Purpose: There is a gap in social science literature addressing issues of access and quality of care for transgender military veterans. Psychologists, medical doctors, and other health professionals are beginning to address some of the barriers present in the Department of Veterans Affairs (VA) system that affect veterans who are also transgender and intersex. Over a 7-year period, between 2006 and 2013, 2600 transgender veterans were served by the VA. Data from several surveys revealed that most transgender veterans perceive the Veterans Health Administration (VHA) to be less than accommodating for their special needs. The goal of this study was to investigate the experiences of a sample of transgender veterans with regard to their experiences with healthcare services provided by the VHA.Methods: Using snowball sampling techniques, we were able to recruit 22 transgender military veterans to participate in our study. A combination of telephone interviews and questionnaires provided data from veterans in various branches of the military throughout the United States.Results: Findings indicate that even though the VHA is working to address issues of inequality for transgender veterans, our participants indicated that there are still some problems with administration of care, proper training of staff and physicians, and availability of comprehensive services for the unique healthcare needs of transgender individuals.Conclusion: Since our data were collected, the VA has worked to bridge the gap by focusing on increased training for VHA providers and staff and establishing LGBT programs at VA facilities. However, we suggest that one key area of importance should continue to focus on how mental health and medical providers and ancillary staff are trained to interact with and provide care for their transgender patients. Of the >125,000 National Guard and >48,000 Reserve members included in that study, about 57% and 46%, respectively, enrolled with the VHA within 12 months of being demobilized.3 These figures illustrate the importance of the VHA to our military veterans.Although there are only an estimated 1.4 million people in the United States who identify as transgender, current statistics shows they are more likely to serve in the military than their peers.4 The goal of the most updated directive, VHA 2013-003, is to also ensure that adequate healthcare is provided to transgender and intersex veterans. Since transgender individuals are at greater risk of experiencing depression, suicidal behaviors, substance abuse, violence, and homelessness than other people in the population, the authors contend that available evidence suggests the same health risks can be applied to transgender military veterans.6 Other findings indicate that between 2006 and 2013, the VHA served a total of 2600 transgender veterans.4In 2007, the VHA began focusing on local policies regarding provision of care to transgender veterans and then on June 1, 2011, VHA Directive 2011-024 was issued in an attempt to standardize treatment services for transgender veterans.According to participants in this research study, negotiating their gender identities within the intersection of public/private spaces such as mental health and medical environments is both stressful and complicated. As the data indicate, some of the transgender veterans were faced with a multitude of issues directly related to their gender identity when seeking heathcare with the VHA. Our study adds to the literature dealing with transgender individuals, specifically transgender military veterans who need quality mental health and medical services. As the review of the literature reveals, there is little qualitative research on this subject matter.* and other procedures needed for transition; an issue for some of the veterans interviewed in our study. Although a step in the right direction, we suggest that the VA continue to focus on training mental health and medical providers and ancillary staff to provide proper care for their transgender patients, including provision of better access to a system that is steeped in bureaucratic rules and regulations relating directly to gender identity. The review of the literature that follows specifically addresses the experiences of transgender veterans with healthcare providers at the VHA as well as some of the current programs that are being utilized to positively impact transgender patient care.Our data focus on the participant's experiences with the Department of Veteran Affairs (VA). It is important to point out that our participants were interviewed during and after the VHA Directive 2013-003 was released by the Department of Veterans Affairs on February 8, 2013. The Directive was written to ensure that adequate healthcare is provided to transgender and intersex veterans. Since our interviews were conducted, the VA has made improvements by providing more training and resources to VHA staff and outreach to transgender veterans. However, at the time of our interviews and currently, the VA does not provide sex reassignment surgery (SRS)7 They found that the use of the VHA was higher among transgender veterans than the general population of veterans. Approximately 9.3% of the sample sought mental health services from the VHA and the population had higher than average levels of depression when compared to the general nonmilitary population. Veterans in the study also reported suffering from post-traumatic stress disorder (PTSD) along with the need for gender identity counseling. Documented barriers included the cost of health services and the veterans' perceived reaction by medical providers to their gender identity.In a study including 141 trans women veterans, the researchers examined healthcare utilization and potential barriers to accessing care for this population.8 Focusing on the data collected in 2008 from the University of California, Santa Barbara, the researcher found that in regard to the VA, there were patient care problems and lack of cohesive polices as these relate to transgender veterans. For instance, some patients were given inconsistent information about SRS, were not addressed using pronouns that aligned with their gender identity, and in some cases, were refused service by the VA. Overall, transgender veterans reported that they dealt with many mental and medical health restrictions.One study directly explored discriminatory practices aimed at transgender military personnel.9 The sample consisted of 202 VA providers and 58 LGBT veterans. One finding indicated that 93% (n=54) LGBT veterans thought that it was somewhat or very important for the VA to be welcoming, while 81% (n=164) of the VA providers shared this same view. Despite these findings, only a minority of both VA providers and LGBT veterans viewed their hospital as welcoming. The researchers concluded that providers should have more training in working with sexual and gender minority clients along with exposure to literature documenting risk factors for LGBT patients.Researchers conducted a study in two VA centers in Oklahoma and Texas in hopes of improving service to LGBT veterans.10 The researchers recruited and interviewed 11 transgender veterans from the 2013 Southern Comfort Conference in Atlanta, Georgia. Like the current study, the researchers found that transgender veterans continued to face challenges when seeking healthcare. The VHA not only lacked knowledge in transition-related care but some transgender veterans also experienced insensitivity, harassment, and violence in some VA clinics. Findings also indicated that providers lacked a general knowledge about transgender people.One study explored the experiences of transgender veterans with the VHA.11 Utilizing a 3-year pilot program, the researchers found in the first cohort of learners (n=33) that the SCAN-ECHO was very successful in delivering training on transgender care. In fact, respondent's confidence levels to treat transgender veterans increased from 59% to 83% after completing the 14-session, 7-month program. In addition to SCAN-ECHO, the VHA has implemented a 3-year nationwide interdisciplinary e-consultation program on transgender health. The goal of this program is to give consultation to providers who treat transgender veterans at the VHA. In over a 17-month period, 303 e-consults were given to providers on transgender-related care issues to include hormones (n=125), primary care concerns (n=97), mental health (n=63), and psychotherapy (n=18).12 Overall, e-consultation complements SCAN-ECHO and other trainings offered by the VHA. With regard to total integration of these policies, much rests with the ability of large bureaucratic organizations to minimize barriers and maximize opportunities for advancement of important initiatives such as those discussed above.Regarding clinicians and staff who provide mental health and medical care services to transgender veterans, framers of Directive 2013-003 address the need for more knowledge and education about transgender health issues and specifically state that transgender veterans should be treated with dignity and respect. Professional consultation on complicated cases within Veterans Integrated Service Networks (VISN) or facilities is encouraged by the VA along with cultural awareness and sensitivity education for field staff. For instance, the VHA Transgender SCAN-ECHO  was introduced in 2011 to train VHA providers on transgender patient care. According to one study, “This program is innovative in its national scope, interdisciplinary team model, and multihub structure.”13 The type of legitimate authority found in the bureaucratic state is based on rational–legal domination, where legitimacy is derived from both legal and rational components. The legal component implies “rules or laws which are accepted by subordinates” and the rational component which suggests that “rules are effective and efficient in accomplishing specific and immediate objectives.”14 Based on rational–legal domination, an administrative staff carries out tasks and outcomes that are calculated, efficient, and predictable. The end result of bureaucratic, rational calculation is that it fails to sympathize with the human condition in all its forms. Rational action becomes the rule rather than the exception.Max Weber is well known for his sociological perspectives on bureaucracy and its effects on modern-day capitalist societies. Writing in the late 19th and into the early 20th centuries, Weber witnessed the rise of industrialization, with particular attention to institutions reflecting the modern bureaucratic state. Some characteristics of bureaucracy include an explicit division of labor with clear lines of authority, an office hierarchy, written rules and regulations, and management by rules that are emotionally neutral.15 In other words, the VA is a bureaucracy that functions with strict adherence to rules and regulations. It is part of the military industrial complex that falls under the oversight of highly bureaucratized rational authority figures. Organizations need regulations for order and efficiency; however, the human element or “subjective rights of the individual” are often lost in the bureaucratic shuffle.15 Purely rational calculation often results in irrational consequences, particularly for transgender and intersex veterans. For instance, the VHA Directive 2013-003 was passed to address the healthcare needs of transgender and intersex veterans and provides for hormonal therapy, mental healthcare, preoperative evaluation, and medically necessary postoperative and long-term care following SRS. Although a step in the right direction, there are no provisions for SRS services or reconstructive surgery; both important elements in the bigger picture of transgender healthcare.17The Department of Veteran's Affairs might be described as a ruling organization that “exists insofar as its members are subject to domination by virtue of the established order.”Powers of governance in an organization such as the VA invest authority figures such as doctors and other administrators with a set of rules that are not easily adjusted or changed. These external laws or rules give officials legal–rational authority since there is a demand for expert administration in large institutions such as the VA. Government bureaucrats receive a fixed salary and get jobs based on specific technical skills located within a well-defined hierarchical institution. Rules and official documents are binding and powerful forces against any kind of compromise. In order for the VA to provide the best healthcare for transgender veterans, authority figures at the top levels of the VA hierarchy must re-evaluate the policy limitations set forth by Directive 2013-003. Although programs such as SCAN-ECHO and e-consultation are steps in the right direction, allowing for all transition-related care would be most beneficial for transgender veterans.Utilizing Weber's theoretical perspective on bureaucracy, the researchers in this study chose to investigate the experiences of transgender veterans impacted by rational–legal domination as set forth by policies that impact their healthcare. Challenging the authority of bureaucratic decision-making, the transgender veterans in this study communicate to the reader their experiences with the VA, including suggestions on how the administration can become more accountable by supporting policies that address transgender care of veterans.A total of 22 transgender veterans were interviewed from August 5, 2012, to July 15, 2013. Participants served in the following military branches: Navy, Air Force, Marines, Army, Army National Guard, and Coast Guard. Each individual was given a pseudonym and any other identifying details were disguised or altered to ensure anonymity. Data collection included phone interviews and questionnaires. Fifteen of the participants stated that they had accessed the VHA within 1 year of the interviews. Five of the participants who submitted questionnaires had not used VHA services during the time of the interviews. Two of the participants did not comment when asked about their experiences with the VHA. Refer to Using nonprobability sampling, participants were recruited via purposive and snowball sampling techniques. The lead investigator made contact with a member of Service Members, Partners, Allies for Respect and Tolerance for All (SPART*A), an advocacy organization for lesbian, gay, bisexual, and transgender veterans and active duty service members, their families, and their allies. This individual sent out requests for participants through transgender chat rooms and other online sources. People who were interested then contacted the lead investigator through e-mail to set up a phone interview or to request a questionnaire. One participant (Brittany) elected to do both. Each participant gave his/her consent verbally. The purpose of a verbal versus a signed consent form was to ensure confidentiality and to increase recruitment because of the unwillingness of many transgender individuals to speak out about their military experiences. The use of verbal rather than signed consent was approved by a university institutional review board (IRB).NVivo, an updated version of the NUD*IST  software program, was utilized to organize the transcribed and questionnaire data. Reading through each interview, the lead investigator used NVivo to organize phrases that corresponded to specific research questions. From this analysis emerged themes that would structure the Findings section of this research project. From the 22 interviews, a total of 12 transcripts were transcribed and uploaded into NVivo. An additional 10 questionnaires were uploaded into NVivo. Examples of research questions include the following: (1) Why did you join the military? (2) Tell me about your experiences as a service member when you were actively serving our country. (3) When you were serving, how did you negotiate your gender identity within the gender expectations of the military? (4) As a transgender veteran, have you been denied healthcare or turned down for any treatment that you needed because of your gender identity? (5) What kind of policy changes do you think could be added or eliminated that impact transgender veterans and those who are currently serving? After the data were organized into themes, a node report was generated providing the researchers with the raw data used to construct the Findings section.Our data focus on the participant's experiences with the Department of Veteran Affairs. Our participants were interviewed during and after the release of VHA Directive 2013-003 by the Department of Veterans Affairs on February 8, 2013. The Directive was written to ensure that adequate healthcare is provided to transgender and intersex veterans. It is important to note that at the time of our interviews, the VA had still not addressed the issue of funding SRS and other surgical procedures; an issue for some of the veterans interviewed in our study. The data reveal that in some cases, there was confusion about what types of healthcare would be covered by the VA at the time of interviews. There were also some concerns about what might happen in the future for transgender veterans who need care at the VA. This uncertainty is a possible reflection of the transitional policies being drafted during the time frame of data collection. The comments that follow focus on transgender care and general healthcare issues.18 These negative experiences can influence whether or not transgender veterans decide to seek healthcare and continue with ongoing and preventative treatment. The VHA Directive 2013-003 has ensured that VA medical personnel are educated about the needs of the transgender and intersex population.19 Brittany (trans woman) discusses her first appointment with the VA:So I went to the VA for my first appointment with my primary nurse or whatever they call it. Actually I was sitting there in a skirt and stuff, and the lady came out and called . I wasn't about to jump up then. There were two sections so she went to the other section. So when she did that, I stood up. And when she turned back around, I kind of motioned to her. Now she wasn't disrespectful. She just wasn't trained in etiquette I guess. I was trying to find out if there were other transgender people coming through. And at that point, I think I said “Are there other people coming through like me?” And she said “You mean drag queens?” I said “No, I'm not a drag queen.” Other than that, it's been pretty nice… Back to my first visit with the nurse, I asked her if there was anything they could do about electrolysis. She said she would ask.Transgender individuals, and particularly transgender veterans, are often misunderstood by healthcare providers.According to Brittany, she does not believe the nurse was intentionally disrespectful. Had she received a minimum of training, she would have known to ask Brittany how she self-identified her gender. This behavior would align with policies detailed in the VHA Directive 2013-003. Brittany also requested information about electrolysis, a necessary procedure for transgender women that positively contributes to the transition process. Although the VA does provide some healthcare relating to gender identity, electrolysis is currently not part of the healthcare plan for transgender veterans.Anna  was the only intersex veteran interviewed for this study. She served as a Petty Officer First Class in the Navy. During her service, Anna presented as male, although her gender was often times questioned by others. According to Anna, “Intersex people typically have to deal with being the brunt of jokes because everybody can see that the boy looks more like a girl. The girl acts more like a man.” In other words, many times it is not easy to gender a person as either male or female who has an intersex condition. Anna was laid off from her civilian job and lost her health insurance, so she enrolled at the VA. During her enrollment, she was asked about the name discrepancy on the DD Form 214 (DD214), a document for identifying the veteran's status at discharge. She explained that she underwent SRS and had changed her name. The receptionist in the lobby asked loudly, “Did you go to Thailand to get the sex change?” Humiliated, Anna walked out of the VA and refused to return. She went on to say that she filed a complaint stating the receptionist needed more training to deal effectively with transgender veterans.As far as I know I'm listed in the system. I legally had my name changed to Katherine before I was discharged and the VA's records should say only Katherine. I'm sure they've got papers that say otherwise but for the most part my record was gone back through and Kenneth was scratched out with Katherine written right above it.Another veteran, Katherine (trans woman), was concerned about getting her name changed in the VA system:20 The veteran must also provide legal documentation to include either a new or amended birth certificate or passport.At the time of our interviews, transgender veterans can ask to have their sex and gender changed in the Computerized Patient Record System (CPRS); however, they must provide a letter from a physician providing specific information as outlined by the Office of Informatics and Analytics.No, I have never been turned down from the time that I began my transition. I started with black market hormones until I had found out that the Phoenix VA was providing treatment back in 2005.Whether identifying as trans women or trans men, most of the veterans in this study agreed that having access to hormone therapy was very important. At this time, the VA does provide cross-sex hormone therapy. Before getting hormone therapy, the individual must be evaluated and monitored over time to ensure positive results. When asked if she had ever been denied healthcare or treatment as it related to her gender identity, Amy (trans woman) had this to say:Before going to the VA, Steven (trans man) educated himself on the transition process. He wanted to make sure that he was prepared to make informed decisions about his care.Of course I did my research so I had read, studied, printed and highlighted the standards of care. I put together precisely what was needed for them to give me. So I got my referral to endocrinology. I went to endocrinology with everything that they needed. They didn't even have to interview me. I gave them a written, signed document that they just stick in my file so that they didn't have to bother to go through it. You know outlining what I understood about testosterone and all of that. I was on T within 3 weeks. Within 2 months I had my hysterectomy and within 6 months I had my top surgery done. Right now I'm still busy losing weight and saving money for lower surgery because of course being a vet, SRS is excluded.21 For Steven, going into the VA with knowledge about transition helped speed up the process. He stated that he was denied SRS by the VA:They will take care of everything except SRS. Actually a trans man can go get phalloplasty and then come back to the VA and get their erectile device paid for and installed by the VA because you're then just a dude who can't get it up because your transition is considered complete.As Steven mentioned above, he utilized the “standards of care” as an educational resource on transgender care. The overall goal of World Professional Association for Transgender Health (WPATH) standards of care is to provide clinical guidance for health professionals to assist transsexual, transgender, and gender-nonconforming people with safe and effective ways to achieve overall health, psychological well-being, and self-fulfillment.So for instance, if one does have genital reconstruction but there is no urethral extension, the urethroplasty that is done so that they can stand and void, the VA will provide that surgery because it's not SRS. It's just male enhancement surgery. They will put in an erection pump because you have erectile dysfunction and it enhances your life as a man. So it's interesting that they are willing to provide certain surgeries but not others. I'd like to see them at least begin to refer people out of network to surgeons who are proficient in those surgeries and cover then partially or fully depending on status of the vet.David (trans man) explained a postoperative procedure mentioned by Steven in this way:22As mentioned previously, transgender care as we know it may change in the near future since the VA is currently considering providing SRS to transgender veterans. In addition, since the time of Steven's and David's interviews, more is being offered to transgender individuals who seek to transition. Some U.S. health insurance companies now cover transgender care to include employer provided group plans, Medicare/Medicaid, and some state insurance plans.They got me some real good therapy going for my PTSD. Now that I've transitioned, they have been very good about that. They won't change your name on your military records or when you were considered female. But like all of my records now are under my male name of Pete. Because of my PTSD, they take care of my…I see a psych doc regularly and they give me my anti-depressant medications. They give me my testosterone which I was shocked that they would do that.Pete (trans man) was diagnosed with severe PTSD. He had this to say when asked about the healthcare he had received from the VA:23 Although it is not guaranteed that the Board of Corrections for Military Records will accept a correction, this procedure is currently the best way to request a name change. At the time of the interview, this option was not available for Pete. Debbie (trans woman) stated that the Veterans Benefits Administration (VBA) was “dragging their feet” to get her on disability.The Department of Defense (DoD) allows name changes in the Defense Enrollment Eligibility Reporting System (DEERS) and on the Military Discharge Record or DD214 by submitting DD Form 149, which is an application for correction of Military Record.I was in from 2005 to July 2008. Honorably discharged and all of that. I did my time and got out. I was a paratrooper in the military. I've got a lot of jumps under my belt and I was over in Iraq for 15 months. I did 20 or so jumps. I got blown up like three times or something. So I'm not going in to scam the system or any type of thing. And I know a lot of people do but I have a harder time turning my neck. I can't touch my chin to my shoulder like not even close. I can't touch my chin to my chest. I've lost a lot of motion in my neck. I have the ringing in the ears and went in for PTSD also. I have a lot of issues I guess. So I'm just trying to work out a lot of things and seeing what happens. So it's just one of those things where I'm fighting them and they are just dragging their feet about it.As David stated, regardless of transgender status, it can “take months to years” to get on disability. At this time, the backlog in disability claims continues to be addressed by the VA in an effort to provide care for all veterans.Well you know one of the lies that they told us when we went in back in 1960 was you serve us for 4 years and we'll take care of you for the rest of your life. And that has been not a complete truth.What these experiences illustrate is that regardless of gender identity, transgender veterans need access to healthcare. As Brittany (trans woman) mentioned in her interview regarding the DoD:It is evident that more needs to be done to ensure that our transgender and intersex veterans are provided adequate healthcare by the VA that includes SRS and other surgeries needed to complete the transition process. As David (trans man) mentioned above, “it's interesting that they are willing to provide certain surgeries but not others.”No in fact I keep saying that I need to go register at the VA because I know they need the body count to get their funding, to keep their funding. But I just haven't registered at the VA so…And my husband is 11 years older than me and he has not registered at the VA either so we need to both go there.Another trend found in the data is that some transgender veterans either had not enrolled or chose not to seek healthcare at the VA. For Julia (trans woman), it was just a matter of taking the time to enroll. When asked if she used the VA, she had this to say:I haven't been brave enough to venture into that yet but I think I'm getting close. I've got an appointment on Friday so we will see. Paying for hormones out of pocket is getting very expensive and I think I'm going to see what the VA can do for me.Katy (trans woman), on the contrary, felt apprehensive about the VA. However, because hormonal therapy was becoming too much of a financial burden, she finally made an appointment to enroll:I don't use the VA healthcare system. All my private physicians know about my transition and I've never had anything but positive reactions with no change in my healthcare quality.Donna (trans woman) stated “I've never sought treatment at the VA related to my gender identity” while Paul (trans man) said “I don't have anything to do with the veteran's administration and that's my choice.” Michelle (trans woman) stated that she did not use the VA because “I do not trust them.” Jenny (trans woman) chose to use her own private physician rather than use the VA for her healthcare:18 Programs such as SCAN-ECHO and e-consultation are just two ways that the VA is educating providers about transgender care. However, some of the comments made by our participants illustrate the necessity of ensuring that programs such as these continue to be developed to provide a comprehensive approach to transgender care needs within the VA.What this dialogue illustrates is that some transgender veterans are not getting all of the treatment that they need and they do not appear to have full confidence that the VA is a reliable source of healthcare. There is a need for more community-based research that brings together the transgender community and VA staff so as to “diminish prejudice, reduce stigmatization, and encourage transgender veterans to obtain needed healthcare.”24 As some of our participants illustrate, getting the transgender care can still be a struggle for them. When asked what policy changes should be added and/or eliminated, our participants made several recommendations. Amy (trans woman) suggested that full treatment, to include SRS, would help to reduce anxiety.When it comes to the issue of gender identity, the question remains as to why the VA provides some treatments for transgender veterans while excluding others. Whether or not transgender people should be allowed to have SRS and other transition-related procedures has been questioned not just by the VA but began back in the 1960s when Johns Hopkins became the first academic institution to perform SRS.I think it would behoove them (the VA) to provide for reassignment surgery because the cost would outweigh the costs of treating the individual for a number of suicide attempts and mental health issues.Robin (trans woman) commented that “retired personnel should have insurance coverage and access to the full spectrum of transgender healthcare” through Tricare while Jenny (trans woman) mentioned that it was important to “allow transgender vets the same access and rights to services as all other vets.” These recommendations indicate that many of our participants were concerned that all treatments to include SRS be made available to transgender veterans. Otherwise, inadequate healthcare will continue to be an issue.Transfriendly doctors and staff who can address an array of medical issues were important for participants in this study. Some of our respondents elected not to utilize the VA for their healthcare needs and others have been less than satisfied with the care they have received in that system. The general consensus among the people in our study was that medical practitioners who are familiar with and open to working with transgender people is a key factor in seeking out treatment from the VA or other healthcare institutions. When a transgender patient sees a doctor who is not informed or does not care about their unique needs, improper treatment may result in ongoing health consequences. For military veterans, unequal medical treatment is not only unethical but it also amounts to a broken promise to people who served their country.10The average age of participants in our study was 50. Their collective years of service were 237 and most had at least some college education. There was one intersex respondent, 16 trans women, and 5 trans men. Although the sample is too small to generalize to the broader transgender veteran population, the comments we received indicate that the VA should be an important health resource for anyone who has served in the military. Even though not all transgender veterans in our sample utilize the VA for healthcare, the data we collected reflect many of the same concerns relative to comprehensive and inclusive medical care for transgender veterans in other similar studies.In ongoing attempts to address concerns of transgender and intersex veterans, the VHA published Directive 2013-003 to establish policy regarding delivery of healthcare to transgender and intersex veterans enrolled in the VA healthcare system. Although this was a step in the right direction, more must be done to ensure that veterans get the healthcare they need. Along with denying SRS, the VA should also consider other treatments necessary to complete the transition process such as electrolysis and reconstructive surgery.25The limitations of the study are attributed to the use of purposive and snowball sampling techniques, which do not require random sampling; therefore, the findings cannot be generalized to all transgender veterans. More specifically, this sample of transgender veterans is not representative of the transgender population in terms of race and ethnicity, class, disability, and age due to the small sample size. Since the time of our interviews, another important limitation that should be mentioned is that our participants were interviewed from August 5, 2012, to July 15, 2013. Although VHA Directive 2011-024 was in place when some of the interviews were conducted, the new VHA Directive 2013-003 was released by the Department of Veterans Affairs on February 8, 2013, to ensure that adequate healthcare is provided to transgender veterans. Therefore, the data reflect a time before and shortly after the 2013 directive was released. Since the time of our interviews, more has been done by the VHA to improve healthcare services for transgender veterans.26 If approved, this new rule would replace VHA Directive 2013-003, alleviating some of the issues faced by many of the participants in our study.However, as mentioned above, there are still no provisions for SRS services or reconstructive surgery; both important elements in the bigger picture of transgender healthcare. This may change in the near future since the Department of Veterans Affairs is currently considering providing SRS to transgender veterans.16 Even though Directive 2013-003 does not currently support the full spectrum of transgender care for those seeking SRS and other surgical treatments necessary to complete transition, the VA has made some significant headway in recent years in treatment protocols and policies for transgender patients. For instance, there is now Veterans Care Coordinators located at VA facilities that support LGBT programs. The Department of Veterans Affairs lists the websites of 28 VA facilities with LGBT programs in several states.27 Providing such programs ensures that LGBT veterans have a supportive and safe environment when seeking the care that they need. Two additional VA clinics that specialize in transgender care have also recently opened in Cleveland, Ohio, and Tucson, Arizona. Like the larger clinics and medical centers in the VHA system, these clinics do not provide SRS and other surgical treatments related to transition. Therefore, bureaucratic powers of governance must continue to re-evaluate policy limitations set forth by Directive 2013-003. While the clinic in Tucson serves >100 transgender veterans, some VA hospitals continue to turn away transgender veterans. Policy changes can and will result in better healthcare along with incentives for transgender veterans to use VA facilities.28Medical doctors and other healthcare providers agree that transgender individuals who receive the full spectrum of services experience better quality of life than those who do not.29 As mentioned earlier in this article, SCAN-ECHO and e-consultation have been very successful in delivering training on transgender care to practitioners. The VA also focuses on outreach to LGBT veterans, which is also a step in the right direction.30 At this time, the Department of Veterans Affairs is considering providing SRS to transgender veterans. This new “rule” would remove barriers to transition-related care. The review process could take up to 22 months.26 If approved, this new rule would replace VHA Directive 2013-003, alleviating some of the issues faced by many of the participants in our study.The VA also provides trainings to VHA staff regarding LGBT healthcare.31 However, we suggest that one key area of importance should continue to focus on how mental health and medical providers and ancillary staff are trained to interact with and provide care for their transgender patients. As our participant Donna (trans woman) so eloquently states, the VHA “needs to have clear cut policies to ensure that trans veterans are receiving fair treatment and are being treated with dignity and respect.” With 84% (96 of 114) of all VHA clinics achieving Leadership Status in the 2016 Human Rights Campaign's Healthcare Equality Index (HEI), perhaps the end of bureaucratic, rational calculation is in sight.31 As our data illustrate, transgender veterans have a great deal of knowledge and experiences to share with the VHA that can contribute to shaping current and future polices as these relate to their care. Whenever change occurs, especially in large bureaucratic organizations such as the U.S. military, it is particularly important for social researchers to continue collecting and recording empirical data in an effort to ensure that progress continues and does not stall.With regard to further research, as military organizations begin to re-evaluate their policies, more transgender service members will come out of the shadows. This affords an opportunity for increased dialogue with researchers about personal experiences with the VA and the military in general. Considering the time frame in which our data were collected, more has been done to improve VHA relationships with LGBT veterans. The VHA has made and continues to make system changes that advance LGBT inclusiveness and clinical competence.


Routine neonatal circumcision remains a controversial topic. The most recent Canadian Paediatric Society does not recommend routine circumcision of every newborn male . And altThe association between lack of circumcision and penile cancer has been well documented . A meta-Despite its rarity, there are several common factors that have been implicated to cause penile cancer. Factors resulting in phimosis, balanitis, and smegma have been associated with penile cancer in a meta-analysis . This haAs a driving factor for penile carcinogenesis, the prevalence of HPV induced penile cancer has been shown to be approximately 50% of penile malignancies worldwide . CircumcPrevention of HPV infection and subsequent penile cancer risk can also be accomplished with vaccines. There are two prophylactic HPV vaccines have been developed that can play an important role in the prevention of HPV transmission. This includes the quadrivalent and 9-valent HPV vaccines (Gardasil and Gardasil Penile carcinoma can be preventable through advocating daily hygiene and HPV prevention. Although circumcision can help reduce risk factors for penile cancer development, this does not necessarily warrant a requirement for circumcision as a neonate given the rarity of disease and alternative strategies in prevention. Through improved efforts in modifiable behaviors and implementation of HPV vaccination, this can curb penile cancer risk until the patient can make an informed decision regarding circumcision later in life.


Human papillomavirus (HPV), especially HPV16 genotype, is associated with oropharyngeal squamous cell carcinoma (OPSCC). We aim to determine the prevalence and characterize the high-risk (HR)-HPV genotypes in head and neck SCC (HNSCC) in a South-East Asian multi-ethnic society in Singapore and examine its prognostic significance.digene HPV genotyping RH test). P16 protein overexpression was identified using immunohistochemistry (IHC). Statistical correlation between clinical outcomes were performed between HPV-positive and negative HNSCC patients.159 HNSCC archival tissue samples were retrieved and tumour DNA was screened for 18 HR-HPV genotypes using a PCR-based assay . HPV-DNA PCR assay yielded a high sensitivity (96%) but low specificity (11%) when compared to p16 immunohistochemistry as the reference standard.Six HR-HPVs  were detected in 90.6% of HNSCC; and 79.9% had multiple HPV genotypes detected. HPV31 and HPV45 were the most prevalent ; and HPV16 was predominantly found in OPSCC (p = 0.005); and p16-positive OPSCC exhibited improved overall survival compared to p16-negative OPSCC (p = 0.022). Similarly, smoking and alcohol consumption were poor prognostic factors of overall survival  in OPSCC patients.P16-positive HNSCC was predominantly observed in OPSCC (73.7%; HR-HPVs were identified in 90.6% of HNSCC patients using the HPV-DNA PCR assay. This test had a poor specificity when compared to p16 IHC; making it an unreliable detection technique in selecting patients for radiation dose de-escalation treatment protocol. P16-positive tumor was predominantly found in the oropharynx these patients demonstrated better overall survival than those with p16-negative OPSCC. There is firm evidence that human papillomavirus (HPV) is implicated in the pathogenesis of a subset of patients with head and neck squamous cell carcinoma (HNSCC) who are typically younger and without the traditional risk factors of smoking and alcohol consumption –4. Most There are approximately 18 oncogenic HPV genotypes which have been implicated in the causation of cancers in humans , 6. The This study aims to determine the prevalence and characterize the HR-HPV genotypes in HNSCC in Singapore, which constitutes a multiethnic, urban South-East Asian society; and to explore the prognostic implication of HPV detection in patients with HNSCC in this population.p < 0.001). Additionally, multiple co-existing HPV genotypes (2 and more HPV genotypes) were detected in 79.9% of the tumor specimen and they were equally detected across all subsites. In OPSCC, 74.2% of tumor specimens demonstrated 2 or more HPV genotypes and only 19.4% had only one HPV strain detected .159 HNSCC patients with sufficient archival tumor specimen were accrual for HPV detection. The distribution of HPV subtypes according to the head and neck subsites is summarized in Table p = 0.005). Of the remaining 5 cases with p16-positive tumor, 2 were detected in oral tongue SCC; 1 from laryngeal SCC and the remaining 2 cases were from primary sino-nasal SCC. All p16 IHC-positive cases were HPV-positive except for one patient with OPSCC who was HPV PCR-negative. Using p16 IHC as a reference test for HPV detection, the sensitivity and specificity of HPV-DNA PCR detection were 96%, and 11%, and 93% and 5.9% for HNSCC and OPSCC, respectively.There was strong concordance of p16 IHC detection with the presence of HR-HPV detected using the PCR assay across all sites of HNSCC (94.7%) Table . RepreseSince p16 IHC was predominantly identified in our OPSCC cohort, comparison of clinical features between p16-positive and p16-negative OPSCC patients was performed. There were no statistical differences in age, gender, T- or N classification and overall AJCC stage among patients with p16-positive or p16-negative OPSCC Table .p = 0.022, Hazard Ratio = 0.21; 95% Confidence Interval (CI) = 0.05–0.84)   compared to p16-negative patients (92.9% versus 58.8%) (log rank test: ) Figure . Additiodigene HPV genotyping RH test kit because it detects a panel of HR-HPV genotypes which is associated with cervical cancer [In this study, we have chosen the l cancer , 6, 12. l cancer . Therefodigene HPV genotyping RH test kit, the overall prevalence of HPV-associated HNSCC was 90.6%. Six out of the 18 HR-HPV genotypes were detected  in our cohort of HNSCC patients. Among these HPV genotypes, only HPV16, HPV18, HPV31 and HPV45 had been previously reported in HNSCC. The detection of these HR-HPV genotypes is consistent with the epidemiology data of HPV-associated cervical cancer where HPV31 and HPV45 have been reported, but HPV16 and HPV18 remain the predominant genotypes [Using this enotypes –17. The digene RH test kit has a sensitivity limit of detection for HPV31 and HPV45 at 10 and 23 copies/PCR, respectively; compared to the PapilloCheck high-risk HPV kit  which has a higher detection level of 50 and 90 copies/PCR. Additionally, the type of primer employed by the different commercial test kits may play a role in the detection rate of HPV. The GP5+/6+ used in our study had a higher sensitivity than the MY09/11 primer set in detecting HPV45 [The ng HPV45 . Lastly,ng HPV45 .digene RH test kit only targets the L1 region of the HPV genome, it does not have the capability to differentiate integrated versus episomal forms of the virus, and hence may not identify biologically active HPV transcripts. Therefore, some authors have advocated the use of transcriptionally active HPV-E6/E7 mRNA expression as the gold standard of HPV detection [To our knowledge this is the first report of HPV56 and HPV68 being associated in HNSCC. Interestingly, HPV56 and HPV68 have been reported in cervical smears of healthy Singaporean women although these genotypes are not seen in the malignant counterparts in Singapore , 15, 17.etection , 23. To etection , 25. Theetection . TherefoMultiple HR-HPV genotypes are typically detected in our HNSCC cohort with HPV31 and HPV45 genotypes being the most prevalent. HPV16 genotype is hardly detected in non-oropharyngeal sites and is primarily detected in OPSCC. This information is compatible with our Western cohorts where HPV16 genotype is the commonest genotype detected in OPSCC. Specific to the Asian context, our findings differ from the HPV genotype expression in China where HPV18 and HPV16 exists in equal proportions among patients with OPSCC . BesidesP16 immunohistochemistry (IHC) detection is a useful surrogate marker for HPV-related oncogenic pathway as it represents active HPV oncogenic activation. In the HPV-driven oncogenic pathway, HPV integrates into the host genome and increases HPV-E6 and E7 gene transcription. E7 onco-protein specifically results in phosphorylation of the retinoblastoma tumor suppressor which releases E2F transcription factor. This in turn increases p16 expression –31. TherAdditionally, using p16 IHC as a surrogate in OPSCC allows easy clinical translations as it is much easier to perform and less time consuming compared to HPV-DNA PCR assay. As with other studies, p16-positive OPSCC patients have consistently demonstrated improved clinical outcomes compared to patients with p16-negative OPSCC . This obdigene test kit which may not distinguish HPV16 from HPV31 and HPV18 from HPV45; since these paired HPV genotypes are genetically closely related [One limitation in our study was the relatively small sample size of OPSCC. Therefore, we are unable to correlate the presence of HPV16 with p16 IHC expression. The correlation between HPV16 and p16 was 42.8% (6/14) in our study; as compared to more than 80% reported in other studies , 35. Fur related . Be thatIn conclusion, our HNSCC cohort from an urban, multi-ethnic, South East Asian society, demonstrated a high prevalance of HR-HPVs  using a commercial PCR-based test kit. HPV16 and P16 IHC expression were predominantly detected in the oropharyngeal subsite. Patients with p16-positive oropharyyngeal HNSCC exhibited a superior overall survival compared to those with p16-negative HNSCC; making p16 IHC a convenient surrogate marker for HPV detection in HNSCC. Routine clinical use of commercial HPV DNA based test kits should not be utilized given the high percentage of false positive results.digene HPV Genotyping RH test amplification kit, Germany) followed by a reverse hybridization procedure . P16 was detected using immunohistochemical staining of the tissue slides . Approval by our institution review board was obtained prior to the commencement of the study.Tissue blocks of patients diagnosed with primary HNSCC were retrieved from the archives of the Department of Pathology at the National University Hospital Singapore. These tissue blocks were verified to be squamous cell carcinoma by one experienced pathologist (FP) and the DNA was extracted from the tissue blocks using the QIAamp DNA Formalin-Fixed, Paraffin-Embedded (FFPE) tissue kit as per manufacturer protocol . One hundred and fifty-nine tissue blocks contained adequate DNA and were accrued for HPV detection. HPV was detected using a Polymerase Chain Reaction (PCR)-based amplification assay  system for 30 mins. Slides were visualized using Ventana Ultra View DAB kit with DAB  for 8 mins. Tissues sections were counterstained with hematoxylin for 2 mins. The specimens were considered to be p16 positive when at least 80% of the tumour cells showed both cytoplasmic and nuclear p16INK4a staining.For the immunohistochemistry analysis, an anti-human p16INK4a monoclonal antibody  was used. In brief, tissue sections were cut into 3-μm slices and deparaffinized. Antigen retrieval was done with pH9 CC1 buffer  for 36 mins at 100°C. The tissues sections were incubated with peroxidase blocking reagent for 30 mins and then with 10% goat serum for 60 mins. The tissues were then stained with anti-p16INK4Genomic DNA was extracted using Qiagen QiaAmp DNA FFPE kit . Briefly, 5–10 μm thick tissue sections were deparaffinized with xylene and residual xylene were extracted by ethanol. The tissue was then lysed and digested at 56°C followed by 90°C for 1 hr. Cells were lysed and DNA was extracted using DNA column. DNA was then eluted and stored at −20°C until further testing.digene RH test was used to detect 18 carcinogenic HR-HPVs. These 18 HR-HPVs are HPV 16, 18, 26, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66, 68, 73 and 82. The test comprised of an amplification and detection phase. Briefly, 10 μl of the extracted DNA was mixed with 25 mM MgCl2 and the commercial master mix solution containing GP5+/6+ primer sets and an internal control primer set. L1-segment PCR amplification was carried out with cycle parameters as follows; initial activation step, 94°C for 5 mins; 3-cycling steps comprising denaturation at 94°C for 20 sec; annealing at 38°C for 30 sec; extension at 71°C for 80 sec. This 3-cycling-step was repeated 40 times using PE Applied Biosystem GeneAMP PCR system 9700 machine .In this study, the After GP5+/6+ PCR amplification, detection was carried out according to the kit protocol. In brief, 10 μl of the amplicon was denatured and hybridized with the specific immobilized probes on the nitrocellulose strip in parallel lines, including an internal control- human β-globin amplicon. The nitrocellulose strip was then incubated with a conjugate and a substrate and produced a color band for visualization and identification of the specific HPV strain.p values < 0.05 were considered statistically significant.The statistical association between HPV-DNA PCR positivity, p16 expression and clinico-pathological was evaluated using Chi-square Fisher’s exact test . Clinical outcome measures include overall survival and disease specific survival. Overall survival was defined as time of diagnosis to death from any cause or last follow-up and disease-specific survival was defined as the time to death from cancer or treatment related complications. The survival outcomes were estimated using Kaplan-Meier and survival curve were compared using Mantel-cox log-rank tests (GraphPad Prism version 6). All calculated


Large numbers of post-deployment U.S. veterans are diagnosed with post-traumatic stress disorder (PTSD) and/or traumatic brain injury (TBI), leading to an urgent need for effective interventions to reduce symptoms and increase veterans’ coping. PTSD includes anxiety, flashbacks, and emotional numbing. The symptoms increase health care costs for stress-related illnesses and can make veterans’ civilian life difficult.We used a randomized wait-list controlled design with repeated measures of U.S. military veterans to address our specific aim to test the efficacy of a 6-week therapeutic horseback riding (THR) program for decreasing PTSD symptoms and increasing coping self-efficacy, emotion regulation, social and emotional loneliness.n = 15) or a wait-list control group (n = 14). The wait-list control group experienced a 6-week waiting period, while the horse riding group began THR. The wait-list control group began riding after 6 weeks of participating in the control group.Fifty-seven participants were recruited and 29 enrolled in the randomized trial. They were randomly assigned to either the horse riding group .The PCL-M as well as other instruments including, The Coping Self Efficacy Scale (CSES), The Difficulties in Emotion Regulation Scale (DERS) and The Social and Emotional Loneliness Scale for Adults-short version (SELSA) were used to access different aspects of individual well-being and the PTSD symptoms.P ≤ 0.01) as well as a statistically and clinically significant decrease after 6 weeks of THR (P ≤ 0.01). Logistic regression showed that participants had a 66.7% likelihood of having lower PTSD scores at 3 weeks and 87.5% likelihood at 6 weeks. Under the generalized linear model(GLM), our ANOVA findings for the coping self-efficacy, emotion regulation, and social and emotional loneliness did not reach statistical significance. The results for coping self-efficacy and emotion regulation trended in the predicted direction. Results for emotional loneliness were opposite the predicted direction. Logistic regression provided validation that outcome effects were caused by riding longer.Participants had a statistically significant decrease in PTSD scores after 3 weeks of THR (The findings suggest that THR may be a clinically effective intervention for alleviating PTSD symptoms in military veterans. Therapeutic horseback riding (THR) is defined as a horseback riding program in which the primary goal is rehabilitation , 2. ThisOur study was designed using the conceptual framework of social cognitive theory (SCT). SCT explains psychosocial determinants of behavior in terms of triadic reciprocal causation  . In thisAccording to Bandura, in SCT, a major factor of motivation, affect, and behavior is self-efficacy. Persons have a level of confidence, known as perceived self-efficacy, that influences behavior . The actBandura  noted thPTSD is an anxiety disorder that occurs after exposure to a life-threatening event or injury . PTSD isPTSD has been associated with poor quality of life and increased use of health care services, and a variety of co-morbid physical and psychological conditions ; most noResearch suggests that a common coping response to PTSD among veterans is excessive alcohol use –15. FurtAdditionally, fears of stigma may prevent veterans with PTSD from admitting to symptoms, seeking assistance, or following medical advice. PTSD and chronic pain are commonly comorbid, with each reinforcing and exacerbating the effects of the other . VeteranCognitive behavioral therapy (CBT) is often a recommended treatment for veterans , 23. CogDespite empirical evidence of the prevalence and potential negative impact of PTSD on veterans, research that examines innovative interventions is scarce. Experts have advocated for research on complementary and alternative therapies  –32. AnimClinically, THR is a standardized horse riding program for people with disabilities in which the primary goal is their rehabilitation , 2. THR Physical benefits of THR include improved sitting posture , motor fThere are limited studies involving THR in the treatment of PTSD or TBI. One important case study of a 44-year-old veteran with a spinal cord injury demonstrated both functional improvements, including increased regular PA, along with increases in self-esteem, self-control, and sense of independence . VeteranHorses and THR have been previously used in treating PTSD; however, there are no randomized controlled trials studying the effectiveness of horses in reducing levels of PTSD . THR mayThe study had the approval of the VA Research and Development Committee at the Harry S. Truman Memorial Veterans Hospital, the University of Missouri Health Sciences Institutional Review Board and the Animal Care and Use Committee of the University. All participants completed the VA Research Consent Form, which had been approved by the Health Sciences (HS) Institutional Review Board(IRB). Fifty-seven veterans were assessed for eligibility. Nineteen veterans were excluded . Our primary means of recruitment occurred through letters and postcard invitations. Two invitations were mailed to veterans identified through VA electronic medical records as having met the inclusion criteria of a diagnosis of PTSD, or PTSD and traumatic brain injury (TBI), and who lived within a 50-mile radius of the THR sites. Veterans were also recruited through referrals from VA clinicians and by advertising the study throughout the VA hospital. These veterans contacted the study office to volunteer to participate.Other inclusion criteria were: age 18 years or older, no longer in active military service (including reserves), weight of 220 pounds or less, able to walk at least 25 ft without the assistance of a person, and willing to interact with and ride a horse. The weight limit of veterans in our inclusion criteria was determined because the horses could not accommodate heavier participants.All horses working at the riding center were eligible to participate in the study if they were able to accommodate a veteran of up to 220 pounds. The horses that worked in our study were selected by the Professional Association of Therapeutic Horsemanship (PATH)-certified riding instructor for their fitness and experience of being ridden by adults. As part of the ethics approvals, the VA Research and Development Animal Component of Research Protocol (ACORP) involved a visit by a VA-affiliated veterinarian to the riding centers to verify the welfare and husbandry conditions for the horses.n = 5; injuries, n = 2; logistic issues, n = 2). All participants were aware that they would be assigned to one of the two groups prior to providing informed consent. They were randomly assigned to either the riding group (n = 15) or a wait-list control group (n = 14) based on their identification number . When they converted to the treatment group, another set of baseline data was collected from them. This increased the baseline treatment group data to 23 participants. For the 3-week data, there was also data from 23 participants. However, in the 6-week data, due to attrition, there was data from only 19 participants. This waitlist control design has been successfully employed in an equine program research with 131 adolescents in an 11-week equine facilitated learning program . I am always happy around him and I think he responds well to me also.” It may be that loneliness scores at six weeks were related to the veterans’ anticipation of the completion of the program.Interactions between the veterans and the riding center and study staff were positive. Other investigators have advocated for an individualized approach to selecting “talkative volunteers” or less talkative volunteers as a match with each veteran’s needs, which may enhance veterans’ THR experience . We atteAt the beginning of our study, some veterans expressed reluctance to participate on receiving our first invitation letter. They were more receptive to the subsequent postcard that we sent with the study logo on it. One gentleman who was a Vietnam war veteran said that he did not want to participate, but his wife encouraged him to come. However, after his first session , he thought that it was too bad to have to wait 2 weeks to do this again. This veteran not only completed the study, he expressed interest in continuing to volunteer at the riding center after completion of the study.Veterans expressed interest in participating in the study to try something new or rekindle a childhood experience. Many had ridden horses as children and recalled this fondly. Fortunately, veterans who wanted to continue THR were able to do so after the study was completed. This occurred via prescriptions for THR facilitated by VA recreation therapists or when the veterans volunteered at the therapeutic riding centers. Volunteering has been found to be a meaningful activity for veterans, particularly those who served in combat .Accessibility to riding centers may be an issue for veterans who wish to participate in THR. One veteran said “I had to drive an hour to and from the horse center”, and we know this was one of the challenges of the program. THR may be more accessible to rural veterans who may have to travel long distances to reach VA treatment programs. THR is clearly not a replacement for conventional therapies used to treat PTSD, but as a complementary therapy, riding centers may be a readily accessible resource to veterans in rural areas.Our sample size was small, which limited our power to detect changes in the dependent variables. Nevertheless, we did find statistical and clinical significance in lowering PTSD symptom levels. The sample size was limited by the local VA requirement that we could only recruit participants being treated at the VA through which the study was approved. Originally, we partnered with two THR centers in the St. Louis area. Only 4 of the 38 participants who were treated at the Columbia, Missouri VA lived close enough to these two riding centers to be able to travel there.The study was logistically complex to implement because of the need to balance VA requirements and approvals, the riding center schedules, university calendars, and the busy lives of the participants. One previously mentioned veteran drove 1 h, while others drove 40 min to attend the classes, which occurred during normal business hours to accommodate the riding center. Three volunteers were needed per veteran . The volunteers are essential to the daily operation of the riding center, increasing the complexity of scheduling. We required an Occupational Therapist and a PATH-certified riding instructor to administer the detailed riding curriculum. We also had study staff present to monitor fidelity to the riding curriculum, oversee data collection, and monitor participant safety. The staff also phoned the veterans each week to confirm their attendance at each session or apprise them of changes in class schedule due to inclement weather.We did not use a longitudinal follow-up. It would have been helpful to identify to what extent the PTSD symptoms remained lower after time passed since the riding program ended. Furthermore, the length of our THR program may have been too short. Lanning and Krenek  used a 2Those planning THR programs would do well to address some of the lessons we learned from our study. While our findings showed beneficial outcomes in PTSD at the 3-week measurement, longer participation (six weeks) was beneficial for self-efficacy and emotion regulation. We identified steps that could be taken to minimize veterans’ attrition. For example, assisting with transportation to the riding center and providing child care during the classes may be beneficial. Climate controlled riding centers would maximize comfort for participants, volunteers, staff and horses, while minimizing the likelihood of having to cancel a class session. Horses able to carry riders weighing more than 220 pounds would increase the numbers of veterans who can participate. In addition, provision of THR class scheduling options may decrease the number of classes missed by veterans due to scheduling conflicts.Our findings have honed the existing knowledge base on THR, a beneficial intervention for veterans with PTSD, by identifying a clinically meaningful dosage of THR. A 3-week THR program was effective, and a 6-week program produced clinically significant outcomes in PTSD levels. Older veterans, such as the majority in our sample, may have been diagnosed with PTSD decades ago; THR may be particularly promising for them. It may be important for health systems to recognize such promise by supporting THR as a reimbursable complementary therapy.


Human papillomavirus (HPV) vaccination coverage in the Netherlands is low (~60%) compared to other childhood vaccinations (>90%), and even lower among ethnic minorities. The aim of this study was to explore the possible impact of ethnicity on the determinants of both HPV vaccination intention and HPV vaccination uptake among parents/guardians having a daughter that is invited for the HPV vaccination.In February 2014, parents/guardians living in Amsterdam were invited to complete a questionnaire about social-psychological determinants of their decision making process regarding the HPV vaccination of their daughter and socio-demographic characteristics. This questionnaire was sent approximately one month before the daughter was scheduled to receive her first HPV vaccine dose. Their daughters’ HPV vaccination status was retrieved from the national vaccination database. We distinguished four ethnic groups: Dutch (NL), Surinamese, Netherlands Antillean, and Aruban (SNA), Middle-Eastern and North-African (MENA), and Other. To assess the impact of determinants on both intention and uptake, linear and logistic regression analyses were used respectively. Missing data were imputed using multiple imputation by chained equation.2:0.56) and lower in the other ethnic groups (pseudo-R2 varied between 0.23 and 0.29). The lower explained variance can be attributed to the relative large proportion of participants with a positive intention that finally did not go for vaccination in the SNA-group (11%) and MENA-group (30%). Explained variance (R2) of intention varied between 0.66 and 0.77 across ethnic groups, and was best explained by the proximal social-psychological determinants. The strength of association of these determinants with both intention and uptake were largely similar across ethnic groups.In total 1,309 parents/guardians participated (33% participation rate). In all groups we found the mothers’ intention to be the strongest predictor of their daughters’ HPV vaccination uptake. Explained variance of uptake was highest in the NL-group  contains supplementary material, which is available to authorized users. The national immunization program (NIP) of the Netherlands is a voluntary and free-of-charge program with an overall high childhood vaccination uptake that exceeds 90% . HoweverIn previous studies investigating HPV vaccination acceptability in the Netherlands, common predictors of HPV vaccination intention appeared to be attitude, concerns about vaccination effectiveness and safety, feelings of ambivalence towards HPV vaccination, anticipated regret, and normative beliefs , 10, 11.In the Netherlands the bivalent HPV vaccine (Cervarix) is offered since 2009 by the National Immunization Programme (NIP). In 2014, the vaccination schedule was reduced from three to two doses in line with guidelines of the European Medicine Agency. All girls are invited for HPV vaccination in the year they turn 13 years old. Girls are invited to come to a large public venue designated by the Public Health Service (e.g. sports centers and child-and parent centers). During this vaccination moment no personal consultation is offered and no consent of the parent is necessary to obtain the HPV vaccination.All parent/guardians with a daughter born in 2001 living in the district of the Youth Health Service of the Public Health Service of Amsterdam were invited for this study (2014). They were selected from a database of the Youth Health Service of the Public Health Service of Amsterdam which contains the most up to date information of all girls residing in Amsterdam. The selection of the girls eligible for this study was done one month before they received the invitation of the HPV vaccination. Parents/guardians were able to participate only once. A computer tablet was raffled among participants. The study was approved by the Ethics Committee of the Academic Medical Centre (W2013_257).Parents/guardians were invited to complete a questionnaire approximately one month before their daughter was scheduled to receive her first HPV vaccine dose ). Based on the country of birth of the parents/guardians, the research assistants were matched to the language presumably spoken by the parents/guardians. The assistants attended a comprehensive training in which they were instructed about: (i) the general background of the HPV vaccination, (ii) what information they were allowed to provide to the parents/guardians in order to not influence parents’/guardians’ responses to the questionnaire, (iii) the content of the questionnaire, and (iv) relevant other logistic aspects concerning data collection and the HPV vaccination of their daughter. The assistants called parents/guardians to explain the rationale behind the study, to offer assistance with completing the questionnaire if necessary, and/or to remind them in case of an initial non-response. If parents asked information regarding the HPV vaccination, the research assistants only gave the information that was provided in the invitation letter for the HPV vaccination and the information leaflet of this study .. [The questionnaire was based on a questionnaire initially developed by van Keulen et al.  which wa. , Social . , and the.  constitun = 109), Turkish (n = 67), and Egyptian (n = 21) ethnicity. The category Other consisted mainly of Indonesians (n = 35), Ghanaians (n = 32), and Germans (n = 15).The ethnic background of participants was based on country of birth of both parents as described previously . In shorConsent to retrieve the vaccination status of the daughter was obtained by posing a separate question asking whether the parent/guardian agreed if we would retrieve the HPV and childhood vaccination status of their daughter from Praeventis. Praeventis is a national database for monitoring childhood vaccinations in the Netherlands . HPV vacproximal determinants versus distal determinants.According to their theoretically expected cause-effect relation, socio-psychological determinants of HPV vaccination intention and HPV vaccination uptake were subdivided into Proximal determinants are theoretically expected to impact the parents’/guardians’ intention most directly. These were: general attitude, vaccination-related beliefs, negative outcome expectancies, risk perception (e.g. perceived susceptibility when not vaccinated against HPV), anticipated regret, perceived relative effectiveness of the HPV vaccination, subjective norms, descriptive norm, and self-efficacy.distal determinants on the parents’/guardians’ intention are expected to be indirect. These were: knowledge, confidence in authorities, ambivalence about the HPV vaccination, habit strength, information processing, amount of information processed, subjective evaluation of the received information about the HPV vaccination, past experience with cervical cancer (self or close relative), and socio-demographic characteristics. Socio-demographics included age, gender, educational level, religion , and ethnicity. For analyses religion was dichotomised into religious versus non-religious.The impact of See Additional file Response rate was explored by country of birth of the mother of the girl invited for the HPV vaccination; the country of birth of the mother was grouped into the same four groups as used for ethnicity of the participant. We compared HPV vaccination status, HPV vaccination intention, past childhood vaccination status, and socio-demographics by ethnicity. Descriptive analyses were only computed for complete cases.Bivariable and multivariable analyses were based on multiple imputed datasets. We used multiple imputation to account for missing data . The perp < 0.25. To obtain a parsimonious model, backward selection was executed during each step in which a new set of determinants was offered to the model (p < 0.05). Multivariable analyses with HPV vaccination intention as the criterion variable were executed according to the same theory-based order. In both final multivariable models we tested for interaction effects between ethnic group and each determinant (p < 0.05). Statistical analyses were performed in Stata 14 [HPV vaccination uptake and HPV vaccination intention were both used as criterion variable (i.e. outcome variable). Determinants of these two criterion variables, HPV vaccination intention and HPV vaccination uptake, were investigated using linear and logistic regression analyses, respectively. The multivariable analyses with uptake as the criterion variable were executed in three steps, by consecutively adding the following three sets of determinants: intention, proximal determinants, and distal determinants. Determinants were only included if the bivariable association with the criterion variable was Parents/guardians of 4,216 girls were invited to participate in this study. The response rate was highest among mothers born in the Netherlands (37%), followed by the Other group (34%), SNA group (31%) and with the lowest response rate among mothers born in one of the Middle East or North African countries (20%)  returned a questionnaire of whom 1,317 (96%) returned the questionnaire before the first scheduled HPV vaccination day. Ethnicity was missing for 8 participating parents/guardians. This resulted in a net-response of 1,309 participants , median age was 45 years (inter quartile range (IQR): 42–48), and 28% received higher education. On average, the NL-group was oldest (median 46 (IQR: 43–49)) and reported the highest level of education (34%), 74% of the NL-group reported not to be religious, while in the MENA-group 80% reported to be Islamic, and, finally in the NL-group only 7% received help when completing the questionnaire while this was 43% in the MENA-group (see Table p < 0.001), with the lowest intention found in the MENA-group . In the NL-group we found the largest fraction of daughters receiving two HPV vaccine doses (84%), while the SNA-group had the largest fraction of daughters that received only one dose (13%) and the MENA-group had the largest fraction of non-vaccinated daughters (41%). Overall past childhood vaccination uptake was 95%  but differed across ethnic groups  of HPV vaccination was 0.56 in the NL-group ; 0.28 in the SNA-group ; 0.23 in the MENA-group ; and 0.29 in the Other-group .The explained fraction (pseudo-Rp = 0.020) and Other-group (p = 0.042) when compared to the NL-group; the association of subjective norms with uptake was significantly lower in the SNA-group (p = 0.025) and MENA-group (p = 0.002) when compared to the NL-group; habit strength was significantly lower in the MENA-group when compared to the NL-group (p < 0.001).We tested whether the magnitude of the association of determinants of HPV vaccination uptake differed significantly in the non-Dutch groups compared to the NL-group Table . The assp = 0.002), 30% in the MENA-group (MENA- vs NL-group p < 0.001), and 11% in the Other-group (Other- vs NL-group p < 0.001) ultimately did not go for the HPV vaccination (inclined abstainers). Among the participants with a negative intention we observed that that 11% in the NL-group, 7% in the SNA-Group (SNA- vs NL-group p = 0.606), 6% in the MENA-group (MENA- vs NL-group p = 0.621) and 19% in the Other-group (Other- vs NL-group p = 0.298) did go for the HPV vaccination (disinclined actors).Among the participants with a positive intention to vaccinate against HPV we observed that 4% in the NL-group, 11% in the SNA-Group  att att1: Ta2) was: 0.77 in NL-group , 0.66 in SNA-group , 0.75 in MENA-group , and 0.66 in Other-group .The explained fraction of HPV intention (Rp = 0.001) and Other-group (p = 0.001); when having taken the decision regarding the HPV vaccination for an older daughter, parents/guardians from the NL-group would more often take the same decision for the daughter now invited for vaccination, compared to parents from the SNA-group (p = 0.024) and MENA-group (p = 0.020). Religion plays a significantly stronger role on HPV vaccination intention in the NL-group when compared to the Other-group (p = 0.029).We tested whether the magnitude of the association of determinants of HPV vaccination intention was significantly different in one of the non-Dutch groups when compared to the NL-group Table . The assWhen comparing analyses restricted to the complete cases with analyses done on the imputed dataset we found similar results which did not affect our main conclusion  when compared to the other ethnic groups (pseudo-R2 of around 0.20). This discrepancy can be attributed to the higher proportion of participants with a positive intention who ultimately did not go for the vaccination (inclined abstainers [In the NL-group intention explained a much larger fraction of the variability in HPV vaccination uptake (pseudo-Rstainers ) in the stainers . One migWe did not find large differences between the strength of association of the determinants on intention across ethnic groups, except for a couple of subtleties in which we found subjective norms and having an older daughter for which they already decided about the HPV vaccination to play a significantly stronger role in the NL-group. We found similar proportions of explained variance and similar determinants as in a previous study on native Dutch : i.e. atThis is the first study to present longitudinal results regarding the association of intention to vaccinate against HPV and actual (rather than self-reported) HPV vaccination in an ethnically diverse sample. Since illiteracy and language barriers are known issues in our study population , we antiWe recommend future communication strategies to target social-psychological determinants of HPV vaccination intention and uptake. Furthermore, we may conclude that intervention programs should target the same determinants for all ethnic groups, although some determinants will have stronger effects in some groups than in others. Within the scope of these determinants, the process of HPV vaccination decision making has shown to be mainly attitudinally driven, however, we should not neglect processes of internalization of social normative/media influences. As HPV vaccination intention and uptake in the different ethnic groups is based on similar determinants, future interventions could employ similar behavior change methods . However, the selection and design of concrete applications or strategies to reach parents in the different ethnic populations may need tailoring. For example, a prerequisite for a successful communication is that non-Dutch speaking parents/guardians are able to understand and process the HPV vaccination information offered to them. In the MENA-group we found that 18% completed the questionnaire in another language than Dutch and that 43% completed the questionnaire with help of someone, we therefore advise to translate this information into their native language, and to use interaction-based approaches  so parents/guardians are able to ask questions and exchange their concerns about the HPV vaccination. This also provides opportunities to correct possible misperceptions before they become salient within a particular ethnic community. Further research is needed to understand the discrepancy between intention and behavior among the non-Dutch groups. Our research stresses the need of measuring actual uptake instead of intention in an ethnical diverse population.We confirmed that social-psychological factors are most decisive when parents/guardians make a decision regarding vaccinating their daughter against HPV. Based on the measured determinants we conclude that intervention programs can focus on the same determinants for all ethnic groups. Furthermore, we found the MENA-group and SNA-group to contain a large fraction of parents/guardians that had a positive intention to vaccinate yet ultimately decided not to go for the HPV vaccination of their daughter.


PAX1, ZNF582, SOX1, and NKX6-1 and HPV genotyping. Combined with HPV-16/18 genotyping, both a dual methylation test for PAX1/ZNF582 and testing for ZNF582 methylation demonstrated 100% association of methylation with pathology results, indicating carcinoma in situ or squamous cell carcinoma. The sensitivity and specificity of the dual methylation test for PAX1/ZNF582 as a reflex test for identification of CIN3+ lesions were 78.85% and 73.55% , respectively. This strategy could reduce the number of patients referred for colposcopic examination by 31.3% compared with cytology, and thus provide a feasible follow-up solution in regions where colposcopy is not readily available. This strategy could also prevent unnecessary anxiety in women with hrHPV infection.In 2015, the American Society for Colposcopy and Cervical Pathology and the Society of Gynecologic Oncology issued interim guidance for the use of a human papillomavirus (HPV) test for primary screening, suggesting triage of women positive for high-risk human papillomavirus (hrHPV) by HPV-16/18 genotyping and cytology for women positive for non-16/18 hrHPV. The design of the present study was based on this interim guidance and analysis of the methylation status of specific candidate genes, which has been proposed as a tool to reduce unnecessary referral following primary HPV screening for cervical cancer. We performed a hospital-based case-control study including 312 hrHPV-positive women. hrHPV genotyping was performed by nested multiplex PCR assay with type-specific primers.Residual cervical cells from liquid-based cytology were used for extraction of genomic DNA for assessment of the methylation status of  Infection with a human papillomavirus with a high-risk genotype (hrHPV) is the only known etiologic risk factor for the development of cervical cancer and a necessary step in carcinogenesis –4. The dThe American Cancer Society (ACS) and the American Society for Colposcopy and Cervical Pathology (ASCCP) recommend that either co-testing every 5 years or cytology-only screening every 3 years should be implemented for women aged 30–65 years, whereas cytology alone should be performed for women between 21 and 30 years of age . SeveralIn China, cervical cancer remains the seventh-leading cause of death from cancer among females, and there were almost 62,000 new cases and 30,000 deaths due to cervical cancer in 2012 . In urbaThe use of HPV testing has increased dramatically in China in recent years because of the achievement of consistent and repeatable results. The lack of clinical validation of the majority of HPV tests used in hospitals has led to confusion and huge numbers of women undergoing colposcopic examination. The aims of cervical cancer screening are to discover high-grade lesions or cancer, rather than to detect the virus, whose presence does not necessarily indicate malignant disease and can thus lead to unnecessary anxiety for the women tested. Therefore, there is an urgent need for an objective, reproducible, and accurate method of screening for cervical cancer in China.PAX1), sex determining region Y-box 1 (SOX1), zinc finger protein 582 (ZNF582), and NK6 transcription factor-related locus1 (NKX6-1) have reported their potential as biomarkers for cervical cancer screening and for triage of cytological diagnoses and high-risk HPV infection groups [PAX1 (PAX1m), SOX1 (SOX1m), ZNF582 (ZNF582m), and NKX6-1 (NKX6-1m) have been extensively documented as highly sensitive biomarkers for detection of cervical intraepithelial neoplasia at grade 3 or higher (CIN3+) [ZNF582m has been found to have clinical potential for cancer detection. Clinically, ZNF582m demonstrated 70% sensitivity and 82% specificity for CIN3+ lesions in a Taiwanese case-control cohort [ZNF582m also has good sensitivity and specificity for triage of low-grade squamous intraepithelial lesion (LSIL), with values of 73% and 71%, respectively [ZNF582 has been reported to exhibit a high methylation rate in cervical adenocarcinoma, which has been difficult to diagnose [One such screening possibility arises from the field of epigenetics . Numeroun groups –25. In f (CIN3+) . In partl cohort . ZNF582mectively . Moreovediagnose .PAX1, ZNF582, SOX1, and NKX6-1 and liquid Pap smears, were compared and analyzed. We propose that this approach would be useful in outpatient departments of hospitals in highly populated countries, where screening is not readily accessible through community-based settings.Here we report a case-control study of diagnosis of CIN3+ in hrHPV-positive cases negative for HPV 16/18 by detection of methylated DNA, HPV genotyping, liquid-based cytology, and combinations thereof. The results were analyzed following the recommended primary HPV screening protocol algorithm . The anaZNF582m, and PAX1m/ZNF582m was 66.5%, 77.4% and 86.5% for CIN3+ lesions in the HPV positive patients (n=312), respectively. The specificity of HPV16/18, ZNF582m, and PAX1m/ZNF582m was 77.1%, 77.7%, and 65.0%, respectively. The referral rate of HPV16/18, ZNF582m, and PAX1m/ZNF582m was 44.6%, 50.0%, and 60.6%, respectively. There were 173 HPV-16/18-negative hrHPV patients, of whom 81 (46.82%) had normal pathologic results, 13 (7.51%) were CIN1, 27 (15.60%) CIN2, 35 (20.23%) CIN3, 4 (2.31%) CIS, and 13 (7.51%) SCC. The demographic characteristics, reflex testing results (including cytology), most common HPV genotypes in Chinese women, and methylation results for PAX1, ZNF582, SOX1, and NKX6-1 are presented in Table PAX1m, ZNF582m, SOX1m, and NKX6-1m, respectively.In total, 461 patients were enrolled in this study, of which 12 were excluded, 137 were hrHPV-negative, and 312 were hrHPV positive , 71.2%–86.7%). The sensitivity and specificity of ZNF582m in the identification of CIN3+ lesions were 76.92% and 80.99%, respectively.The sensitivity, specificity, and AUC results for the different molecular reflex tests for detection of CIN3+ lesions, individually and in combination, are presented in Table PAX1m/ZNF582m, PAX1m/SOX1m, and ZNF582m/SOX1m resulted in AUC of 76.2%, 77.6%, and 75.9%. The highest odds ratios for combinations of two methylated genes were those for PAX1m/ZNF582m and PAX1m/SOX1m, at 10.37 and 12.19, respectively.Dual DNA methylation testing of positivity for either gene in pairs of PAX1m/ZNF582m, PAX1m/SOX1m, and SOX1m/ZNF582m were the most discriminatory. ZNF582m combined with either PAX1m or SOX1m achieved 100% positivity for histology results indicating cancer; however, the positive rate decreased to between 62.86% and 74.29% for the CIN3 category  (Table PAX1m/SOX1m or PAX1m/ZNF582m) as reflex tests, the positive rates for CIN3+ histology were 6.94% higher than for PAX1m alone, and use of the SOX1m/ZNF582m dual test increased positive rates for CIN3+ by 11.56% compared with ZNF582m alone, as shown in Figure Colposcopy is a diagnostic procedure performed by physicians to examine the cervix, vagina, and vulva closely for signs of disease. Due to the high cost and insufficient availability of colposcopy, it is desirable to reduce the number of colposcopy referrals, while not missing women with severe dysplasia or carcinoma . Our results indicated that the referral rates for colposcopy after reflex tests using the individual methylated genes HPV infection is a major cause of cervical cancer. HPV genotype testing, based on the identification of HPV DNA in the cervix, has the potential to improve HPV testing and demonstrates higher sensitivity for high-grade CIN than cytology testing, based on the identification of changes in cellular morphology. DNA testing for high-risk HPV types is frequently performed in parallel with cytology to detect high-grade dysplasia and cervical cancer, particularly in women over 30 years of age. HPV DNA testing is a relatively simple process and therefore advantageous for large-scale screening in China or developing countries, which are lacking cytology infrastructure and expert cytologists. In addition, HPV DNA testing of self-collected vaginal samples can be less uncomfortable for conservative women, and can increase participation rates in primary or routine screening for cervical cancer –32.PAX1, ZNF582, and HPV16/18 that can distinguish between the CIN3+ and CIN2- groups are urgently required. Cytology is considered appropriate for application as a reflex test for hrHPV-positive women [It is believed that only persistent HPV infections are associated with pre-cancerous lesions. Despite its high sensitivity, hrHPV testing cannot distinguish whether or not an HPV-positive result is associated with a clinically relevant lesion, and a positive hrHPV result may lead to over-interpretation of minor cellular abnormalities, thereby reducing the specificity of this technique . Therefove women . For detve women . When usve women . Recent ve women –42.PAX1m/ZNF582m) testing indicate a need for biopsy during colposcopy for diagnosis of CIN3+ lesions. We suggest a 1-year follow-up of patients with positivity for the 12 high-risk HPV genotypes but negative results for HPV-16/18 and methylated gene testing. Because HPV-16/18 and methylation co-testing demonstrated a 100% rate of positivity for cervical cancer identification in this study, we suggest that because of the lower risk for CIN3+ lesions, patients with negative results by hrHPV and methylated gene testing should be followed up after 5 years or more [There is increasing evidence that testing for methylated genes could replace cytology as a reflex test for women positive for the 12 hrHPV types other than HPV-16/18, and interim clinical guidance approves the use of such tests as an appropriate triage tool for hrHPV , 43. We  or more .PAX1m and ZNF582m for detection of high-grade CIN lesions and cervical cancer. We explored the value of triage testing of hrHPV-positive women by combined testing for positivity for HPV-16/18 or the dual methylated markers, PAX1m or ZNF582m. Combined triage resulted in substantially higher sensitivity and specificity for CIN3+, compared with cytology alone, genotyping for the 12 other hrHPV types individually, or cytology with HPV-16/18 testing. The strengths of the present study lie in its use of a delinked case-control study design and the incorporation of all histological results, including those of the normal cytology group.Several previous studies have supported the great promise of analysis of In 2012, the ACS proposed new screening guidelines that included using age-adjusted screening methodology as well as cytological and HPV co-testing. HPV-16/18 typing has been recommended for hrHPV-positive women ; howeverThe huge number of hrHPV patients referred for colposcopic examination and follow-up could be an extra burden in China, should HPV testing be used for hospital-based opportunistic screening. Our data demonstrate that following the current interim clinical guidance would not significantly reduce the referral rate for colposcopy. However, the substantial variability in sensitivity observed among areas with different cytology infrastructures and insufficient numbers of experienced cytologists are other factors reducing the referral rate for colposcopy in China. Comparison of cytology and dual methylated markers as reflex tests indicated that the dual methylated markers resulted in a referral rate for colposcopic examination that was 24.4% of the rate for cytology.The present study has some potential limitations. Not all hrHPV-positive patients were referred for colposcopy examination because of the limited budget of the clinical study and based on the independent judgment of gynecologists in clinic. In addition, 144 hrHPV-ASC-US patient samples were collected during colposcopy because the majority of patients with obvious cervical cancer underwent biopsy immediately following abnormal Pap smear test results and clinical observation in the outpatient department. Other limitations include the small sample size and a lack of extensive and long-term follow-up information. In addition, the use of dual methylated markers as the only triage biomarker for hrHPV-positive women was not analyzed in this study, and this study is not representative of a typical screening population.PAX1m and ZNF582m, combined with HPV-16/18 testing, could help to reduce the number of hrHPV- positive women being referred for colposcopy. Furthermore, implementation of our recommendations could simplify the logistics of follow-up care, especially in regions where colposcopy is not readily available. We propose that the suggested approach would be useful in outpatient departments of hospitals in highly populated countries where screening is not readily accessible through community-based settings.We recommend a new hrHPV flow-chart procedure for primary screening for cervical cancer. Our study suggests that the dual methylated markers, In accordance with the Declaration of Helsinki, all patients provided informed consent for their participation in the clinical study protocol approved by the Institutional Review Board of the Department of Clinical Pharmacology at Xiangya Hospital, Central South University, China. The clinical trial was registered in the Chinese Clinical Trial Registry (ChiCTR-DOD-14005446). Study inclusion criteria were females who were sexually active and not pregnant, had an intact uterus, and had no history of treatment for cervical intraepithelial neoplasia (CIN) or cervical cancer. Patients who had a history of cancer related to the reproductive tract, therapy for cervical lesions, HPV vaccinations, or a current pregnancy were excluded. The recruited subjects were in attendance following referral for colposcopy examination, and consisted of patients with both normal Pap and abnormal Pap smear results, inflammation symptoms, cervical erosion, bleeding symptoms, or suspected cervical cancer.The sub-group of patients who were hrHPV-positive was invited to the colposcopic examination room of the Department of Obstetrics & Gynecology to participate in a blinded study where testers were not provided with clinical data. After patients signed the informed consent form, each underwent a colposcopic examination and biopsy. The cytology results were classified according to the 2001 Bethesda System (TBS 2001). Colposcopy-directed biopsies were performed for histological analysis, according to standard procedures in China. The final diagnosis was based on the results of tissue-proven pathology. Standard guidelines for the management and treatment of cervical neoplasia were followed in all patients . All patIn total, 461 patients were enrolled in the study from November 2011 to March 2013. Twelve patients were excluded based on the exclusion criteria, including five who were excluded because of poor-quality DNA specimens. This study focusses on data from the sub-group of 312 women who were hrHPV-positive following interim clinical guidance [All liquid-based cytology samples were clinician-collected using the Cytoprep Brush  and preserved in CytoFast Solution  in the clinic one week prior to the colposcopy visit. Residual cervical cells from cytological tests were used for HPV genotyping and methylation detection tests for the four genes. All specimens collected were given an identification number and delinked from patient clinical information until final data analysis. All molecular tests were performed at the Institute of Clinical Pharmacology, Hunan Key Laboratory of Pharmacogenetics, China, following Good Laboratory Practice (GLP) guidelines. The cells were centrifuged and stored in phosphate-buffered saline at -20°C from the day of collection. Genomic DNA (gDNA) was extracted from the collected cells using the QIAamp DNA Mini Kit . A BioSpec-nano spectrophotometer  was used to quantify the amount of extracted DNA.PAX1, ZNF582, SOX1, and NKX6-1 were determined using the iStat Biomedical Q-PCR kit on the LightCycler LC480 real-time PCR system , according to the manufacturer’s protocol. The criteria for positive and negative results for the methylated genes were based on the delta crossing point (Cp) values, as determined in previous studies [Briefly, 500 ng of gDNA was subject to bisulfite conversion using EZ DNA Methylation-Gold kits . The methylation levels of  studies , 23, 25.The hrHPV typing testing  was a clinical test performed using a nested multiplex PCR assay that combined degenerate E6/E7 consensus primers and type-specific primers as previously described , 49. ThePAX1m = ΔCp ≤ 9, SOX1m = ΔCp ≤ 8, ZNF582m = ΔCp ≤ 11, and NKX6-1m = ΔCp ≤ 11. SPSS software  was used for all statistical analyses. Both Chi-squared and Fisher’s exact tests were used to analyze the status of the methylated genes or of HPV genotyping in different combinations. Fisher’s exact test is considered more accurate than the chi-squared test when the sample size is smaller than five. A cross-validated receiver operating characteristic (ROC) curve was generated and the area under the ROC curve (AUC) for CIN3+ lesions was calculated for each detection method. The sensitivity, specificity, and odds ratio (OR) with 95% confidence interval (CI) were calculated for lesions classified as CIN3 or worse. All tests were two-sided and differences were considered statistically significant at P < 0.05.The positive cutoff values for methylated genes were determined as