytochemistry is the largest . The figure found that when the optimal cut-off value of HPV E6/E7 mRNA testing is selected at 882.53 copies/ml, the specificity of HPV E6/E7 mRNA testing is improved. Tong-Yu Liu’s and Ye-li Yao’s finding also supported this result , 35. HowBoth biomarker tests, HPV E6/E7 mRNA testing and p16/Ki67 immunocytochemistry, especially p16/Ki67 immunocytochemistry, could be a valuable test in such a setting with higher specificity for the detection of high-grade cervical neoplasia than HPV DNA detection without losing sensitivity. p16/Ki67 immunocytochemistry is promising to be used for the efficient detection of cervical precancer and cancers in triaging women with ASCUS. However, although our results are gratifying, further cost-effectiveness analyses have to be considered when implementing the two strategies for cervical cancer prevention.


The meta-analysis revealed a prevalence of 49% : 35–64%) of any type of human papillomavirus in men and 35% (95% CI: 26–45%) of high-risk human papillomavirus in men. The included studies showed that stable sexual habits, circumcision and condom use are protective factors against HPV. In addition, there is a certain positive association with tobacco use and the early initiation of sexual intercourse. In conclusion, the prevalence of HPV in men is high. The risk factors for HPV infection are sexual promiscuity, early sexual debut, absence of circumcision, lack of condom use and smoking. Further study in this field about the effectiveness of the vaccine and health education should be conducted.Human papillomavirus (HPV) is one of the most prevalent sexually transmitted infections. Although the research focus has been on women, men are also affected. Thus, the aim was to estimate the prevalence of HPV in men and to analyse its risk factors. A systematic review with meta-analysis was performed. The main health science databases were consulted. The search terms were was: “papilloma virus AND  AND men”. The final sample of studies was  Human genital papillomavirus (HPV) is a common sexually transmitted infection (STI) which has become a major source of morbidity and mortality worldwide . More thAmong the low-risk subtypes, the most common serotypes include HPV-6 and HPV-11, which usually cause benign warts and are occasionally associated with non-invasive lesions. In contrast, serotypes HPV-16 and HPV-18 are considered high-risk subtypes, due to their high carcinogenic potential. HPV-16 appears primarily in invasive tumours and in those lesions with a high degree of malignancy, while HPV-18 is associated with poorly differentiated carcinomas and with increased lymph node involvement ,7.Most studies of HPV have analysed diagnosis, treatment and prevention in women. Moreover, strategies in sexual and reproductive health programmes in many countries have focused on epidemiological control in women, but they have tended to overlook the role of men in this infection, despite its high prevalence. In fact, it has been suggested that men may constitute a reservoir for inadvertently transmitting infection to women, due to its asymptomatic nature, thus contributing to the persistence of infection and cancer . A furthSeveral of the risk factors for HPV in men are associated with deficits in hygiene and prevention ,10. In fIn view of the above considerations, this study was undertaken to perform a meta-analysis of HPV prevalence in men and to analyse the corresponding risk factors. The question that guided this study was: what is the prevalence of HPV in men and which are its risk factors?A systematic review and meta-analysis of the literature was conducted, following the PRISMA  recommenThe analysis included quantitative studies on HPV in men published in English or Spanish, unrestricted by year. Studies lacking quantitative statistical information were excluded, as were studies with mixed samples without independent data on HPV in men, studies that associated HPV with other diseases, and studies focused on the genetics of the virus.A data collection notebook was used. For longitudinal studies with more than one HPV evaluation, the first measure obtained in the study population was recorded. The following variables were collected on the characteristics of the sample: year of publication, country of study, type of publication  and study design .Once the preliminary results obtained from each database, the title and abstract were read. The full text was then read. After this, a forward and reverse search was carried out within the studies included to locate as many documents as possible. Finally, the studies were read critically before the inclusion. The methodological quality was assessed with the checklist proposed by Ciaponni. The levels of evidence and degree of recommendation used were those proposed by the Oxford Centre for Evidence-based Medicine .2 index.Data analysis was performed with the StatsDirect meta-analysis package . The first step was to perform a sensitivity analysis to ensure that none of the studies included produced significant changes in outcomes when excluded. Publication bias were then assessed using Egger’s linear regression test. Prevalence and confidence intervals were calculated by random-effects meta-analysis, one for each burnout dimension. The heterogeneity of the sample was analysed by Cochran’s Q test and the In = 568 records were excluded because they were not related to the study topic, the sample also included women, the study design was not appropriate for this review, or due to the language of publication. After reading the full texts, the final sample was n = 16 after eliminating duplicates and applying the inclusion and exclusion criteria. The search and selection process is described in In total, 945 relevant studies were located. After reading the titles and abstracts, The meta-analytical estimation of the prevalence of any HPV in males was 49% with a 95% confidence interval of 35–62%. The forest plot is shown in p < 0.001) and the I2 index was 99.7%. For the high-risk HPV meta-analysis the Cochran Q-value was 1118.93 (p < 0.001) and the I2 index 99.1%.In the heterogeneity analyses of the any HPV meta-analysis, the Cochran Q-value was 3555.24 .A significantly higher risk of HPV detection was associated with an increased number of lifetime female sexual partners; OR 6.96–9.01 for non-oncogenic HPV (any type) and for oncogenic HPV among men with ≥50 partners versus only one partner . AccordiIngles et al.  reportedVarious studies have shown that circumcision is a protective factor against HPV infection ,19,20,24According to Tarnaud et al. , the mecMale circumcision is strongly associated with a lower probability of flat penile lesions. Backes et al.  reportedCondom use is considered a protective factor against HPV infection ,28,30. ASeveral studies have concluded that tobacco use aggravates the risk of HPV infection ,26. CurrA stronger association was observed between smoking 10 or more cigarettes per day (compared to smoking 0–9 cigarettes per day) and each of the following outcomes: detection of any HPV (OR 3.0 (95% CI: 1.4–6.4); oncogenic HPV (OR 3.7 (95% CI: 1.6–8.5); and non-oncogenic HPV (OR 2.4 (95% CI: 1.1–5.6]) .According to King et al. , based oEducation is considered to be a factor related to HPV infection . UniversAccording to Ingles et al. , genitalAkogbe et al.  conducteDue to the association between cervical cancer and HPV, men have not been the focus of HPV research . NotwithSexual promiscuity, related to the number of sexual partners, in men is an important risk factor due to multiple sexual contacts facilitate contamination by HPV, which is then transmitted to sexual partners . When seSome authors argue that the prevalence of HPV infection is associated with age, and hence with the pattern of sexual behaviour in the community. Thus, HPV prevalence is greater at earlier ages (15–25 years) with the initiation of sexual intercourse; later, between 25–40 years of age, there is a marked decrease, after which the prevalence stabilises . HoweverRegarding prevention, penile circumcision appears to be a protective factor against high-risk HPV infection. Circumcised men show a lower risk of HPV infection . In unciCondoms present an impermeable barrier to particles the size of the pathogenic microorganisms of STIs, preventing their transmission and acquisition by avoiding contact between the penis and genital secretions, the mucous membrane and the skin of the sexual partner. Therefore, condoms are considered to be a protective factor against HPV infection ,39.The tetravalent HPV vaccine provides effective prevention, as shown by the significant decrease in the rate of immune infection . HoweverSmoking is considered to induce added negative effects to HPV infection because cigarette smoke contains chemicals that damage the genetic structure of the human cells. As a result of this damage to the DNA, cells turn into oncogenic cells, which accelerates the appearance of genital lesions and evolution towards cancer .This study has some limitations. First, the number of studies is low because not all the studies did inform about the prevalence of HPV in men or its risk factors without mixing the data with female populations or using other diseases as main subjects of the study. Second, the analysis of the risk factors has been done individually, lacking multivariable analysis. Some variables may influence the other HPV risk factors, e.g., the educational level has been identified by some authors as a predictor of condom use, HPV awareness and HPV vaccination in males ,43. FinaTo our best knowledge, this is the first meta-analysis addressing the prevalence of HPV in the male population and associated risk factors. We have observed a prevalence of 49% in any HPV and of 36% in high risk HPV. Stable relationships, circumcision, condom use and non-smoking are potentially beneficial factors in preventing the development of HPV infection and thus, they should be given serious consideration. The issue of HPV infection in men warrants further research, particularly studies aimed at promoting the provision of a vaccine for this population group and educational campaigns aimed to change sexual behaviour, especially among young men.


Approximately 50% of cases of penile carcinoma (PeCa), a rare neoplasm worldwide, are associated with human papillomavirus (HPV). However, the detection of HPV-DNA is not sufficient to consider it the etiological factor in the development of this type of cancer. Currently, the overexpression of P16INK4A is used as a surrogate biomarker of HPV carcinogenesis. Information on PeCa in Mexico is scarce, particularly regarding cases related to HPV and genotype frequency.To evaluate the presence of HPV, its genotypes, and the presence of multiple genotypes, and the expression of P16INK4A, as well as its clinical and histopathological parameters.For HPV-DNA detection and P16INK4A expression, we used the INNO-LiPA® test and immunohistochemistry, respectively.Sixty cases of PeCa were evaluated, of which 75% were HPV-non-related histological variants. We found that 58.9% (33/56) of PeCa cases were HPV-DNA positive, while 30.9% of the cases evaluated (17/55) were positive for P16INK4A. HPV16 was the main genotype in 42.9% of the cases, followed by HPV52 in 7.1% and HPV18 in 5.4%. Within the HPV-positive cases, 27.3% had multiple genotypes. All HPV-positive patients under the age of 45 years were positive only for HPV16.HPV16 was the most commonly detected genotype in PeCa. HPV 31, 35 and 39 were infrequent; however, they were related to a single infection and P16INK4A overexpression; thus, they seem to be relevant in PeCa carcinogenesis. Our results suggest that P16INK4A overexpression could be useful for the classification of HPV-related PeCa. The role of multiple HPV genotypes in the development and prognosis of PeCa is still not completely understood. Thus, it is necessary to define criteria to establish reliable ways to classify HPV-related PeCa that could lead to optimal therapeutic approaches. Penile carcinoma (PeCa) is a globally rare neoplasia that mainly affects men between 50 and 70 years of age. The incidence reported in 2012 was 26,000 new cases worldwide . In our work, we identified the following genotypes in descending order: 16, 52, 18, 45, 31 and 11. The distribution of high-risk genotypes different from HPV16 varies with geography and anatomical site . AccordiThe presence of multiple HPV genotypes has been explored mainly in cervical cancer and its precursor lesions , 26–28. Regarding the patient’s age, we found that 57% of patients under 45 years were HPV positive, and all of them had a single HPV16 infection. In contrast, 76% of older patients (> 76 years) were HPV positive, and only one half of them presented a single genotype (HPV16 or HPV35). These data suggest a relationship between genotype and age, possibly due to fast progression caused by a variety of events such as chromosomic instability and viral integration associated with the oncogenic capacities of E6 and E7 –36. AlthCDNK2A, as well as loss of heterozygosity [Although P16INK4A has been defined as a surrogate marker for high-risk HPV-induced transformation, differences have been observed in the concordance between HPV presence and P16INK4A expression , 38. Somzygosity . Therefozygosity .In our study, HPV was detected in almost 60% of PeCa cases, although three quarters were classified as non-HPV-related tumors. HPV16 was found to be the most frequent genotype present, followed by HPV52, 33, 35, and 39. Most of these could be prevented by nonavalent HPV vaccination. The role of multiple HPV genotypes in PeCa is of interest. The etiological attribution to HPV in PeCa may be stratified with more certainty by using biomarkers such as viral oncoprotein mRNA and surrogate markers such as P16INK4A.


INK4a was observed in 93.3% of cases and was significantly correlated with high-risk HPV infections. Furthermore, p53 and NRAS mutations were detected in five (16.7%) and one (3.3%) cases, respectively, whereas no EGFR, KRAS or BRAF mutations were observed. These results demonstrate that high-risk HPV infection may be an important oncogenic factor for the development and progression of cervical neuroendocrine carcinoma.Neuroendocrine cervical cancer is a rare subtype of cervical cancer with a highly aggressive malignancy. This study was conducted to analyse the human papillomavirus (HPV) infection and molecular abnormalities in Vietnamese neuroendocrine carcinomas of the uterine cervix. HPV genotyping and p53 mutations were examined using polymerase chain reaction (PCR)-based direct sequencing. Mutations of epidermal growth factor receptor (EGFR), Kirsten rat sarcoma (KRAS), neuroblastoma RAS viral oncogene homolog (NRAS) and v-Raf murine sarcoma viral oncogene homolog B (BRAF) were identified using commercial kits. Four high-risk HPV genotypes were identified in 26 (86.7%) out of a total of 30 tumours. The prevalence of HPV 16, 18, 31 and 45 was 20.0%, 50.0%, 20.0% and 36.7%, respectively. Overexpression of p16 Cervical cancer is the second most common type of cancer and the fourth leading cause of cancer mortality in women worldwide, with an estimated 530,000 cases per year and a disproportionate impact on developing countries . CervicaHuman papillomavirus (HPV) infection plays a critical role in the pathogenesis of cervical cancer. HPV can be categorised into high-risk and low-risk HPV types based on each type’s correlation with cervical cancer and precursor lesions. High-risk HPV genotypes—such as HPV 16, 18, 31, 45, 56 and 68—are detected in 99% of cervical cancers and approximately 70% of cervical cancers are related to the presence of HPV 16 and 18  neuroendocrine cervical tumour tissue blocks were collected from Nov 2016 to May 2018 from the patients who were diagnosed and underwent surgical resection at National Cancer Hospital K in Vietnam. All hematoxylin and eosin (H&E)-stained slides were evaluated by pathologists to confirm the histological diagnosis of neuroendocrine cervical cancer. The clinical characteristics of the patients were obtained from surgical and pathological records. Informed consent was obtained from the patients via a written form and the study was approved by the guidelines of a local ethics committee in Vietnam.INK4a was performed using the BenchMark XT system . The pattern of p16INK4a positivity was scored on a four-tier scale from 0 to 3+ as follows: 0 , 1+ (blush staining in fewer than 10% of tumour cells), 2+ (medium staining in 10%–50% of tumour cells) and 3+ (strong diffuse nuclear and cytoplasmic staining in more than 50% of tumour cells)  . Tumoursβ-globin gene.DNA was isolated using the QIAamp DNA FFPE Tissue Kit . The quality of the DNA samples was assessed via polymerase chain reaction (PCR) targeting the The high-risk HPV strains 16, 18, 31 and 45 were detected using PCR with specific primers for each genotype. PCR products were sequenced using the Bigdye Terminator Kit  to exactly confirm each HPV genotype.Mutations in p53 were examined from exon 5 to exon 8 by directly sequencing PCR products using the Bigdye Terminator Kit . Epidermal growth factor receptor (EGFR) mutations were detected using the Therascreen EGFR RGQ PCR kit . v-Raf murine sarcoma viral oncogene homolog B (BRAF) mutations were detected using the Cobas 4800 BRAF V600 mutation test , whereas KRAS mutation were examined using the Cobas KRAS mutation test . NRAS mutations were detected using the Therascreen NRAS Pyro Kit 24, V1 , based on the pyrosequencing technology of the PyroMark Q24 system.P < 0.05.The statistical analysis was carried out using SPSS software . The potential correlations were analysed using Fisher’s exact test. The statistical significance for all analyses was set at P = 0.037), suggesting HPV infection was more commonly found in neuroendocrine cervical cancer with advanced stage disease than in those with early stage disease.The HPV DNA of four high-risk genotypes were identified in 26 (86.7%) tumours, of which 17 (56.7%) cases were only infected with one HPV type (single infection), six (20.0%) were co-infected with two HPV types and three 10.0%) were co-infected with three HPV types. HPV 18 was the most common genotype , followed by HPV 45 , HPV 16  and HPV 31  . Accumul.0% were P = 0.002), but there was no correlation between HPV infections and p53 or NRAS mutations. In the future, the precise mechanisms of these relationships should be elucidated.The features of HPV infection in cervical cancer, including cervical neuroendocrine carcinoma, have become increasingly clear, whereas the characteristics of the mutations remain inconclusive. Genetic abnormalities of p53 and NRAS were detected in five (16.7%) and one (3.3%) tumours, respectively, whereas serval mutations—such as EGFR, KRAS, and BRAF—that often occur at a high frequency in many cancers were not found in this study. Out of a total of five p53 mutations identified from exon 5 to exon 8, two of the mutations were found in exon 8 and only one mutation each existed on exons 5, 6 and 7. Five point mutations in the p53 sequence resulted in a single amino acid substitution at codons 140 (Thr-Ile) of exon 5, 205 (Tyr-Cys) of exon 6, 237 (Met-Lys) of exon 7, and 278 (Pro-Ala) and 300 (Pro-Thr) of exon 8. p53 is one of the most frequently mutated tumour suppressor genes observed in many types of human cancer. However, the mutation of the p53 gene is a rare event in carcinomas of the cervix, only accounting for approximately 5% of all cases , 19. In INK4a overexpression. Notably, the p16INK4a IHC results showed a significant association with the presence of HPV (P = 0.014)  cases were found to be positive for p16= 0.014) , whereasmutation . This daINK4a expression. We identified a low prevalence of p53 and NRAS mutations that do not appear to be associated with any specific HPV genotype, whereas the NRAS mutation correlates with the p53 mutation. However, further studies with larger sample sizes should be conducted to confirm the frequency and distribution of genetic alterations, as well as their relationship with HPV infection.In conclusion, our findings indicate that cervical neuroendocrine carcinomas are universally correlated with HPV infection, especially high-risk types, and a corresponding high frequency of p16


In a transition period of prevention strategy against HPV infection and cervical cancer in Saudi Arabia, it becomes necessary to appraise physicians’ preparedness to undertake the inherent actions and responsibilities, by evaluating their knowledge and opinions regarding HPV infection and vaccine.A cross-sectional study carried out between Jan 2017 and Nov 2018, included 2000 physicians working in 21 public centers from the five regions of Saudi Arabia. A self-administered questionnaire was used to assess physicians’ perception about HPV infection prevalence (1 item), knowledge about HPV infection and vaccine (9 items), and opinions and attitudes toward vaccine (4 items). A knowledge score (range 0–9) was calculated and adequate knowledge was assumed for a score ≥ median. Factors associated with opinions and attitudes were explored and multivariate regression was used to analyze independent factors of inadequate knowledge (score < median).p = 0.003), practice level , and non Ob-Gyne specialty ; in addition to disparities across region and age. Among the participants, 7.6% were immunized and 41.2% accepted to receive the vaccine, while majority were favorable to have their children vaccinated (77.6%) and to include HPV vaccine in the local immunization program (69.6%). Self-perceived underexposure to HPV infection (58.5%), lack of knowledge about the vaccine (21.1%) and being sexually inactive (14.7%) were the most frequently reported reasons for refusing the vaccine. Overall negative attitude regarding vaccine was typically associated with male, older, Saudi, senior consultant in other than Ob/Gyn specialty. Inadequate knowledge level as well as lowly perceived prevalence of HPV infection were associated with less favorable attitude to vaccination.Majority of the participants replied correctly to all knowledge questions, and 63.0% perceived HPV infection as a frequently encountered infection. Median knowledge score was 8 and 62.0% had adequate knowledge (score ≥ 8). Inadequate knowledge was independently associated with Saudi nationality (OR = 1.51, More specific educational interventions are warranted to trigger physicians’ active engagement in the fight against HPV infection and cervical cancer. Such interventions should demystify the HPV vaccine by exposing its efficacy, availability and safety, along with providing practical information about the vaccination procedure and goals to achieve successful prevention strategy. Human papilloma viruses (HPVs) are considered common pathogens that are disseminated through sexual intercourse. Several genotypes of HPV were identified, which are classified into low- or high-risk depending on their oncogenic potential on the infected cells. Low-risk genotypes are generally associated with anogenital warts, while high-risk ones are associated with malignant tumors of the genital tract, notably cervical cancers and, to a lesser extent, endometrial and ovarian cancers –4.Global observations suggest that 75% of sexually active adults would contract an HPV infection during their lifetime . ConsequThe major threat of HPV infections remains the cervical cancer among infected women. Cervix cancers ranked fifth most fatal and fourth most common cancers worldwide by the early 2010’s . They ocTherefore, the globally adopted strategy in cervical cancer prevention includes systematic HPV cervical screenings and vaccination against HPV. Majority of the developed countries recommended preventive vaccination against HPV as a routine vaccine for 11–12 year-old population. As well, the vaccine was recommended for unvaccinated females and males who were between 13 and 26 and 13–21, respectively , 17.However, the implementation of such strategies is slow to appear in several developing countries, which is thought to explain 85% of the global burden of mortality from cervical cancer today , 18–20. On the other hand, raising awareness among the general population as well as among specific high-risk subpopulations is considered the most effective method to prevent HPV infection. This requires active involvement by the physicians, and specifically the obstetric gynecology, family medicine and primary health care physicians, to promote the vaccine and other preventive measures among the community , 22. HowThis cross-sectional study involved physicians from all regions of the Kingdom of Saudi Arabia, and was conducted between January 2017 and June 2018. Eligibility criteria applied for both gender, all nationality residents and junior and senior physicians, who were qualified by the Saudi Commission for Health Specialties and currently working at a governmental hospital or clinic in Saudi Arabia. Physicians who were employed as part time or locum, as well as those who were on leave at the time of data collection were excluded. The study was approved by the Institutional Review Board of the Saudi Ministry of National Guard Health Affairs (MNG-HA) and the committee board of the Saudi Ministry of Health (MOH).P = 50%) of physicians with adequate knowledge, with ±0.03 precision, 80% statistical power and 95% confidence level, among a target population of 10,641 physicians working in Saudi Arabia as estimated in 2017 [2 * P(1 - P))/e2.Sample size was calculated to detect an unknown percentage ,P is expected true proportion = 0.5, as true proportion unknown,e is desired precision = 0.03 .where:n = 971, which was doubled (n = 1942) and up rounded to n = 2000 to enable more powerful subgroup analysis.The sample size was calculated as n = 2000).A stratified clustered sampling was used to randomly select a number of clinics (clusters) from each of the five regions (strata) of the Kingdom. The number of clinics by region was proportional to the number of big cities and to the total number of clinics as follows: Eastern , Western , Northern , Southern  and Central region . Thus, a total 21 clinics were selected, expecting approximately 100 participations from each clinic to fulfil the target sample size  and Shapiro-Wilk statistics  indicated non normal distribution of the variable. Consequently, factors associated with knowledge were analyzed using two methods: 1) nonparametric tests  to compare raw knowledge scores between different factor’s categories; 2) chi-squared test to compare the percentage of participants with adequate knowledge level, defined as knowledge score ≥ median value, between the factor’s categories. Results are presented as mean, median and range for nonparametric tests and frequency and percentage of individuals for chi-squared test. A multivariate binomial regression model was carried out to analyze independent factors of inadequate knowledge, defined as knowledge score < median value. Further, chi-square test was used to analyze the association of opinions and attitudes regarding HPV vaccination with demographic factors as well as with knowledge level. A p value of < 0.05 was considered to reject the null hypothesis.Statistical analysis was performed with the Statistical Package for Social Sciences version 21.0 for Windows . Descriptive statistics were used to summarize participant’s charactereistics and patterns of answers to knowledge and opinion and attitude items. A knowledge score (range = 0–9) was calculated as the sum of correct answers given by the participant regarding the 9 knowledge questions. Analysis of knowledge score distribution using Kolmogorov-Smirnov statistics . Participants’ demographic characteristics showed that more than half were females (52.8%), two-third were Saudi (67.0%), and more than three-quarter were aged 20–50 years  and married (79.4%). Professional characteristics showed majority of senior (50.2%) and Obstetrics-Gynecology (58.6%) consultants. Distribution by region showed highest percentage of Western region (57.4%), followed by Eastern and Central regions with 15.4% each , sexual transmission mode (95.6%), and whether HPV infection may be asymptomatic (95.0%) totalized the highest correctness rates. The lowest correctness rate was observed for the item whether HPV vaccines protect against all HOP serotypes (67.0%). Further, 63.0% of the participants perceived HPV infection as a frequently encountered infection Fig. . RegardiBoth nonparametric tests analyzing knowledge score and chi-square test analyzing the percentage of adequate knowledge level showed statistically significant differences in all demographic and professional factors Table . These sp = 0.003), practice level , specialty , and region ; as well as age  was independently associated with the physicians’ nationality  included the following: not being under the risk of HPV infection (58.5%); lack of knowledge about the vaccine (21.1%); being sexually inactive (14.7%); vaccine not being reimbursed by the government (8.7%); and fear from vaccine side effects (8.4%); while 10.0% had no reason for refusing  and less than half opined that they agree to be vaccinated 41.2%); however, majority were favorable to vaccinate their children or own children (77.6%) and to include HPV vaccine in the local immunization program (69.6%). Reasons for HPV vaccine refusal among participants who disagreed to be or to have their children vaccinated , single (10.4%) followed by married (7.6%) ones (p = 0.044), those from Ob/Gyn specialty  and those from the Western regions  compared to their counterparts. Additionally, those who perceived HPV infection as being frequent had higher rate of immunization (8.6% versus 5.9%) compared to their counterparts (p = 0.032). On the other hand, no association with knowledge level was found (p = 0.317). Other opinion and attitude parameters showed statistically significant associations with all or almost all demographic factors. Most notably, overall negative attitude regarding HPV vaccination was typically associated with male, older, Saudi, senior consultant in other than Ob/Gyn specialty. Additionally, participants with inadequate knowledge level were less favorable to vaccinate their children  and to inclusion of the vaccine in the local immunization program . Interestingly, participants who perceived HPV infection as being prevalent were remarkably more favorable to vaccinate their children  and to inclusion of HPV vaccine in the Saudi immunization program  (Table The rate of HPV immunization was higher among younger 12.5%) followed by older (10.7%) physicians , at the time of the survey, or favorable to receive the vaccine (41.2%), which contrasted with the majority being favorable to vaccinate their children (77.6%) and to include the vaccine in the local immunization program (69.6%). The combination of these observations suggest that physicians are partially aware about the imminent threat of HPV infection on the Saudi population, expecting a shift to an endemic trend of the disease. This emerging awareness was more perceptible among young and female physicians, as well as among non-Saudi and particularly among Ob/Gyn specialists. This issues the hypothesis of an increasing number of cases that are diagnosed in the clinical practice, by these categories of physicians. Further, findings of this study enabled profiling the typical physicians with certain reluctance to systematic vaccination against HPV.A few studies have assessed the level of physicians’ knowledge about HPV infections and vaccines in conservative or Middle Eastern communities as Saudi Arabia. A recent study 2018) among Saudi primary health care physicians showed high knowledge scores about both HPV infection and vaccine, along with positive attitude regarding the necessity of the vaccine. However, only 16.5% declared routinely providing vaccine recommendation to their patients; and this was associated with higher knowledge score and better perception about the necessity of the vaccine 8 among S. RegionaComparable observations were reported in international literature. For example, a Mexican nationwide study demonstrated high levels of awareness among physicians about HPV-cervix cancer causal relationship with significantly better knowledge among Ob-Gyn specialists than GPs, notably regarding the pathogenic mechanisms of this relationship. The same study demonstrated favorable opinions among the physicians regarding the pertinence of educating women about the causal relationship of cervical cancer with HPV; however, a significant proportion opined that such information might cause disruptions between partners, confusions or unnecessary anxiety among the population . In IndiStudies that were carried out among other healthcare providers showed comparable results. An Indian study involving healthcare providers from 232 hospitals and 80 PHCs demonstrated that less than half respondents recommended HPV vaccine to their patients although majority had adequate knowledge about the causal relationship with cervical cancer and were aware about the importance of the vaccine . In TurkRegardless of using different tools and methods to assess knowledge and opinions, conclusions from the previous studies converge towards two major issues: 1) a generally poor engagement among physicians and health providers in HPV vaccination among their patients despite a good theoretical knowledge; 2) a positive effect of knowledge on such engagement. This indicates that increasing knowledge and awareness among the physicians about HPV infection and vaccination is crucial to enhance a positive attitude, but not sufficient to obtain active and efficient engagement in the clinical practice. Further, the combination of the previously cited data suggests that the efficacy of the vaccine in preventing cervical cancer may be a powerful argument to convince physicians to promote the vaccine among their patients and the society.p < 0.001) or to have their children vaccinated  had significantly higher propensity to refuse including HPV vaccine in the Saudi immunization program (Results not presented). Consequently, the presence of such barriers among physicians constitute not only barriers to accept the vaccine at the individual level, but indicate a general adverse attitude towards HPV vaccine that could downplay the physicians’ role in promoting the vaccine among patients. Therefore, identifying and alleviating these obstacles is the second most important step in physician’s education in HPV vaccination.The other determining factor of physicians’ engagement in promoting HPV vaccine among their patients may be the presence of eventual barriers that stand between the theoretical knowledge and practice. In the present study, participants raised a number of reasons to explain their refusal to receive the vaccine or to have their children vaccinated. These included lowly perceived risk for HPV infection, lack of knowledge about the vaccine and concerns about the vaccine safety and cost. Similar to our findings, previously cited Indian study among medical and paramedical students showed that almost half of the respondents were reluctant to receive the vaccine, presenting concerns about side effects and cost of the vaccine or declaring being underexposed to the risk of contagion . LikewisFurther, the issue of vaccine cost and its reimbursement by the government appear to be of significant weight on doctors’ attitude in the routine practice. A study from Hong Kong, showed that two-third primary health care physicians involved in vaccination program for schoolgirls were favorable to HPV vaccine becoming fully paid by the government, stressing that this would considerably enhance the acceptance of the vaccine and the coverage rate among the population .The present study highlighted male, older, Saudi, and senior consultants in specialties other than Ob/Gyn as the typical physician profile associated with adverse opinions regarding HPV vaccination and a presumably lower propensity to take part in vaccination campaigns and to promote HPV vaccination among patients and the population. Similar observations were reported locally, where better attitude regarding the vaccine was evidenced among female and younger PHC physicians, while male and older ones had comparably poorer perception of the importance of HPV vaccine . Such agThe major strengths of this study was its sampling methodology including large sample size and physicians’ recruitment from the five regions of the country. However, it had some methodological issues that may limit the generalization of its findings. First, the cross-sectional design does not enable establishing a causal relationship between physicians’ levels of knowledge and opinions and attitude regarding the vaccine and vaccination program. Second, only physicians working in public sector were included, while those from private sector or academic filed were not included, which may have a significant impact on knowledge level as well as in vaccination and vaccine recommendation practice. Third, the questionnaire included basic questions and did not probe into advanced issues, which may result in overestimation of the knowledge level. Additionally, the questionnaire failed to explore the physicians’ practice in recommending the vaccine to their patients.Despite these limitations, findings from this study contribute in characterizing the unmet needs in physicians’ education regarding HPV infection and vaccination and shed light on some barriers and misconceptions that may hinder the physicians’ engagement in promoting the vaccine. It highlights efficient and targeted educational interventions among physicians as being the next crucial step for a successful prevention strategy against HPV infections and cervical cancer. On the other hand, the Saudi government and health authorities should tackle the ambient challenges including the societal and cultural barriers by means of effective communication strategies to break the apprehensiveness wall between physicians and patients, as well as a minimum subsidy to overcome vaccine cost among the most eligible groups of the population.Majority of physicians from the Saudi public sector have adequate basic knowledge about HPV infection and vaccination, which was associated with more positive attitude regarding vaccine. A generational shift in opinions and attitude regarding HPV vaccine was observed, denoting an emerging awareness about the HPV threat to the society among younger physicians.On the other hand, lowly perceived risk for HPV infection, lack of knowledge about the vaccine and concerns about the vaccine safety and cost were frequently reported as reasons to refuse the vaccine by the physicians and were indicative of a general adverse attitude towards HPV vaccine that could downplay the physicians’ role in promoting the vaccine among patients.Therefore, more specific educational interventions are warranted to trigger an active engagement among physicians in the fight against HPV infection and cervical cancer. Such interventions should demystify the HPV vaccine by exposing its efficacy, availability and safety, along with providing practical information about the vaccination procedure, local policies and goals to achieve via the vaccination and screening campaigns. Additionally, exposing an accurate epidemiological picture that highlights the imminent threat of the HPV infection to the Saudi society would raise greater awareness about the necessity of the vaccine.Finally, the present study highlighted male, older, Saudi, and senior consultants in specialties other than Ob/Gyn as the typical physician profile associated with adverse opinions regarding HPV vaccination and a presumably lower propensity to take part in vaccination campaigns and to promote HPV vaccination among patients and the population. Therefore, physicians with such “adverse profiles” should be the target of more intensive educational interventions, which should aim at fixing the misconceptions about the HPV vaccine and updating the physician’s roles in a changing society that is increasingly exposed to sexually transmitted diseases.Additional file 1. Physician’ Knowledge and Opinions on Human Papillomavirus Vaccination: A Cross-Sectional Study from Saudi Arabia.


Human papillomavirus (HPV) is the main cause of genital warts and some anogenital cancers in male and female subjects which is commonly transmitted by sexual contacts. The objective of this cross-sectional study was to examine the prevalence of HPV genotypes in 10,266 Iranian male and female population, according to their age. Samples were collected from the penile and anal sites of male subjects and the vagina and cervix of female subjects in a time period between 2011 and 2016. HPV DNA was detected in PCR using the MY09 and MY11 primers, and the INNO-LiPA assay was applied for HPV genotyping. To investigate the relevance of HPV infection and age, the samples were classified into 4 age groups .P=1).Totally, the most common low risk HPV genotypes detected in the studied male and female subjects were HPV-6 (77.7% and 43.3%) and HPV-11 (13.7% and 11.4%), and more frequent high risk HPV genotypes were HPV-16 (5.5% and 16.6%) and HPV-52 (3.2% and 9.6%), respectively. High burden of the HPV infection was observed at ranges of 30 and 44 years (51.8%) with a peak at ranges between 30 and 32 years. No considerable statistically significant correlation was found between HPV infection and age (This study gave an epidemiological overview of circulating HPV genotypes in Iranian population to develop future vaccination policies, though the findings of prevalent HPV genotypes in female subjects were inconsistent with the previous studies reported in Iran. Human papillomavirus (HPV) genotypes have been detected so far. Adequate documents show that there is a relationship between HPV and some human cancers such as cervix cancer, head and neck carcinoma, penile cancer, oropharyngeal squamous cell carcinomas and, anal cancers  females and 1,915 (18.7%) males from different provinces of Iran who were referred to Keyvan Laboratory of Tehran for HPV DNA testing and genotyping between April 2011 and April 2016 were enrolled. Almost all subjects were sexually active with an age range from 13 to 74 years old. The HPV DNA testing and genotyping were ordered by clinicians. Male samples were collected from anal and penile biopsies and female samples from the vagina and cervix through the use of either Dacron swabs or brush. For DNA extraction, QIAamp DNA Mini kit  was utilized in accordance with the manufacturer’s directions and the concentration of extracted DNA was evaluated by spectrophotometry in 260 nm. Genomic DNA of the samples was used in PCR using the MY09 and MY11 primers. DNA quality in samples was verified using beta-globin PCR assay.Detection of HPV DNA and genotypes were carried out using INNO-LiPA assay . The INNO-LiPA HPV genotyping is based on the principle of reverse hybridization. Briefly, a part of L1 region called SPF10 of HPV genome was extended by specific primers; then, the biotinylated amplimers were denatured and hybridized with specific oligonucleotide probes. Furthermore, a primer pair of the human HLA-DPB1 gene was added to confirm the quality and extraction of DNA. Next, streptavidin-conjugated alkaline phosphatase was added. After incubation with BCIP/NBT chromogen, the results were interpreted visually.Statistical analysisIBM SPSS statistics version 22  was usedINNO LiPA HPV genotyping determined 32 HPV genotypes; HPV-16, -18, -31, -33, -35, -39, -45, -51, -52, -56, -58, -59, and -68 as high risk HPV genotypes (HR HPV), HPV-26, -53, -66, -70, -73, and -82 as possible high risk HPV genotypes (pHR HPV), and HPV-6, -11, -40, -42, -43, -44, -54, -61, -62, -67, -81, -83, and -89 as low risk HPV genotypes (LR HPV). This HPV classification was based on IARC Monographs (12). During the study period, the total HPV prevalence was 49.5%; hence, among 10,266 participants, 49.5% (n=5085) were HPV DNA positive and 50.5% (n=5181) were negative. Totally, among 5,085 samples from 31 Iranian provinces, four most common HPV genotypes were HPV-6, -11, -16, and -52 in male subjects and HPV-6, -16, -11, and -52/-53  in female subjects, in a descending order. Among HR and pHR HPV genotypes, after HPV-52/-53 (9.6%), HR HPV-51 (8.0%) had the highest frequency in female subjects and also other genotypes with prevalence less than 8% included pHR HPV-66 (7.7%) and HR HPV-31 (7.0%). In male subjects, HR HPV-51 (2.8%) was the third most common HR HPV genotypes. The common HPV genotypes was observed in both single and mixed HPV infections and the genotypes which had high rates in the single HPV infection were also frequent in the mixed infections  and 2. TOverall, the mixed HPV infections were found in 14.0% (n=174) of HPV DNA positive males, while 42.2% (n=1620) of HPV infection in females were caused by mixed HPV genotypes. Concomitant infection with at least two LR HPV genotypes was detected in 3.3% (168/5085) of all positive cases. In both sexes with mixed HPV genotypes, concurrent infection by at least one HR and one LR HPV genotype was commonly observed than other forms of mixed infections . Only 0.P=1); however, 90.6% and 86.4% of the HPV infections in female and male subjects were seen in subjects aged 13 to 44 years, respectively. The highest rate of HPV infection occurred in positive subjects aged 30-44 years . In male subjects, the subjects aged 30-44 years had the first and second most frequent HR HPV and LR HPV genotypes: HR HPV-16, -51/-52 , and LR HPV-6, -11. In the participants >60 years of age, HPV-6 and -11 were the most detectable HPV genotypes (data not shown). th or higher in our study. Vaccinated female may reduce the HPV infections in male subjects, due to herd immunity (th most common cancer among Iranian female (HPV distribution varies in different countries and populations and even though in various parts of a country. For example, HPV-16 in Zhejiang province, HPV-52 in Guiyang and Guangdong provinces, and HPV-6 in Xi'an province of China are predominant genotypes , 15. In immunity ; hence, immunity . Accordin female . On the n female  has genen female  play a kn female . D is thn female . TherefoMen with HPV infection only accounted for 12.1% of all HPV DNA cases. Among this, 86.0% had a single infection. The predominant three HPV genotypes found were in complete agreement with those were reported by Salehi-Vaziri in 2015 . Only alThe age distribution of HPV infection in both sexes showed a mono summit graph with a peak age of 30-32 years. There was the highest incidence in the age of marriage and after beginning of the sexual activity due to some cultural habits, religious beliefs such as not to have illegitimate sex, increasing the age of marriage, and absence of immune protection against HPV in the lack of national vaccination program. Also, the rate of mixed infections is increased with the rise in the frequency of sexual contact and high-risk sexual behaviour . The steAn important limitation of this study was the lack of cytology information of positive samples for correlation with the results of HPV genotypes. Another limitation, this rate of HPV prevalence may not reflect the distribution of HPV genotypes in the country since this study recruited only the samples with normal pap-smear, cell collections from vagina, and cervix in female subjects, and bulge of penile and anal sites in male subjects. Hence, similar studies need to be conducted on cancerous samples. No demographic data of participants such as marital status, job, and the number of sex partners were available. In spite of the male subjects’ limitations, this study gives an insight into the present status of the distribution of HPV genotypes in the target population and is helpful for setting policies.


Chlamydia trachomatis (CT) is considered a potential cofactor in the development of cervical intraepithelial neoplasia (CIN), although different studies have produced contradicting information . The objective of this cross-sectional study was to determine the prevalence and association of HPV-Chlamydial coinfection with cervical intraepithelial lesions and other risk factors for cervical intraepithelial lesions at a hospital in south western Uganda (MRRH). Human Pappilloma Virus (HPV) is the necessary cause of cervical cancer. A number of risk factors are believed to influence the role of HPV in the development of cervical cancer. This is so because majority of HPV infections are cleared and only a few are able to result into cancer. Chlamydia trachomatis antigen testing. Data was entered in Microsoft excel and analysed using STATA 12 software. With the help of spearman's correlation at the 0.05 level of significance, bivariate and multivariate analysis were done by logistic regression, to determine associations of risk factors to cervical lesions. The study included 93 participants, with an age range of 25 to 80 years, from whom cervical specimens were collected and enrolment forms were completed upon consent. Experienced midwives collected one cervical smear and two endocervical swabs from each participant. The swabs were used for HPV DNA and The results showed the prevalence of HPV-Chlamydial coinfection to be 8.6% (8/93). Positive Pap smear results were found in 60.22% (56/93) participants, most of whom had low grade squamous intraepitherial lesion (LSIL) (54.84%). HPV-Chlamydial coinfection showed a significant correlation with a positive cytology result and only relatively significantly correlated with LSIL grade of cytological positivity. HPV was found to be the risk factors associated with cervical intraepithelial lesions at MRRH. HPV, Chlamydia, and HPV-Chlamydial coinfection are prevalent infections and there is a likelihood of association between HPV-Chlamydial coinfection and with cervical intraepithelial lesions. This study recommends general sexually transimitted infections (STIS) screening for every woman that turns up for cervical cancer screening and a larger study, probably a multicentre study. Cervical cancer is the fourth most common cancer in women , and theThe World Health Organisation (WHO) reported an estimate of 266,000 deaths from cervical cancer worldwide in 2012, accounting for 7.5% of all female cancer deaths. It is further emphasized that almost nine out of ten (87%) cervical cancer deaths occur in the less developed regions. Mortality varies 18-fold between the different regions of the world, with rates ranging from less than 2 per 100,000 in Western Asia, Western Europe, and Australia/New Zealand to more than 20 per 100,000 in Melanesia (20.6), Middle (22.2), and Eastern (27.6) Africa [The transformational changes start from a normal cell, turning it into a tumour cell in a multistage process, typically a progression from a precancerous lesion to malignant tumours. Because precancerous lesions can be detected by screening, vigorous screening programs have been put in place all over the world and have yielded results of reduced mortality in developed countries and a few developing countries , 4. In U et al., 2016). With increasing prevalence of HIV, there is a projected increase in risk to STIs in the developing world . Research studies conducted by Kakaire et al. (2015) showed a prevalence of STIs among women living with HIV to be 11.1% (95% CI 7.8-14.4) and individual prevalence for Trichomonas vaginalis, Neisseria gonorrhoea, and Chlamydia trachomatis being 5.9%, 5.4l%, and 0.9%, respectively.A number of sexually transmitted infections (STIs) have been linked to cervical cancer or precancerous lesions of the cervix. Some of these include HPV, HIV, and Chlamydia , while more than 290 million women have an HPV infection . In view of this, Nwankwo and Magaji (2014) studied the prevalence of Chlamydia trachomatis in Nigeria and found out that the prevalence was 9.6% in Kano. They further noted the fact that a very high percentage of the patients, 95.2%, were completely ignorant of the existence of Chlamydia infection and consequently did not know of its associated complications.Chlamydia trachomatis (CT) is considered a potential cofactor in the development of cervical intraepithelial neoplasia (CIN) although different studies have produced contradicting information [ Chlamydia trachomatis play a central role in the etiology of cervical intraepithelial neoplasia and subsequent cancer. It has also been noted that multiple non-HPV infections correlate significantly with low grade intraepithelial lesions . There are correlations not only between STIs and hrHPV infection but also between a positive cervical cytology and STIs .ormation –9. It isSo, this study aimed at studying the relationship between Chlamydial infection, HPV infection, and HPV-Chlamydial coinfection and cervical intraepithelial lesions at MRRH.The study was carried out in the cervical cancer clinic and the pathology department of Mbarara Regional Referral Hospital (MRRH) and Mbarara University of Science and Technology (MUST), respectively. MUST and MRRH are located in Mbarara district, south western Uganda. It is approximately 260 km from Kampala, the capital city of Uganda and 2 km from the town centre on the Mbarara-Kabale highway. MUST and MRRH are public institutions and the university has over 3000 students while the hospital is the regional referral hospital of south wersten Uganda with a capacity of 1200 beds. The cervical cancer clinic together with MUST clinical and research laboratory were chosen because they receive clients from the entire western region of Uganda; hence the sample size was representative of a big number of Ugandans. (Ref: MUREC 1/7).The principal investigator sought approval from the Department of Medical Laboratory Science, the faculty of medicine research committee and the Mbarara University Institutional Research and Ethics CommitteeClearance was also sought from the Department of Obstetrics and Gynecology before commencement of the study.Respondents were given verbal explanation about the study and were given a consent form, well translated into Runyankole-Rukiga,  where they signed, having known the relevance of the study.Confidentiality was highly observed as respondents were identified with study numbers and not names.The participants were recruited from the cervical cancer clinic in the months of June and July 2017. The study covered a total of 105 participants from whom samples were collected and then analysed. The study included all patients that presented at the cervical cancer clinic and consented to participate in the study. However all moribund patients and those that presented at the cervical cancer clinic during their menstrual periods were not considered for the study.The study employed an enrolment form to collect qualitative data and laboratory methods. Three cervical specimens  were collected from every participant and labelled with unique study numbers. Chlamydia trachomatis antigen testing and HPV DNA testing  was used for qualitative detection of Chlamydia trachomatis DNA in cervical specimens  using the Amplisens HPV HCR genotype-Eph PCR kit. This Amplisens HPV HCR genotype-Eph PCR kit utilizes consensus primers, directed to a conserved L1 gene during amplification for hr HPV genotypes . In this multiplex PCR, the Data from 93 participants was entered in Microsoft excel and analysed using STATA 12 software, with the help of spearman's correlation at the 0.05 level of significance. Bivariate and multivariate analysis were done by logistic regression, to determine associations of risk factors to cervical lesions.The study enrolled a total of 105 participants, of whom 12 were excluded from the study, because of unsatisfactory Pap smears. Eventually 93 participants were included in the data analysis. The mean age of participants was 41.92 years (minimum 26 and maximum 80), 55.91% of participants falling within the 25-40-year age bracket . Chlamydia trachomatis infection was found in 8.6% (8/93) participants. And the HPV-Chlamydial coinfection was also in 8.6% (8/93) (HPV infection was found in 63.4% (59/93) of participants, whereas% (8/93) .For those women who were coinfected with HPV and Chlamydia, the mean age was 36 years of age, with a minimum of 29 and maximum of 52 years of age. Contraceptive use was reported in 2 out of 8 women (25%) whereas 25% of them reported to have a history of a positive Pap test. Smoking was reported in 1 out of 8 women (12.5%). HIV tests were negative in all the eight women who showed HPV-Chlamydial coinfection.Bacterial vaginosis was reported in 88 out of 93 participants (94.6%). However, all women, 8 out of 8 (100%) who showed coinfection with HPV and Chlamydia, were positive for bacterial vaginosis. Chlamydia trachomatis antigen test result, all of them had a positive Pap result  and significantly correlated only with LSIL grade of cytological lesion . ASCUS and HSIL had no significant correlation with HPV-Chlamydial coinfection  (The eight women who had a positivep result . On furtctively) . P value <0.001), but specifically with LSIL  unlike other grades of lesions.59 women tested positive for HPV DNA, and of them, 47 had LSIL, 2 had HSIL, and 1 had ASCUS, while 9 were negative for intraepithelial lesion or malignancy. Generally, HPV DNA positivity was significantly associated with Pap smear positive findings . This could not be assessed on logistic regression because there were only 8 observations, which were below the required minimum (10). Chlamydia trachomatis infection, and HPV-Chlamydial coinfection.Eight (8) risk factors were assessed for any association with cervical intraepithelial lesions. These included age, smoking, contraceptive use, HIV status, family history of cervical cancer, HPV positivity,HIV infection, Chlamydia and HPV-Chlamydial coinfection had very few observations and therefore had insufficient data for logistic regression. P value <0.001)   . et al. [ et al. [ et al. [ et al. [The prevalence of HPV-Chlamydial coinfection in this study was 8.6%, way above the reported rates of 0.7% as reported by Bhatla et al. . Bhatla  et al.  also rep[ et al.  and de P[ et al. , respect[ et al.  reported et al. [This study found a significant association between HPV-Chlamydial coinfection and the cytological diagnosis of LSIL , unlike de Castro-Sobrinho et al.  who repo et al. [Studies by da Silva Barros et al.  confirmeThis study investigated five risk factors for an LSIL. These included age , HPV infection, Chlamydia infection, and HPV-Chlamydial coinfection. However, only HPV infection was found to be significantly associated with cervical intraepithelial lesions, specifically LSIL. This was additional information in terms of risk factors for cervical lesions. However, this was in disagreement with some authors who had earlier reported associations of some other sort. For example, many authors reported smoking, HIV, contraceptive use, and family history of cervical cancer as possible risk factors for cervical cancer. All these were found insignificantly associated with cervical intraepithelial lesions in our study. et al. [For instance, Kjellberg et al.  conducte et al. [In agreement with our findings, Bhatla et al.  conducte et al. [Just like from our study, Castellsagué et al.  concludeBased on the correlation results  there is a likelihood of an association between HPV-Chlamydial coinfection and LSIL. The only risk factor for cervical intraepithelial lesions that was confirmed was HPV infection.From this study, we recommend routine STI screening for all women of child bearing age, that turned up for cervical cancer screening. This screening is also encouraged in mass screening programs as outreaches.We also recommend a larger study, with greater power, that can gather information on more risk factors, which could have been confounders in this study.