A vaccine targeting high-risk human papillomavirus (HPV) strains can effectively prevent HPV-associated cervical cancer risk. However, many girls and women do not receive the vaccine, more often those impacted by health disparities associated with race and/or socioeconomic status. This same disparate population has also been shown to be at higher risk for cervical cancer. Many of these women also rely on the emergency department (ED) as a safety net for their healthcare. This study sought to gather information pertaining to HPV and cervical cancer risk factors, awareness of HPV and the vaccine, as well as HPV vaccine uptake in female patients presenting to an ED.We obtained 81 surveys completed by female ED patients. Demographics included age, race, income, insurance status, primary care provider status, and known cervical-cancer risk factors. Subsequent survey questions explored respondents’ knowledge, familiarity, and attitudes regarding HPV, cervical cancer, and the HPV vaccine, including vaccination uptake rates. We analyzed data using descriptive statistics and Fisher’s exact test.Approximately one in seven respondents (14.8%) had never previously heard of HPV and 32.1% were unaware of the existence of a HPV vaccine. Minority patients, including those who were Black and Hispanic patients, low income patients, and uninsured and publicly insured patients were less likely to be aware of HPV and the vaccine and likewise were less likely to be offered and receive the vaccine. More than 60% of all respondents (61.3%) had never previously been offered the vaccine, and only 24.7% of all respondents had completed the vaccine series.Female ED patients may represent an at-risk cohort with relatively low HPV awareness and low HPV vaccine uptake. The ED could represent a novel opportunity to access and engage high-risk HPV populations. Human papillomavirus (HPV) is the most common sexually transmitted infection and the most common cause of cervical cancer and cervical cancer-related deaths in the United States (US).6Most alarming, significant disparities in HPV vaccine knowledge and vaccine series completion exist among those with the highest rates of HPV infection, including racial minorities, the under- and uninsured, and the economically disadvantaged.Due to challenges accessing primary care, disparate populations may benefit from engagement in HPV awareness and screening in alternative venues.16We sought to measure and characterize HPV and cervical cancer awareness, risk factors, and HPV vaccination history among adult females presenting to an urban, academic ED. We hypothesized that female patients in the ED would demonstrate lower rates of HPV awareness and vaccination uptake than the general population.am to 4 pm (a convenience sample).We performed a cross-sectional, survey-based study at the University of Alabama at Birmingham (UAB), an urban, academic ED in the Southeast US with an affiliated emergency medicine residency program and an annual volume of approximately 65,000, from June 1–August 31, 2016. The UAB Institutional Review Board approved the protocol. Adult female patients, aged 18–32, were recruited to participate in the survey during their ED visit. We selected the age of 32 as the upper age limit for survey engagement to align with the availability of the HPV vaccine and vaccine administration guidelines beginning in 2006.We collected 81 completed surveys in this pilot study to demonstrate feasibility. Exclusion criteria included the following: individuals who did not speak English or for whom communication barriers were present ; were medically or psychiatrically unstable; or who were undergoing active emergent evaluation or treatment; and individuals presenting for sexual assault. Subjects were recruited and administered the surveys in private rooms, and written informed consent was provided. RAs verbally delivered the survey questions in a casual interview style.The 21-item survey instrument was developed by the study authors with the intent to be expansive, capturing general HPV awareness as well as HPV vaccine uptake information, in addition to demographic factors to include pre-existing cervical-cancer risk factors. While created de novo, the survey was influenced by previously published HPV awareness and attitude survey studies.What do we already know about this issue?Human papillomavirus (HPV) is the most common sexually transmitted infection in the US, responsible for over 90% of cervical cancer cases. HPV vaccine uptake is low.What was the research question?Evaluate and characterize HPV awareness and vaccine rates in female patients presenting to the emergency department (ED).What was the major finding of the study?The female ED population represents a unique, at-risk cohort in regard to HPV and cervical-cancer risk factors.How does this improve population health?This study challenges emergency physicians to consider an expanded role as HPV educator and vaccine advocate, particularly for “at-risk” patients.We selected a $20,000 annual household income threshold to reflect the 2016 Federal Poverty Line (FPL), the minimum amount of gross income that a family needs for food, clothing, transportation, shelter, and other necessities, as determined by the Department of Health and Human Services.We collected the project data in a secure REDCap database.A total of 81 surveys were completed. Participant demographics are displayed in Selected survey question responses with demographic delineation are displayed in p = .016) and those with higher annual income . Among respondents with awareness of HPV, however, additional understanding of its designation as a common sexually transmitted infection and its association with cervical cancer was generally mixed. White patients were more likely to be aware of the HPV-cervical cancer association . HPV vaccine awareness was higher among patients with an established primary care provider  and lower among those publicly insured or uninsured  and those with lower annual household income .HPV awareness was significantly higher among White respondents  for HPV vaccination, 76% reported they would be comfortable initiating and receiving HPV vaccination in the ED.A number of known cervical-cancer risk factors were also identified among respondents: inconsistent use of barrier contraceptive during sexual activity (59.5%); tobacco use (27.2%); pregnancy prior to age 17 (12.3%); and a family history of cervical cancer (7.6%) .In general, our survey respondents demonstrated a population with low HPV awareness and low HPV vaccination rates along with significant risk factors for HPV exposure and cervical cancer. The HPV vaccine series completion rate reported in this unique ED population was far below national vaccination rates. In 2017, 49% of US teens had completed the HPV vaccine series as compared to less than a quarter of our survey respondents.11In addition, it is alarming to note the number of vaccine-eligible women who reported they had never been counselled about nor offered the HPV vaccine, potentially because they did not have a primary care physician or the resources to see one regularly. Equally concerning was the nearly 13% refusal rate by those who had been offered the vaccine. Reasons given for declination were variable but predominantly included “I feel the illness is not severe or ‘bad enough’ to warrant vaccination,” and “The preservatives in the vaccines are dangerous.” This may represent lack of an established or primary care relationship to offer sufficient education regarding HPV, HPV’s direct association with cervical cancer, and vaccines in general.25The study population also demonstrated a low awareness of HPV’s direct correlation with cervical cancer along with a significant risk profile for cervical cancer. Thirty-five percent were unaware of the HPV-cervical cancer connection, with even higher percentages noted among Black patients (44.9%) and the uninsured (57.1%). This has been demonstrated previously and may represent the influence of socioeconomic determinants of health, which can affect primary care access and utilization of preventive health services.The survey results highlight several potential engagement opportunites for emergency physicians (EP) to consider, including HPV awareness campaigns and ED-initiated HPV vaccine administration. Physician recommendation has been shown to be the most effective indicator of HPV uptake among vaccine-eligible women and may be equally effective in an ED setting.EPs, however, as demonstrated by our survey responses, may represent the only healthcare encounters many women ever have. EDs, as part of the national public health safety net, frequently encounter subsets of the general population who often have minimal interaction with or limited access to the traditional primary care-based healthcare system.Initiating a conversation with EPs and primary care partners and/or community resources may be an important consideration as the healthcare system attempts to address HPV from a public health perspective.The ED setting has more recently been used as a venue for universal screening of other sexually transmitted viral infections, including HIV and HCV. Well established, opt-out screening programs of this type have resulted in the successful linkage of care of previously undiagnosed and untreated HIV- and/or HCV-positive patients, many of whom may have had no other access to testing or definitive care.Future, more expansive studies, considering additional evaluation of the adolescent age range as well as male patients, would be useful to further characterize the at-risk population presenting routinely to the ED setting.This is a single-center study, and as such, results may not be universally applicable to all ED settings. In addition, the results represent a convenience sample collected as a pilot study and may not reflect overall ED demographics. In certain instances , small response numbers may limit data interpretation. Likewise, demographic data on excluded patients, as well as those who refused participation, was not collected in this feasibility study, which may also limit its general applicability. Exclusion of population subsets, particularly non-English speaking individuals, may result in skewed data; however, the majority of non-English speaking patients presenting to our ED represent underserved minorities who face similar healthcare inequities to other minority groups, including decreased rates of HPV vaccination. Thus, we suspect that inclusion of their responses and data would likely contribute to disparities noted in the discussion above. Because survey answers were self-reported by respondents, there was a potential for recall bias and/or social desirability bias, which could have contributed to over- or under-inflation of personal medical history and/or risk factor reporting in particular. Finally, pediatric patients, an important patient subset, were excluded from our study. A nearby children’s hospital caters to the majority of local pediatric emergency patients, limiting pediatric patient exposure in the study ED.HPV awareness and vaccination rates in this female ED population were low while cervical-cancer risk factor rates were high, identifying a particularly vulnerable population. EDs may be a high-yield venue for HPV and cervical cancer prevention including education, screening, and vaccine initiation.


Introduction: A healthcare provider's ability to give a strong recommendation for the HPV vaccine is of utmost importance in increasing HPV vaccination. To reduce the burden of HPV-related cancers, there is a critical need to develop and implement theory-based interventions aimed at strengthening healthcare providers' communication about the HPV vaccine.Methods: We used Intervention Mapping (IM) steps 1–5 to develop and implement a provider-level intervention that aligns with the priorities and needs of a large, urban Federally Qualified Health Center (FQHC).Results: In step 1, a diverse planning group identified barriers to HPV vaccination in clinical settings and generated process maps and a logic model of the problem. Step 2 outlined outcomes and provider performance objectives of the intervention and identified knowledge, skills, self-efficacy, outcome expectations, and normative beliefs as modifiable targets that need to change for providers to deliver strong recommendations for the HPV vaccine to parents and patients. In step 3, the planning group mapped the methods of persuasive communication, information, and modeling and skills training to behavioral targets and outlined the program practical applications (strategies) components, scope, and sequence. In steps 4 and 5, the planning group produced the intervention and planned for program implementation. The iterative and participatory process of IM resulted in modifications to the initial intervention that aligned with the needs of the FQHC.Discussion: IM provided a systematic, participatory, and iterative approach for developing a theory-based provider-level intervention aimed at strengthening healthcare providers' ability to provide a strong recommendation for the HPV vaccine to eligible patients and parents served by a large FQHC. IM assisted with the identification of behavioral targets and methods that move beyond HPV knowledge and reminders to create behavior change. IM can help researchers and planners describe the processes and rational behind developing interventions and may help to facilitate implementation in real-world clinical settings by tailoring intervention components to the needs of the population. Human papillomavirus (HPV) is one of the most common sexually transmitted infections in the United States with an estimated 70% of individuals acquiring the infection at some point in their lifetimes , 2. PersStudies assessing HPV vaccine uptake consistently highlight the importance of healthcare providers' recommendations –20. ReceDespite being one of the strongest predictors of HPV vaccination uptake and coverage, few interventions focus on improving HPV vaccine communication. There is a need for theory-based interventions aimed at strengthening healthcare providers' communication about HPV vaccination for children. Provider-targeted interventions are often limited to delivering provider education about the vaccine or alerts in electronic medical records . ImportaIM is a theory-driven planning framework that provides a systematic process and detailed protocol for effective multi-level intervention development, implementation, and evaluation , 35. IM FQHC leaders collaborated with researchers to develop a multi-level intervention to increase the proportion of FQHC age-eligible patients (11–26 years) initiating and completing the HPV vaccine series in accordance with Advisory Committee on Immunization Practice Guidelines. The overall program goal was to increase the percentage of age-eligible patients initiating the HPV vaccine within 1 year to 30%. For this paper, we describe the use of IM to develop a provider-level intervention aimed at strengthening FQHC providers' recommendation for the HPV vaccine. We define providers as physicians, nurses, medical assistants, and other medical professionals who speak about vaccination with patients and/or parents during in-person clinic visits. This study was approved by the University of Texas Health Science Center at Houston Institutional Review Board.IM provides a systematic framework for program development and planning that can help increase the use of effective practices in healthcare settings –48. IM gIn our study for step 1, we formed a planning group and stated the current behaviors and determinants associated with failures in providing strong recommendations for HPV vaccinations to parents of HPV-vaccine eligible patients. In step 2, we stated the desired intervention outcomes, specified the provider health promoting behavior (making a strong recommendation) and performance objectives necessary to achieve the behavior, and identified psychosocial factors that influence strong provider recommendations (determinants) , 35. PerWe established a diverse planning group comprised of stakeholders with expertise in cancer prevention and control, intervention development, HPV, and FQHC leadership. Specifically, our planning group included FQHC program leaders, 3 intervention design experts, cancer prevention and control researchers, a gynecological oncologist, and 2 program staff. The planning group, led by a cancer prevention and control researcher, met weekly during the development phase and bi-weekly during the implementation phase of the provider-level intervention. Using facilitated discussion and synthesis of provider and staff surveys, the planning group identified barriers to HPV vaccine uptake and failures in FQHC providers delivering a strong recommendations for HPV vaccinations to eligible patients and parents. The group listed barriers to uptake identified through their previous research examining HPV vaccination among adolescents –53, and The behavioral outcome was stated as “FQHC providers deliver a strong recommendation for the HPV vaccine and administering the vaccine.” We identified seven performance objectives for the outcome .Based on previous research, the process map, and surveys from FQHC providers and staff, we identified the following determinants, or cognitive factors that would need to change to achieve the outcome: (1) knowledge; (2) skills and self-efficacy; (3) outcome expectations; and (4) normative beliefs. For example, behavior change models such as Social Cognitive Theory suggest that self-efficacy is a direct and proximal determinant of behavior change and is most predictive of behavioral outcomes –58. To iUsing the matrix of change objectives generated in Step 2, the team developed the program components of the HPV provider training by mapping the change objectives and determinants to change methods, and selected practical applications to deliver the methods . The teaIn step 4, we designed, produced, and pre-tested the components of the intervention. Program materials included detailed flowcharts, synopses and scripts, and text and image vignettes delivered via PowerPoint. The team enlisted support from cancer prevention researchers with experience designing materials for providers to produce program materials.A gynecologic oncologist—a member of the planning group with experience conducting HPV vaccine educational sessions—delivered the in-person provider intervention, which included three components: (1) didactic instruction and education, (2) interactive role-play opportunities using tailored messages, and (3) take-home materials. The didactic educational component highlighted up-to-date research on HPV, HPV-related cancers, vaccination rates, and guidelines, and it emphasized the importance of bundled communication. Additionally, the educational component reviewed individual provider and staff roles during adolescent clinic encounters and outlined processes in place to improve HPV vaccine uptake, such as recall reminders and pre-clinic huddles. The role-playing component was designed to strengthen the communication skills required to provide a strong HPV vaccine recommendation to patients aged 11–12, 13–18, and 18–26 years in accordance with vaccination guidelines. We incorporated communication strategies that staff could use when dealing with hesitant parents and patients, as well as responses to frequently asked questions. The one time, in-person program was designed for 1 h to fit into providers work schedule and all providers also received take-home materials including a fact-sheet about HPV vaccination and clinical guidelines.Based on feedback, the in-person intervention was recorded as a webinar so that providers and staff could view the program at any time. Two versions of the webinar were audio-recorded and delivered based on provider role . First, the planning group identified stakeholders for the implementation of the provider training and developed detailed summaries of the tasks that implementers would need to do to successfully implement the provider training. The planning group identified two primary implementers of the program: a clinic champion, and a gynecological oncologist/HPV vaccine advocate. The gynecological oncologist was a member of the planning group and was involved in the entire planning process. Performance objectives, or sub-behaviors that the gynecological oncologist needed to do for implementation included the following: (1) preparing for the training sessions, (2) coordinating with clinic HPV champion to schedule the in-person provider training, and (3) conducting the in-person provider training. The clinic champion implementation performance objectives included: (1) overseeing implementation efforts, (2) identifying providers who needed the training, (3) providing feedback to planning committee, and (4) interacting with the research team, FQHC providers and staff as necessary to share and address barriers to implementation and provide suggestions for overcoming these barriers. Next, we identified factors  influencing implementation behaviors and crossed these with performance objectives. After completing the matrix of change objectives, we identified change methods to influence the determinants of implementation behaviors and operationalized these into implementation strategies that included coordination meetings with clinic champions, gynecological oncologists training and practice presentations. Between March and August 2016, the gynecological oncologist delivered the intervention to 57 FQHC providers and staff members from four clinics including 10 physicians, 32 medical assistants, 2 nurses, and various administrative staff.Following initial in-person sessions, we conducted interviews and surveys with staff and providers who completed the in-person training to identify areas for improvement. Providers and staff participating in the surveys and interviews requested revisions to the training in order to improve program delivery by decreasing barriers to participation be creating flexible time options for completing the training and to better tailor the content. For example, results from the interviews recommended that we redesign the presentation to fit a webinar format to minimize disruption to busy clinic schedules. Additionally, the providers requested two separate modules for providers and medical assistant and a reduction in the number of slides to decrease the overall training time in hopes of reaching a larger amount of providers. FQHC providers also requested additional information around special populations including men who have sex with men, transgender, and HIV positive individuals. Through analysis of surveys and interviews, we identified additional barriers and facilitators to HPV vaccination that we used to inform the development of case studies that demonstrate how to make a strong HPV vaccine recommendation and address common parental/adolescent concerns. These case studies were incorporated into the revised webinar training.Participatory approaches used throughout both the planning and implementation of the intervention, also informed intervention modifications at each stage of delivery. Modifications to the intervention were based on feedback and guided by of the iterative process of IM, resulting in changes to the content (to increase relevance for the specific needs of the patient population) and delivery of the HPV provider training (to increase provider participation). For example, we updated performance objective four and incorporated new information that outlined the vaccination recommendations for all eligible age groups as well as specific populations frequently served by the FQHC including the LGBTQ community, transgender people, and people with HIV. The FQHC team agreed to distribute the webinar using their online learning management system. Between April 2017 and March 2018, 133 FQHC staff competed the webinar training.Improving healthcare provider communication is one of the most highly prioritized goals in the national movement to increase HPV vaccination rates . Yet, heIn this article, we provided a comprehensive and detailed description of how we used IM to systematically develop a theory-based HPV provider intervention tailored to the needs and preferences of an FQHC. Using IM, we were able to identify and establish sub-behaviors needed to achieve the behavioral outcome and provided a blueprint to map methods and strategies to address multiple determinants derived from theory, including knowledge, skills, self-efficacy, outcome expectations, and normative beliefs. IM encourages participatory approaches to engage stakeholders as part of an iterative process of program planning, development, and implementation . By explThe iterative and participatory process of IM may facilitate the implementation to real-world setting and support reach a larger provider audience. FQHC staff and leadership, responsible for implementing the intervention, served as members of the planning committee and played a critical role throughout the process by ensuring that the intervention addressed the needs and fit of the organization. Having staff and leadership directly involved in the process played a critical role in modifying the intervention to fit the population and demonstrated the organizations commitment which, in turn, may increase sustainability . We acknThere are limitations to our development process using IM. While tailoring to the FQHCs data and delivery platforms improved reach and relevance, this may limit broader intervention scale-up to other clinics and organizations. Additionally, the provider training was designed for providers serving low income, minority communities and may not be effective in other settings since materials may not resonate for other race/ethnicity populations. However, IM provides a blueprint that allows for adaptations to different populations and settings by retaining core program components . FurtherImproving provider communication remains a prioritized goal to increase HPV vaccination coverage. This paper may provide useful insights for researchers, planners, and organizations interested in developing provider communication interventions in highly complex environments by providing a systematic framework that researchers can use to report the processes and rationale behind developing interventions. As a result, the process of IM may help to advance the development of HPV interventions at multiple levels by aiding in the interpretation of intervention findings, helping to identify causal mechanisms driving change, and assisting with adaptation by identifying core components of the intervention.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation, to any qualified researcher.The studies involving human participants were reviewed and approved by University of Texas Health Science Center at Houston Institutional Review Board. The patients/participants provided their written informed consent to participate in this study.MF, LS, and LR: study and manuscript conceptualization. JA: background, methods, results, and discussion. SR, LS, LR, and MF: contributed to background, methods, results, and discussion. TM: contributed to methods. All authors: contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Rwanda and Bhutan, 2 low- and middle-income countries, implemented primarily school-based national human papillomavirus (HPV) vaccination in 2011 (Rwanda) and 2010 (Bhutan). We estimated vaccination effectiveness through urine-based HPV prevalence surveys in schools in 2013–2014 and 2017. In Rwanda, 912 participants from baseline surveys and 1,087 from repeat surveys were included, and in Bhutan, 973 participants from baseline surveys and 909 from repeat surveys were included. The overall effectiveness against vaccine-targeted HPV types  was 78% (95% CI 51%–90%) in Rwanda, and 88% (6%–99%) in Bhutan and against other α-9 types was 58% (21–78) in Rwanda and 63% (27–82) in Bhutan. No effect against other HPV types was detectable. Prevalence of vaccine-targeted HPV types decreased significantly, as well as that of other α-9 types, suggesting cross-protection. These findings provide direct evidence from low- and middle-income countries of the marked effectiveness of high-coverage school-based, national HPV vaccination programs. Recent estimates suggest that, in the year 2018, ≈570,000 new cervical cancers cases occurred worldwide  and 2010 (Bhutan) . Both prThe International Agency for Research on Cancer (IARC), in collaboration with the ministries of health of both countries, is conducting long-term studies, including a series of urine surveys, to provide direct evaluation of the population-level impact of HPV vaccination in LMICs  and 2017 (repeat survey) in both countries . In bothIn the repeat surveys, we aimed at recruiting ≈1,000 female students 18–20 years of age in each country from the same high schools included in the baseline survey. In Rwanda, we included secondary schools in the Nyarugenge District of Kigali. Of the 22 schools (8 public and 14 private), all but 1 overlapped with the 21 schools included in the baseline survey. In Bhutan, we included high schools in the capital of the country, Thimphu (n = 7), and in the nearby town of Paro (n = 3). Of these 10 schools (3 public and 7 private), 6 overlapped with the 6 schools included in the baseline survey. School authorities gave full support to the conduct of the study, and no school refused participation. The repeat surveys were performed during March–November 2017 (Rwanda) and September–November 2017 (Bhutan). Students in the targeted age groups were invited by school staff to attend study information and recruitment meetings. The large majority of students present at information and recruitment meetings signed the informed consent form, but exact denominators of students by age in each school were not available. In Rwanda, 50 students 17 years of age and 24 students 21 years of age also attended recruitment meetings and were allowed to join the study. Similarly, in Bhutan, 10 students 17 years of age, 68 students 21 years of age, and 30 students 22 years of age also joined the study.All students who signed an informed consent form received a device for self-collection of urine. Participants were asked to collect first-void urine from the first urination of the day and to return the urine sample on the same morning as collection. Urine samples were recovered at school entry the day after recruitment, and on this day, a short online questionnaire was filled in by a study interviewer (Rwanda) or directly by the student (Bhutan). The questionnaire included information on places of birth and living, history of sexual intercourse, and recalled HPV vaccination status. Urine samples and questionnaires could only be matched through an anonymized identification number.https://novosanis.com) designed to collect the first 14 mL of first-void urine immediately into 7 mL of a urine-conservation medium to avoid DNA degradation , other α-9 types , other α-7 types , and non–α 7/9 types detected by both genotyping tests . We adapted the framework proposed by Halloran et al. .We computed prevalence ratios (PR) for HPV detection and corresponding 95% CIs by using binomial regression models with a log link. Estimates for Rwanda were adjusted for age group, place of birth, and reported history of sexual intercourse (never vs. ever or prefer not to answer). Estimates for Bhutan were adjusted only for reported history of sexual intercourse because of the small number of infections with HPV vaccine–targeted types observed. HPV vaccine effectiveness (VE) estimates and the corresponding 95% CIs were computed as (1 – PR)%. All statistical analyses were performed by using Stata SE 15.1  in Rwanda. Other participants  were excluded because of a lack of a questionnaire or because students could not recall their HPV vaccination status. Data from 1,087 students in Rwanda  and 909 students in Bhutan  were included in the final analyses  Figure.Chlamydia trachomatis detection (4% vs. 2% [p = 0.052]). In both surveys and in both countries, detection of C. trachomatis was substantially higher in participants who reported a history of sexual activity (HPV vaccination was reported by 962 (89%) of study participants in Rwanda and 864 (95%) in Bhutan, and median age at vaccination was 14 years (range 10–18 years) in Rwanda and 12 years (range 10–19 years) in Bhutan. Among vaccinated girls in the studies, 94% in Rwanda and 90% in Bhutan reported being administered >1 dose of vaccine. In the baseline survey, 43% of participants in Rwanda and 92% of participants in Bhutan reported to be vaccinated. In both countries, the distribution of age at vaccination significantly shifted toward younger ages in the repeat surveys compared with baseline surveys. We compared the distribution of participants in the repeat survey by age group and other selected characteristics with the distribution of the same characteristics as observed in the 912 participants in Rwanda and 973 participants in Bhutan in the baseline surveys . Studentactivity  Table 1.The distribution of participants’ characteristics by vaccination history is detailed in We calculated prevalence and crude PR for groups of HPV types according to GP5+/6+ PCR, in both the baseline and repeat surveys in Rwanda , panel AWe calculated the adjusted vaccine impact on groups of HPV types in both Rwanda and Bhutan, as measured by estimates of overall, restricted, and total VE . In bothAccording to testing with the more sensitive E7-MPG protocol, all HPV prevalence estimates were consistently higher, and corresponding HPV VE estimates consistently lower than the corresponding estimate shown for GP5+/6+. Also, in Bhutan, restricted and total effectiveness were not statistically significant in the E7-MPG PCR  Table 3.By comparing type-specific HPV prevalence among young women in repeat surveys, we have assessed the early impact of HPV vaccination at the population-level in Rwanda and Bhutan, 2 LMICs implementing a national HPV vaccination program. In both countries, high-coverage in schools (≈90%) with quadrivalent vaccine has vastly decreased the prevalence of HPV types targeted by the vaccine , as well as significantly decreasing also that of other α-9 HPV types , suggesting cross-protection (58% in Rwanda and 63% in Bhutan). On the other hand, no changes were observed in other HPV types during this period, suggesting that the prevalence reduction observed in both countries is entirely vaccine-driven and not attributable to changes over time in sexual behavior.An important strength of our present study is the comparability of HPV prevalence estimates in the baseline and repeat surveys in both countries. To this end, we adopted the same methods and procedures to recruit, interview, and test young women and used, as far as feasible, the same high-schools to recruit study participants. To account for behavioral changes that might have occurred in the source population, we adjusted our estimates for the reported history of sexual intercourse, which was more frequent in repeat surveys in both countries. The prevalence of non–α 7/9 HPV types (for which no prior evidence for cross-protection exists) did not significantly change over time. We also did not observe any indications of type replacement.In both countries, HPV vaccination had been introduced before the implementation of the baseline surveys; 43% of study participants in Rwanda and 92% in Bhutan were vaccinated in the catch-up campaigns. Hence, our effectiveness estimates are underestimated because of vaccine-induced protection in the reference group. In particular, estimates of overall effectiveness are affected by both direct and indirect protection in the baseline group, whereas restricted and total effectiveness estimates are affected only by indirect protection. Furthermore, some baseline survey participants might have been sexually active and HPV-infected before being vaccinated. In Bhutan, because of the high vaccination coverage, the number of participants positive to vaccine-targeted HPV types in the reference group was tiny; therefore, effectiveness estimates are imprecise. Nevertheless, irrespective of the genotyping method considered, overall effectiveness was >80%. Restricted and total effectiveness were not statistically significant in tests using E7-MPG PCR. By contrast, in Rwanda where vaccination coverage in the reference group was much lower, estimates of effectiveness against vaccine-targeted and other α-9 types are more precise and consistently statistically significant.We used 2 HPV testing methods of different analytical sensitivity (GP5+/6+ PCR and E7-MPG) to enable us to compare possibly different estimates of VE by assay and overcome the possible problem of the relative lack of sensitivity of HPV detection in urine. Significant overall effectiveness was shown with both methods used. However, we estimated stronger overall VE (78% in Rwanda and 88% in Bhutan) when HPV was measured by using GP5+/6+ PCR. The lower estimated VE might relate to the increased detection of low-level HPV DNA by E7-MPG that might have no clinical significance  (In summary, our study provides direct evidence from LMICs of the marked effectiveness of a high-coverage national catch-up HPV vaccination program. The full impact of vaccination of the routine target cohorts, vaccinated before sexual debut, remains to be measured; continued monitoring will be necessary to assess the sustained impact of the 2-dose HPV vaccination schedule recently introduced in both countries. The reported findings will also be instrumental in supporting a long-term stakeholders’ commitment to HPV vaccination, to inform future budget allocation exercises, and to adapt local cervical cancer screening programs to vaccinated populations. Furthermore, the repeat urine-based survey approach used in Rwanda and Bhutan to monitor the impact of HPV vaccination is well-accepted and remarkably adaptable to a wide range of settings and populations, making this approach particularly valuable to periodically assess the early impact of HPV vaccination.Additional information about impact of human papillomavirus vaccination, Rwanda and Bhutan.


The causative role of human papillomavirus (HPV) in sinonasal squamous cell carcinoma (SNSCC) remains unclear and is hindered by small studies using variable HPV detection techniques. This meta-analysis aims to provide an updated overview of HPV prevalence in SNSCC stratified by detection method, anatomic subsite, and geographic region. From 60 eligible studies, an overall HPV prevalence was estimated at 26%. When stratified by detection method, HPV prevalence was lower when using multiple substrate testing compared to single substrate testing. Anatomic subsite HPV prevalence was higher in subsites with high exposure to secretion flow compared to low exposure subsites. HPV prevalence in SNSCC followed the global distribution of HPV+ oropharyngeal squamous cell carcinoma. Taken together, this meta-analysis further supports a role for HPV in a subset of SNSCCs.n = 1449) were eligible for statistical analysis estimating an overall HPV prevalence of 25.5% (95% CI 20.7–31.0). When stratified by HPV detection method, prevalence with multiple substrate testing  was lower than with single substrate testing , highest in high-exposure anatomic subsites   vs. low-exposure  and highest in high HPV+ OPSCC prevalence geographic regions (North America)  vs. low (Africa) ). While small sample sizes and variability in data cloud firm conclusions, here, we provide a new reference point prevalence for HPV in SNSCC along with orthogonal data supporting a causative role for virally driven tumorigenesis, including that HPV is more commonly found in sinonasal subsites with increased exposure to refluxed oropharyngeal secretions and in geographic regions where HPV+ OPSCC is more prevalent.Human papillomavirus (HPV) drives tumorigenesis in a subset of oropharyngeal squamous cell carcinomas (OPSCC) and is increasing in prevalence across the world. Mounting evidence suggests HPV is also involved in a subset of sinonasal squamous cell carcinomas (SNSCC), yet small sample sizes and variability of HPV detection techniques in existing literature hinder definitive conclusions. A systematic review was performed by searching literature through March 29th 2020 using PubMed, Embase, and Web of Science Core Collection databases. Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines were followed by two authors independently. A meta-analysis was performed using the random-effects model. Sixty studies ( Human papillomavirus (HPV) has been identified as an etiological factor in a subset of head and neck squamous cell carcinomas (HNSCC). HPV-driven tumors arise predominately in the oropharynx ), but also in epithelial-derived tumors of the oral cavity, larynx and nasopharynx, albeit at significantly lower prevalence ,2. OPSCCThe first evidence for a potential etiological role of HPV in sinonasal squamous cell carcinoma (SNSCC) tumorigenesis arose in 1983 with the detection of HPV DNA by Syrjänen et al. . Since tA systematic review was performed by a medical librarian (L.C.) following the guidelines of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) .A search of published studies in Medline via Legacy PubMed (1946-), Embase.com (1947-), and Web of Science Core Collection (1900-) was performed on 6 February 2020 to identify relevant articles. Search strategies were customized for each database (Methods S1). Each search utilized a combination of controlled vocabulary and keywords focused on the concepts human papillomavirus, squamous cell carcinoma, and sinonasal. The search was constructed to exclude non-human studies. No filters for language, study design, date of publication, or country of origin were used in the search producing 1177 articles . All refStudies examining both SNSCC and HPV status in adult patients were considered eligible for inclusion. Included SNSCC histology subtypes were non-keratinizing, keratinizing, papillary, and basaloid squamous cell carcinoma. SCC in which the histological subtype was not further specified was also considered eligible. Other SCC subtypes such as adenosquamous and multi-phenotypic sinonasal carcinoma were excluded along with studies not listing HPV detection methods or discussing cancers originating from the nasopharynx, nasal vestibule, nasal ala or skin.Extracted data comprised geographic region of the study, histology, anatomic subsite, HPV status, HPV genotype, and HPV detection method. During the screening, any study written in a language other than English, Dutch, Arabic, or German (languages spoken by the authors) were excluded. Titles and abstracts were screened by two authors independently (K.C.S.P. and T.M.) for full text review. The same two authors independently conducted the full text review. Any disagreements in the screening process were settled by discussion and consensus between the two authors. Disagreements that could not be settled in this manner were settled in consultation with a third author (D.F.). All eligible studies were screened for duplicate data by comparing authors, timeframe of data collection, and outcomes. After full text screening, 69 studies remained for the quantitative synthesis.2 statistic [p < 0.05 to be significant.Comprehensive Meta-Analysis (CMA) v3  was used to conduct the statistical analysis. To minimize distortion of the results by outliers the CMA program excludes all studies with a sample size of one patient. Using the random-effects model, HPV prevalence estimates including 95% confidence intervals (CI) were computed from sample size and event rates. In case of an event rate of 0% or 100%, the CMA program adds 0.5 to event and non-event values for computation of logit event rates and its variance. HPV prevalence was stratified by detection method, anatomic subsite, and geographic region for descriptive comparison. Subsequently, separate meta-regressions were performed to test the association of each study characteristic with HPV prevalence estimates. Interstudy variability and between-study variance were assessed by Cochran’s Q statistic . The pertatistic , Begg antatistic , and Duvtatistic . A sensiA total of 69 studies were included in the meta-analysis containing a total of 1458 patients with SNSCC (Table 1). There were 324/1458 HPV-positive cases comprising a crude HPV prevalence of 22.2%. After removal of all studies with a sample size of one patient, 60 studies remained for statistical analysis. Estimated HPV prevalence rates ranged from 5.0 to 94.4%. Using the random-effects model, an overall prevalence rate was estimated at 25.5% (95% CI 20.7–31.0) (Table 1).There were 32 DNA-based studies, six protein-based studies, 12 DNA + protein-based studies, two DNA + RNA-based studies, four RNA + protein-based studies, and two DNA + RNA + protein-based studies. There were no studies using only RNA-based detection methods. DNA-based detection methods included DNA in situ hybridization (ISH), Southern blotting (SB), polymerase chain reaction (PCR), dot-blot-hybridization (DBH), and slot-blot-hybridization (SBH). RNA-based detection methods included RNA ISH and reverse transcriptase PCR (RT-PCR). Protein-based detection methods included p16 IHC and NCL-PVp antibody detection. Due to the small sample size in the majority of the subgroups, we categorized studies as either detecting a single HPV substrate (DNA-based or protein-based) or detecting a combination of HPV substrates . Two studies, Deng et al., (2013)  and Larqp = 0.05, I2 = 70.5%). Since the gold standard for determining a tumor is HPV-driven in OPSCC is RNA-based testing, we also wanted to compare RNA-based to DNA- and protein-based detection techniques. This is of particular importance as the diagnostic utility of p16 overexpression in SNSCC remains unclear. As there were no single RNA-testing studies, we split the multi-agent testing group into either RNA  or no RNA (DNA + protein). Again, in line with our expectations, the RNA group yielded the lowest HPV prevalence .As expected, HPV prevalence was lower when a combination of detection methods was used, reflecting fewer false positives, compared to when DNA- or protein-detection was used alone A—this dievalence B; howeveWe next considered HPV prevalence stratified by sinonasal subsite. These data existed in 20 studies. The remaining 40 studies did not specify sinonasal subsite. We categorized anatomic subsites as either high-exposure to refluxed oropharyngeal secretion flow , or low-exposure . In line with our hypothesis, analysis using the random-effects model yielded the highest HPV prevalence in high-exposure subsites  and lower prevalence in less exposed subsites  A with thData for HPV prevalence stratified by geographic region were available for 59 studies. Three studies were conducted in Africa, 18 in North America, 19 in Asia, and 19 in Europe. No studies were available for analysis from South America or Oceania. Using the random-effects model, the highest HPV prevalence estimate was found in North America , in line with existing literature using the National Cancer Database (32.0%) , and thep < 0.001); I2 = 68.7%. Studies included in the subgroup analysis for detection method (single- vs. multi-agent testing) and anatomic subsite were significantly heterogeneous , but not for geographic regions . Meta-regression results indicated that both detection method , with a border-line significance, and anatomic subsite  were associated with the outcome  when the study by Saegusa et al., (1999)  was remop-value of 0.003 showing significant funnel plot asymmetry  increased HPV prevalence in sinuses with more exposure to refluxed secretion flow from the oropharynx and (2) higher HPV+ SNSCC prevalence rates in regions of the world with higher HPV+ OPSCC prevalence. Using the random-effects model, we found an overall HPV point prevalence of 26%. When sub-stratified by single- vs. multi-substrate testing, we identified a prevalence of 21% for multi-substrate testing, which we posit should represent a more accurate number for estimating HPV-driven SNSCC from the cohort available here. As expected, prevalence decreased in a stepwise fashion when tests with increasing specificity were applied. Unfortunately, there were no studies using gold-standard RNA-based detection techniques alone, which met inclusion criteria for the study. This highlights the need for additional, large cohort studies using RNA-based detection methods. It should further be noted that while p16 overexpression is a widely recognized surrogate marker for high risk HPV in OPSCC, whether p16 is a sensitive and specific marker for SNSCC has not been well established. In our systematic literature review we found 18 studies using both p16 IHC and DNA- and/or RNA-based HPV testing. However, the majority of the studies were too small to make a statement on the reliability of p16 IHC. Eight studies reported correlations of 69–100% between p16 overexpression and positive HPV status ,84,90,97In line with our hypotheses, we found the highest HPV prevalence in sinonasal subsites with the greatest exposure to refluxed secretion flow from the oropharynx and the lowest prevalence in sinuses more remote from routine exposure, i.e., the more anterior, cranial and posterior sinuses, each of which also possess restrictive ostium. High-exposure subsites had HPV prevalence rates more than double low-exposure sites (38% vs. 15%). Interestingly, HPV prevalence rates by subsite mirror reported overall survival rates stratified by subsite with frontal and sphenoid sinuses having the lowest survival and nasal cavity having the highest survival ,105. ConAdditionally, we found considerable variation by geographic region, which aligned with HPV+ OPSCCs rates. For example, overall HPV prevalence was highest in North American studies and lowest in African studies in both OPSCC (using previously published cohorts) and SNSCC, with SNSCC HPV prevalence rates in both cohorts being approximately 50% of the OPSCC rate. Remarkably, when restricting to examination of high risk subsites in North American studies (those most likely to be HPV positive), prevalence rates approximate HPV prevalence rates in the oropharynx in some parts of the US (52%). Additional large cohort studies using RNA-based detection techniques are needed to evaluate if these findings remain true, as sample sizes available for these sub-analyses are small.2 statistics make definitive conclusions challenging . Second, there is significant heterogeneity of HPV testing methodologies, each of which have variable sensitivity and specificity. Additionally, the majority of studies (37/69) use DNA testing alone, which may not represent a tumor driven by HPV but instead contamination or a bystander infection. Small sample sizes and heterogeneity of the datasets, as highlighted by the Q and Illenging . DespiteHere, we provide a new reference point prevalence for HPV in SNSCC, stratified by detection method, along with orthogonal data supporting a causative role for virally driven tumorigenesis in SNSCC. Small sample sizes, high interstudy variability and missing data such as genotype-specific incidence highlight the need for large prospective evaluations of HPV in SNSCC and detailed genomic studies to further clarify the role of HPV in SNSCC.


Despite the fact that cervical cancer is preventable and curable in the early stages, it still remains to be a major public health problem in India. This study was conducted to assess the knowledge and awareness regarding the Human Papilloma Virus (HPV) vaccination among health care professionals working in a tertiary care hospital in urban India.To this aim, we conducted a cross-sectional study among 318 health care professionals working in tertiary hospitals across Chennai, Tamil Nadu, India. Our research group designed a structured questionnaire with 31 items to assess the knowledge and attitudes on cervical cancer, its prevention, and HPV vaccination.Among the 318 respondents, 90.6% were aware of cervical cancer, 83.3% were aware that PAP (Papanicolaou) smear test detects cervical cancer, and 86.2% of the respondents knew that HPV causes cervical cancer. 29.2% of the eligible respondents underwent the screening against cervical cancer, and 19.8% of the study participants were vaccinated for HPV. Only 34.9% know that the HPV vaccine could be given to boys. The most common reason for not being vaccinated against HPV was the lack of awareness. In our study, 77.2% of the respondents were willing to be vaccinated and recommend HPV vaccination to their family members.From this study, it was evident that there is a lack of awareness about HPV vaccination and its importance in preventing cervical cancer among healthcare professionals. Our finding clearly establishes the need to devise intervention programs to promote vaccination against HPV and periodical screening for cervical cancer among healthcare professionals. Cervical cancer is the fourth most commonly occurring cancer among women worldwide . India aHuman papillomavirus (HPV) infection causes cervical cancer, particularly HPV 16 and 18 strains account for 75% of these cases . The risThough screening modalities have been in place for more than fifty years, the burden of the disease has not yet reduced as expected in India. For this reason, reinforcement with another preventive measure through vaccination has now been advocated. Bivalent and quadrivalent HPV vaccines are licensed for use in India . The recWhile the HPV vaccine has been in use for more than a decade in India, it is still not provided as an essential vaccine under the National Immunisation Programme by the Govt of India. Lack of awareness regarding its dosage, schedule, and cost exists even in a highly educated population in the health care sector. Health care professionals need to have complete and updated knowledge regarding the HPV vaccine for their own benefit and thus to promote the same among their patients. This study was conducted to assess the current awareness about cervical cancer and HPV vaccination among the health care professionals working in a tertiary care hospital and to overcome barriers associated with the uptake of the vaccine.A Cross-sectional study was conducted at Panimalar Medical College Hospital & Research Institute, Chennai, India. A random sample of 318 health care professionals working in tertiary hospitals in Chennai, Tamil Nadu, India, was chosen for this study. Those participants who were above 18 years of age and willing to give their written consent to participate in this study have been included. We obtained written informed consent from all the participants. The study protocol was approved by the Institutional Review Board (IRB) of the Panimalar Medical College Hospital & Research Institute, Chennai  and conformed to the requirements of the Declaration of Helsinki (as revised in Seoul 2008).The questionnaire/survey instrument was developed by our research team consists of thirty one questions/statements on demographic characteristics and information pertaining to the cervical cancer preventive methods. The socio-demographic variables embrace Age, Sex, Profession, Educational Qualification, good Experiences, legal status, number of children, and case history of cervical cancer. The demographics were followed by the questions/statements related to the knowledge on Cervical Cancer, preventive methods of cervical cancer and knowledge on HPV vaccination. For data collection through this survey on a 3 point Likert scale, the respondents recorded their response on the scale of Yes/No/’I don’t know’. The survey had one question aimed at exploring the plausible reasons reported for not administering  HPV and the respondents were instructed to record their choices from pre-determined 6 choices/reasons (Lack of awareness/High cost/Fear of side effects/Doubt on efficacy/Lack of interest/Do not know the importance of the administration of HPV vaccine). The participants could choose more than one option among the choices provided. The reliability of the tool was analysed by using Spearman’s brown prophecy formula (r) = 0.9.The sample size was calculated based on the “nmasters” software. With a power of 80%, α-error of 5% and prevalence rate of 74%, we arrived at the sample size of 264. Adding 20% as nonresponse error, the final sample size was computed to be 317. Statistical significance was considered to be at 5% level.All the categorical variables are presented as numbers or percentages. Descriptive analysis was performed using univariate statistics to report the Mean and Standard Deviation (SD) for the continuous variable and frequency distributions for the categorical variables. We performed correlation, T-test, and Analysis of Variance (ANOVA) to compare differences in the continuous variables. Pearson chi-square test was used to identify the differences in distribution. The relationship between preventive behaviors  against cervical cancer and related factors  was evaluated using logistic regression analysis. Odds ratios (ORs) and 95% confidence intervals (CIs) were also calculated. All statistical analyses were performed using Statistical Package for Social Science  for Microsoft Windows, SPSS Inc. USA.The socio-demographic characteristics of health care professionals are discussed as follows. Out of the 318 participants (n = 318), 247 were female (77.7%) and 71 were male (22.3%). Around 39.9% of the respondents had less than six months of professional experience. Overall health care professionals who participated in this study were computed to be 53.5%, of which 5.5% were practicing medicine, and 42.2% were nurses. Apart from this, 73% of the participants were observed to be unmarried, and 27% were married professionals  of the participants were aware of cervical cancer, whereas the awareness index in the studies reported earlier was lesser than that of the participants in this study 0.6% of t. When asA mixed-method study was conducted in Dhaka, Bangladesh in December 2013 among professional women employed by private banks. It revealed that only 26% of the participants had  heard about cervical cancer and 56.6% of them had  heard about HPV vaccination .In the present study, 83.3% of people knew that PAP smear test detects cervical cancer even before the symptoms appear, which were in comparison with similar studies –15. WhilCommon reasons for not getting vaccination reported in our study were lack of awareness regarding HPV vaccination, the importance of the vaccine, and high cost which was in agreement with other studies reported earlier , 22.6 India is considered to be a slightly conservative country where there are still taboos regarding cervical cancer because HPV infection is predominantly sexually transmitted. Providing information regarding HPV vaccination and screening sensitively can help in the proper execution of these interventions. Unlike other vaccines, the HPV vaccine does not provide 95–100% protection against HPV infection and even after completion of a full course of this high-priced vaccine, one must undergo screening for HPV infection lifelong. All these are seen as drawbacks and reasons for poor uptake.The same finding was also observed in the study conducted by Bhuiyan et al. , which aThe profession, knowledge on cervical cancer, HPV screening, and availability of vaccine-associated  have a negative effect on the administration of HPV vaccination. Knowledge on HPV vaccine to pregnant women associated with showed a positive effect on HPV vaccination. When all the variables were put together, only the variables on the knowledge of HPV vaccination in pregnant women were positively associated with the decision to administer the HPV vaccine or recommend the administration of HPV vaccination to the family and friends.There is a large gap that has to be filled to improve awareness about HPV vaccination. This could be achieved by promoting awareness emphasizing that every child needs to be vaccinated, and every mother needs to be screened. In developing countries like India, investing and motivating towards preventive measures like vaccination and screening of cervical cancer could help reduce the burden of advanced disease. Since the HPV vaccine is not available to free of cost currently by the Govt of India, its uptake in society is very poor. This needs to be addressed at the earliest because many countries like Australia have nearly eradicated cervical cancer among their population through vaccination and active screening . ConductOur study was aimed at identifying the level of awareness among health care professionals in a tertiary care centre catering to the urban population. Hence, these results cannot be generalized to the whole population.From our findings in this present study, it is evident that there exists a lack of awareness about HPV vaccination and its importance in preventing cervical cancer among health professionals. This clearly establishes the need to devise intervention programs to promote vaccination against HPV and screening for cervical cancer among healthcare professionals.


We conclude that HPV self-sampling is an acceptable option for women at increased cervical cancer risk who have been long-term screening non-attenders. Educational outreach to enhance Specific Knowledge about HPV, CIN and cervical cancer is critical. Those non-attenders who explicitly avoid gynecologic examinations need special attention. Trial Registry: Clinicaltrials.gov NCT02750124This study aims to investigate acceptance of vaginal self-sampling for high-risk human papilloma virus (HPV) among long-term screening non-attenders at increased cervical cancer risk and to identify leverage points to promote screening adherence among these women. Forty-three long-term screening non-attenders performed home vaginal self-sampling for HPV, had positive HPV results, and subsequently attended gynecologic examination. Sixteen (37.2%) had high-grade cervical intraepithelial neoplasia (CIN2 or 3), and two had invasive cervical cancer. Forty-one of these women completed a questionnaire concerning Specific Knowledge about HPV, CIN, and cervical cancer, potential barriers to screening and views about self-sampling. Results were compared with 479 women treated for CIN2+ who attended gynecologic follow-up and also performed self-sampling. Significant multivariate predictors of long-term non-attender status compared with referents were low Specific Knowledge, high confidence in self-sampling, and potential barriers—refraining from activity to attend gynecologic examination, needing another's help to attend, and long travel time. Non-attenders citing fear/refraining from gynecologic examination as why they preferred self-sampling significantly more often had lowest Specific Knowledge compared with other non-attenders. All non-attenders could envision themselves doing self-sampling again while only 74% of referents endorsed this statement ( Cervical cancer (CC) continues to be a frequent cause of cancer death worldwide, particularly in less developed regions, both within and outside Europe . ThroughLong-term infection with high-risk human papilloma virus (HPV) is the causative agent of the nearly all CC. Compared with cytology, primary screening based upon HPV confers 60–70% greater protection against high-grade cervical intraepithelial neoplasia (CIN2+) and invasive CC . Women wHerein, we examine these questions, focusing on long-term non-attenders to the national organized cervical screening in Sweden, who performed self-sampling with a positive result for HPV and thereafter attended recommended gynecologic follow-up. We assess their Specific Knowledge and explore reasons why self-sampling is or is not acceptable for them. Detailed comparisons are performed with the 479 women treated and followed up for CIN2+. We thereby seek to identify potential leverage points to promote adherence to cervical screening guidelines among long-term non-attenders at increased CC risk and to glean insights into how to best meet the needs of this vulnerable group.The study was carried out within the population-based cervical screening program of Stockholm County. A target group of long-term screening non-attenders is compared with women treated and followed up for CIN2+ (referents). Both groups performed self-sampling for HPV. The target group was selected based on testing positive for HPV and attending gynecologic exam thereafter. Questionnaire data were collected among both groups.Clinicaltrials.gov NCT02750124), 4000 of these women (24.3%) were randomized to receive an HPV self-sampling kit by mail or were invited to order the kit via an eHealth Web application. Further details on the methodology and results of that study are described in Ref. [Approximately 4% of women who were eligible for cervical screening and had received at least 10 annual invitations were considered long-term cervical screening non-attenders. As part of a health services study , Stockholm , 14. NonThe questionnaire was in Swedish. (The English-language translation can be accessed through Ref. ). The qut” tests were employed for some normally distributed continuous variables and Mann-Whitney tests of continuous variables that were not normally distributed. In Ref. [β estimates, standard errors, odds ratios (OR), and 95% confidence intervals (CI) were computed for salient independent variables in adjusted models, with the outcome variable: long-term non-attender to cervical screening with HPV-positive findings from self-collected samples.We performed univariate data analysis for the target group: long-term screening non-attenders with positive HPV results. Detailed comparisons were made between the target group and the referents. Analyses were primarily carried out using Yates' chi-squared test (or Fisher's exact test used if any expected cell was < 5). Two-sample “ In Ref. , factor t” test, p = 0.000).Of the 43 women in the target group, on cone histology fourteen (32.5%) were diagnosed with CIN2 or CIN3, two (4.6%) had invasive cervical cancer, and 27 (62.8%) had normal findings or CIN1. All were age 34 or above  and the majority were over age 40. As described in Ref. , among tFigure The non-attenders unanimously endorsed that HPV self-sampling was easy to perform, sufficient information was provided, and they could envision themselves performing self-sampling prior to the next gynecologic examination. These were all significantly higher endorsements compared with the referents Table . The perχ2 = 4.88, p = 0.027). The two groups did not differ significantly vis-à-vis annual income, civil status , or employment status (over 70% were employed in both groups). Perceived risk of developing cervical cancer without regular gynecologic follow-up (dichotomized at 7 of 10 maximum) was significantly lower among the non-attenders . Most of the preferred sources of further information about HPV, cervical cancer, and preventive measures were endorsed in similar percentages among the two groups. However, a larger percentage of the non-attenders would like more information via postal mail . Significantly more of the non-attenders stated that they refrained from some activity to attend the gynecologic examination. They had also significantly more often taken time off from work and required another person's help to attend. Total time spent traveling was significantly greater in the non-attenders.Fewer of the non-attender group had completed university education (39%) compared with referents (58.2%)  compared with those who did not note this as a reason for self-sampling. Fewer of those who explicitly feared/refrained from gynecologic examination expressed high confidence in Pap smears . Educational level did not significantly distinguish these two sub-groups.Fifteen non-attenders explicitly noted fear/refraining from gynecologic examinations as a reason for readiness to do self-sampling. A zero score on Specific Knowledge was more frequent among those who feared/refrained from gynecologic examination , likely due, in part, to their very low Specific Knowledge. Overall risk among the target group as a whole is lower, since over 60% had normal findings or CIN1, whereas the control cohort all had CIN2+. This difference is a limitation of the present study for comparing the two groups.In our previous analyses of the referents, Specific Knowledge scores showed a significant multivariate association with realistic assessment of cervical cancer risk without regular gynecologic follow-up (score ≥ 7) . HoweverAnother difference between the two groups is age-related. Since invitational screening in Sweden begins at age 23, the youngest women in the target group were 34 years old. Among the control cohort, this age constraint was not in operation and nearly 40% were 30 or younger. These age differences were fully taken into account in the multivariate analysis, as were differences in educational level.We anticipated low educational levels among women who explicitly cited fear or other reasons for refraining from gynecologic examination. However, this was not the case. In fact, six of these fifteen women had completed university education. Strikingly, 40% of these women had zero Specific Knowledge scores, suggesting that they actively avoided information about HPV, CIN, and cervical cancer.The unanimous acceptance of self-sampling and significantly greater percentage of highest confidence in self-sampling compared with referents are also striking findings. These findings, including those of multivariate model 2, suggest that self-sampling could be another leverage point for promoting participation in screening among long-term non-attenders. It should, however, be noted that self-sampling was performed in different settings: at home for the target group versus at the clinic for the referents. Moreover, the referents collected vaginal and urine samples, whereas the target group collected only vaginal samples.Swedish data show that vaginal self-sampling for HPV is an effective strategy for increasing population coverage . The earPractical issues associated with attending gynecologic examination were identified as significant in both multivariate models. Namely, the target group was distinguished from referents by significantly more often requiring another person's help and refraining from activity to attend gynecologic examination, plus more often having traveled over 1 h to and from clinic. These were likely barriers to attending clinic-based screening. Concordantly, among a population-based cohort in Stockholm, a significant association was reported between taking time from work and non-adherence to clinic-based cervical screening . Time coSeveral women who indicated that they feared/refrained from gynecologic examinations explicitly stated that the reason was having suffered sexual abuse. In a study from southeastern Sweden , among 1The present study yields insight into a unique group of cervical screening non-attenders. Their increased cervical cancer risk associated with HPV-positive findings was identified via self-sampled specimens. Subsequently, these women agreed to attend gynecologic examination and 95% completed the questionnaire. The fact that all but two of these 43 women completed the questionnaire suggests that once these women are under care, they may become more proactive. They could potentially serve as a social norm to encourage their family, friends, and wider network to attend screening, or at the very least, to perform self-sampling. Still, this target group is highly selected and represents a very small percentage of long-term non-attenders. Efforts are needed among the larger remainder of non-attenders. In particular, women of ethnic minority background, especially those not fluent in the host country language, are often non-attenders . A reasoFurther studies are needed, focusing on these hardest-to-reach long-term non-attenders. A promising intervention would be to systematically promote self-sampling for HPV within community health programs, concomitant with multi-cultural and multi-linguistic educational efforts through various media. These efforts would likely be most effective if closely coordinated with population-based cervical screening. Therein, knowledge is critical for decision-making regarding screening and for choosing the screening modality that best suits the individual.


Determine the presence and prognostic value of human papillomavirus (HPV), Epstein-Barr virus (EBV), Merkel cell polyomavirus (MCPyV), and cell cycle proteins in head and neck squamous cell carcinoma (HNSCC) of non-smokers and non-drinkers (NSND).in situ hybridization (ISH) was used to screen for the presence of HPV and MCPyV mRNA. Immunohistochemistry was performed for expression of p16 as surrogate marker for HPV, Large T-antigen for MCPyV, and cell cycle proteins p53 and pRb. Positive virus results were confirmed with polymerase chain reaction. For EBV, EBV encoded RNA ISH was performed. Differences in 5-year survival between virus positive and negative tumors were determined by log rank analysis.Clinical characteristics and tumors of 119 NSND with HNSCC were retrospectively collected and analyzed on tissue microarrays. RNAscope All oropharyngeal tumors (OPSCC) (n = 10) were HPV-positive, in addition to one oral (OSCC) and one nasopharyngeal tumor (NPSCC). The other three NPSCC were EBV-positive. MCPyV was not detected. Patients with HPV or EBV positive tumors did not have a significantly better 5-year disease free or overall survival. Over 70% of virus negative OSCC showed mutant-type p53 expression.In this cohort, all OPSCC and NPSCC showed HPV or EBV presence. Besides one OSCC, all other oral (n = 94), hypopharyngeal (n = 1), and laryngeal (n = 9) tumors were HPV, EBV, and MCPyV negative. This argues against a central role of these viruses in the ethiopathogenesis of tumors outside the oro- and nasopharynx in NSND. So, for the majority of NSND with virus negative OSCC, more research is needed to understand the carcinogenic mechanisms in order to consider targeted therapeutic options. Viruses play an increasing role in head and neck squamous cell carcinoma (HNSCC). High-risk human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) has been identified as an entity with a different carcinogenesis than traditional HNSCC resulting from excessive tobacco and alcohol consumption. HPV is also an independent prognostic factor for a better disease free survival (DFS) and overall survival (OS), which has led to a down staging of these tumors in the eighth edition of the American Joint Committee on Cancer (AJCC) and union for International Cancer Control tumor-node-metastasis (TNM) classification –3. Becauvia PD-L1 overexpression, making these patients interesting candidates for checkpoint blockade therapy (in situ hybridization (ISH) . EBV has a strong association with nasopharyngeal squamous cell carcinoma (NPSCC), approaching a prevalence of 100% in these tumors, and is endemic in Southern China, Southeast Asia, Northern Africa, and the Mediterranean basin , 10. It on (ISH) , 11.Lately, besides these acknowledged oncogenic viruses, there is attention for polyomaviruses in HNSCC. Merkel cell polyomavirus (MCPyV) has not only been detected by digital transcriptome subtraction and polymerase chain reaction (PCR) in up to 80% of Merkel cell carcinoma of the skin, but also in non-malignant tonsillar tissue, oral squamous cell carcinoma (OSCC), and pharyngeal cancer, with a reported prevalence of 23, 6.6–29, and 50% respectively –17. AlthTP53 and CDKN2A, leading to cell proliferation and prevention of apoptosis, among others. In HPV-related OPSCC, HPV integration in the host cell DNA genome leads to deregulation of oncoproteins E6 and E7, resulting in inactivity of p53 and retinoblastoma tumor suppressor gene product pRb, respectively. The negative feedback of pRb inactivation leads to p16 overexpression  remain largely unclear, but a significant role of oncogenic viruses would be expected. Indeed, a higher prevalence of HPV in these tumors has been reported in several studies, though in small numbers of patients –25. TherConsecutive patients with HNSCC were selected at the University Medical Center Utrecht (UMCU) and Maastricht University Medical Center (MUMC). In the UMCU, patients were prospectively selected between 1980 and 2004, as described previously . In the The Medical Ethics Review Committee of the MUMC (2018-0567) has approved this study and the principles outlined in the Declaration of Helsinki were followed. All data and tissues were handled according to General Data Protection Regulation.FFPE blocks of either the diagnostic biopsy or tumor resection were retrieved and hematoxylin and eosin sections were digitally evaluated with a senior head and neck pathologist (SW or MH), using Pannoramic viewer . Per patient, three 0.6mm tumor tissue cores and one normal epithelium core were selected, placed in a tissue microarray (TMAs), and cut into 5 μm sections.To screen for the presence of HPV and MCPyV mRNA, the RNAscope 2.5 RED assay kit and HPV-16/18 or V-MCPyV-LT-ST-Ag probe cocktails  were used according to the manufacturer’s instructions. In short, TMA sections were deparaffinized and pretreated with RNAscope Hydrogen Peroxide for 10 min. Antigen retrieval comprised of boiling the slide sections in the provided Target Retrieval Reagents solution at 100°C for 15 min. After washing, the TMAs were dried over night at room temperature and treated for 30 min with RNAscope Protease Plus. In situ hybridization was performed applying four droplets of the provided probes prior to each of the six amplification steps . After each hybridization step, the slides were washed in the RNAscope wash buffer for 2 min at room temperature. Subsequent to alkaline phosphatase Fast Red chromogenic visualization of hybridized probes, the slides were counterstained with hematoxylin and assessed under a bright field microscope at 200x magnification. Tissue with at least 1 red punctate signal dot in the cytoplasm and/or nucleus of malignant cells was considered to be positive, as suggested by the manufacturer for genes with an expression level varying between 1 to >10 copies per cell. Probes for housekeeping gene transcript human peptidylprolyl isomerase B (Hs-PPIB) and bacterial dihydrodipicolinate reductase gene (dapB) transcript were used as positive and negative controls, respectively. As virus specific positive controls, virus positive tumor tissue was used, in addition to the HPV-18 positive cell line HeLa, HPV-16 positive cell lines SiHa and Caski, and MCPyV positive cell lines MKL-1, MKL-2, and WaGa Figure 1EBER-ISH was performed using the Dako fluorescein-labeled EBV peptide nucleic acid (PNA) probe mixture and PNA ISH Detection kit  according to the manufacturer’s instructions. Briefly, following TMA section deparaffinization, target retrieval was performed with the Dako Omnis ISH Pre-Treatment solution for 5 min. Subsequently, enzyme pre-treatment was carried out using two steps of 3 min ethanol 96% application and one step of Dako ISH Pepsin for 15 min. Once the provided EBER RNA CISH probe was applied, denaturation at 66°C for 10 min, and hybridization at 45°C for 90 min followed. The slides were washed with the ISH Stringent Wash Buffer for 3 min, and the provided reagents were applied for staining: CISH Endogenous Enzyme Block for 3 min, Anti-FITc-AP for 30 min, and BCIP-NBT Substrate for 15 min. Sections were counterstained with Nuclear Fast Red and analyzed under a bright field microscope at 200x magnification. Strong blue staining of more than 50% of tumor nuclei was considered to be positive. In parallel, a probe for housekeeping gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used to ensure the presence of mRNA in the TMA and a case of EBV-positive infectious mononucleosis served as a positive control Figure 1Three-μm FFPE TMA sections were subjected to IHC, using primary monoclonal antibodies directed against p16, p53, pRb, and the LTAg of MCPyV Table 1.Of patients with a positive result for HPV RNA-ISH and/or p16 IHC, DNA was isolated from eight 5-μm FFPE whole tissue sections with the Maxwell RSC DNA FFPE kit . DNA concentrations were determined using the Quantus Fluorometer and the QuantiFluor ONE dsDNA system (Promega). Next, 250 ng DNA was added to 1 ml SurePath preservative fluid  and used for HPV-DNA analysis utilizing the COBAS 4800 platform , according to the manufacturer’s instructions. The COBAS 4800 tests specifically for HPV-16, HPV-18, and a combination of 12 other HR-HPV types . Results were considered reliable when the controls were labeled “valid”. Housekeeping gene β-globin was used as a control for the human DNA, in addition to DNA samples of an HPV-positive and HPV-negative tumor.p < 0.05) or near significant (p < 0.1) relationships were evaluated in multinomial logistic regression to assess predictors for DFS and OS. Analyses were performed using IBM SPSS Statistics 25.0  and a p-value of <0.05 was considered to be statistically significant.Patients were considered to be HPV or MCPyV positive when the virus was detected with at least two techniques . EBV presence was based on the EBER-ISH result. Differences in clinical parameters between virus positive and virus negative patients were evaluated by Mann-Whitney U test for age because of a non-normal distribution, Fisher’s exact test for binominal variables , and the Fisher-Freeman-Halton exact test for tumor location, T-stage, N-stage, and p53 because of expected counts of less than five. The 5-year DFS and OS were estimated with Kaplan-Meier curves and differences between virus positive and negative tumors were determined by log rank test. DFS was defined as the last date of treatment until the biopsy date of a histologically proven recurrence or second primary tumor in the head and neck region. OS was defined as the time between the primary tumor biopsy date and death. Censoring took place when patients were lost to follow-up, deceased without recurrent disease for DFS, or at the cut-off point of 60 months. All clinical and pathological parameters were assessed in bivariate analysis regarding survival. Variables with significant  and were mainly women (78%) with a tumor of the oral cavity (80%) and no regional or distant metastases . Thirty-one patients (26%) had recurrent disease within 5 years Table 2.ISH on the TMAs showed HPV-16/18 mRNA expression in tumors of ten patients. All of these tumors showed p16 overexpression by IHC as well, in addition to five tumors with no HPV-16/18 mRNA expression. COBAS analysis on HR-HPV DNA in these 15 patients detected the presence of HPV-16 in ten tumors and another HR-HPV type in one other case. For patient 61, the quality of the DNA was insufficient for COBAS analysis. This resulted in a total of 12 tumors being HPV-positive based on at least two detection techniques Table 3.p = 0.003), oropharyngeal origin , and N2-stage   Table 2.p = 0.33 and p = 0.27, respectively), compared to HPV-negative HNSCC in NSND (p = 0.008), T1 stage (p = 0.0047), and N0 or N1 stage (both p < 0.001) were retained in the best multivariable model as predictors for OS in HNSCC of NSND .Four of the twelve patients with HPV-positive tumors (33%) died within five years, two of which had recurrent disease. For patient 1 there were no details recorded on the cause of death (OS = 59.6 months), patient 5 died of the complications of an aortic valve prosthesis endocarditis (OS = 23.7 months), patient 21 received palliative treatment because of distant metastasis (DFS = 1.6 months), and patient 61 developed liver metastases 20.1 months after initial therapy Table 3.p < 0.001; p53: 83 versus 27%, p < 0.001; pRb: 83 versus 19%, p < 0.001)  Table 5.p < 0.001), resulting in virus positivity of all four nasopharyngeal tumors in this cohort (three containing EBV and one HPV). Two of the three patients (67%) with EBV-positive tumors were below 50 years of age and non-Caucasian (Northern African and East Asian) and they all had regional metastases   showed p53 mutant-type overexpression. This did not differ significantly from cell cycle protein expression in EBV-negative tumors, most probably because of the small number of EBV-positive tumors Table 5.MCPyV was not detected in any of the samples with RNA-ISH or IHC against the LTAg of MCPyV.None of the squamous cell carcinomas of the oral tongue (OTSCC) (n = 39), larynx (LSCC) (n = 9), or hypopharynx (HPSCC) (n = 1) showed involvement of HPV, EBV, or MCPyV. Except for one tumor of the alveolar process, all other OSCC (n = 54) were virus negative as well Table 2.p = 0.021) and N0 or N1 stage (both p < 0.001) were retained in the best multivariable model as predictors for OS in NSND with virus negative OSCC .The three patients with virus negative OSCC containing p16 overexpression all had recurrent disease within 6 months after surgical resection . Two of these tumors showed loss of pRb expression. A poor DFS  was also found in three other tumors in this cohort with some p16 expression (>50%): all OTSCC with a pRb expression above 50%. Apart from one LSCC, all tumors with loss of pRb were OSCC (19/20) without any expression of p16. A younger age at cancer diagnosis (ive OSCC Table 6.Over 70% (76/104) of virus negative tumors showed mutant-type p53 expression, with 0-type mutant expression in 20% (19/94) of OSCC, 56% (5/9) of LSCC, and 100% (1/1) of HPSCC, and mutant-type overexpression in 52% (49/94) of OSCC and 22% (2/9) of LSCC Table 5.The objective of this study was to determine if HPV, EBV, and MCPyV play a role in head and neck carcinogenesis of NSND, the role of cell cycle proteins p16, p53, and pRb regarding viral presence, and the influence of these viruses and proteins on patient survival. In this cohort of 119 NSND, the ten oropharyngeal (100%) and four nasopharyngeal (100%) tumors contained either HPV or EBV. Besides one oral cavity tumor, all other specimens of the oral cavity, hypopharynx, and larynx were HPV, EBV, and MCPyV-negative. Virus positivity did not predict better disease free or overall survival. Regarding cell cycle protein expression, HPV-positive tumors showed more p16 overexpression, wild-type p53 expression, and loss of pRb compared to HPV-negative tumors. OSCC with >70% p16 expression had a poor DFS and OS, with loss of pRb in two of the three cases. The other pRb negative tumors were mainly OSCC as well and did not show p16 expression. Mutant type p53 expression was observed in over 70% of virus negative HNSCC.As the worldwide HPV prevalence is rising, a wide range has been reported in the literature, ranging from 20% in OPSCC patients from Eastern Asia or Central America, to over 50% in Europe and Australia, based on HPV DNA combined with E6*I mRNA or p16 IHC . This isvia structural alterations, or maybe as a result of other oncogenic viruses affecting pRb expression that we are not aware of  of OSCC showed p16 overexpression with IHC, and one of those contained HPV-16 DNA following COBAS analysis resulting in HPV-positivity according to detection by two methods. p16 overexpression could result from loss of pRb function aware of . Lechnere tumors , 37. NSD anymore , 38. The anymore . As the al cells . The thrung NSND .The 5-year OS of HNSCC patients in general is 40–50%, whereas it is 70–80% for patients with HPV-positive OPSCC , 42. In Viral association in all four NPSCC patients of this cohort was as expected. Although the worldwide incidence of EBV related NPSCC has been decreasing over the past decade, the prevalence is still high with almost 100% in endemic regions , and 60–85% in non-endemic regions , 43–46. TP53 mutations  in the current study  patients with OTSCC . AlthougHPV has its clinical relevance in routine practice as a prognostic marker in OPSCC, with a better DFS and OS, in addition to an improved radiosensitivity compared to HPV-negative OPSCC because of altered DNA repair, reduced hypoxic regions, and an increased cellular immune response , 53. WitOne of the limitations of this study was that the definition of when a patient was a non-smoker and non-drinker was collected retrospectively from their medical records. Nevertheless, a strict definition was used , based on a standard history taking template including specific questions on any current or previous tobacco and alcohol consumption, during two separate hospital visits (Head and Neck outpatient clinic and Anesthesiology screening). Secondly, there was a small group of patients with tumors at anatomical sites other than the oral cavity (OSCC). In combination with the strict NSND definition, this resulted in no virus negative OPSCC and NPSCC. However, it has been reported that NSND are mainly patients with OSCC, so a small number of tumors outside the oral cavity was expected , 56. ThiA high prevalence of HPV and EBV was observed in OPSCC and NPSCC of NSND respectively, but not in HNSCC outside these locations. Although a significant role of oncogenic viruses would be assumed in this specific patient group lacking the traditional risk factors for developing HNSCC, HPV, EBV, and MCPyV were not detected in this relatively large cohort of 95 OSCC apart from one case, using clinically relevant cut-off values. This argues against a central role of these viruses in the etiopathogenesis of oral, hypopharyngeal, and laryngeal squamous cell carcinoma in this specific patient group. With ongoing de-escalation trials for HPV-positive OPSCC and trials for immune checkpoint inhibitors in the treatment of EBV-positive NPSCC, the treatment of NSND with tumors at those locations may change in the near future. However, for the majority of NSND with virus negative OSCC, more research is needed to understand the carcinogenic mechanisms in order to consider targeted therapeutic options.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Maastricht University Medical Center (METc 2018-0567). Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.All authors have contributed substantially to the conception and design , acquisition of data , analysis and interpretation of data ; drafting the article  or revising it critically for important intellectual content ; final approval of the version to be published ; and agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All authors contributed to the article and approved the submitted version.SW receives a research grant from Roche, MSD, AstraZeneca, Pfizer, Nextcure, Amgen, and Bayer. BK and ES receive a research grant from Pfizer and Novartis. The funders had no role in the design of the study, in the collection, analyses, or interpretation of the data, in the writing of the manuscript, or in the decision to publish the results.The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.