Every year nearly half a million new cases of cervix cancer are diagnosed worldwide, making this malignancy the fourth commonest cancer in women. In 2018, more than 270,000 women died of cervix cancer globally with 85% of them being from developing countries. The majority of these cancers are caused by the infection with carcinogenic strains of human papillomavirus (HPV), which is also causally implicated in the development of other malignancies, including cancer of the anus, penis cancer and head and neck cancer. HPV is by far the most common sexually transmitted infection worldwide, however, most infected people do not develop cancer and do not even have a persistent infection. The development of highly effective HPV vaccines against most common high-risk HPV strains is a great medical achievement of the 21st century that could prevent up to 90% of cervix cancers. In this article, we review the current understanding of the balanced virus-host interaction that can lead to either virus elimination or the establishment of persistent infection and ultimately malignant transformation. We also highlight the influence of certain factors inherent to the host, including the immune status, genetic variants and the coexistence of other microbe infections and microbiome composition in the dynamic of HPV infection induced carcinogenesis.  Human papilloma virus (HPV) is a ubiquitous double stranded DNA virus that is associated with the development of various types of benign and malignant neoplasms in humans. More than 200 HPV types have been described so far and a number of them are transmitted through intimate skin-to-skin contact during sexual contact . These vAlthough there is a great variation in the prevalence of HPV infection worldwide, it is estimated that nearly all sexually active people around the world may get infected with some HPV type at some point of their lives, and thus HPV infection is considered the most common sexually transmitted infection in the world .HPV infections have been causally associated with the development of various cancers, including, cancers of the cervix, vagina, and vulva in women; cancers of the penis in men; and cancers of the anus and of the head and neck in both women and men ,6,7. In In nearly 50% of cases HPV infections are cleared by an appropriate immune response within six months of exposure and in the majority of cases (~90%), the virus become undetectable within 1–2 years . WhereasSeveral immunological factors, from both the innate and adaptive immune system play important roles in HPV recognition and elimination and in most cases these lines of defense are highly effective. However, HPVs have developed several mechanisms aimed to escape immune response, ranging from unspecific viral strategies, capable of downregulating various components of the innate immune system, to more sophisticated evasion strategies that turn ineffective the adaptive immune response against the virus, which ultimately results in persistent viral infections. In addition, once malignant transformation has started, the virus can hijack the immune system creating a chronic proinflammatory response favorable for cancer development and tumor progression. In parallel to the immune escape mechanisms of the virus, during the host/virus interaction, other important players are also involved. For example, host genetic variants in genes involved in human immune response may influence individual susceptibility to the long persistence of the virus and ultimately could influence HPV-induced cancers or tumor progression. Similarly, the presence of other microorganisms in the host, such as infection with other viruses, or alterations in the microbiota composition (dysbiosis) in the tissues infected with HPV, directly or indirectly may determine the virus infection outcomes. Consequently, HPV-induced carcinogenesis appears to be the result of a breakdown of the balance between the host and the virus, where the host’s defense mechanisms  are overcome by the pathogenic factors of virus leading to a persistent infection that ultimately can cause cancer. In this article, we discuss current understanding on the virus/host interaction during HPV-induced carcinogenesis and highlight recent insights on the immune response against the virus, as well as on the possible implications of microbiome during such virus/human interplay that may be translated into novel preventive and therapeutic approaches against HPV-induced cancers. HPV is a nonenveloped double-stranded DNA virus with a circular genome (approximately 8000 base pairs) that can be physically divided into three regions: an early region of six early viral genes  encoding non-structural proteins; a late region with two late genes (L1 and L2) encoding structural viral capsid proteins required for virion formation and virus spread ,15; and Some HPV early gene products such as E1 and E2 are critical regulators of viral DNA transcription and replication and E2 especially contributes to maintaining the intracellular transport and nuclear accumulation of the DNA during infection . AnotherHPV has evolved to synchronize its infectious cycle to the differentiation program of the target cells (keratinocytes) in the host. Once the virus gain entry into the epithelium via skin microlesions, it infects immature basal keratinocytes and very likely the stem cells ,22 and wTypically, during its viral cycle, HPV genome does not integrate into the host DNA, however, for yet undefined mechanisms, in some circumstances the HPV genome integrates randomly into the host DNA, in a process that includes the disruption of E2 expression, and since E2 is a negative transcriptional regulator of E6 and E7 viral oncogenes, when E2 is lost, E6 and E7 become actively expressed, promoting malignant transformation, which is further promoted by the ability of E6 and E7 oncoproteins to disrupt the tumor-suppressor genes p53 and pRB, respectively . As a reCervical cancer, the most frequent malignancy associated with HPV, typically develops from precancerous lesions over 10 to 20 years. About 90% of cervical cancer cases are squamous cell carcinomas, 10% are adenocarcinomas, and a small number are other types. The classification of cervical carcinoma precursor lesions has been modified several times over the years. For example, the world health organization classification system categorized cervical lesions into mild, moderate, or severe dysplasia or carcinoma in situ (CIS). Thereafter, the term cervical intraepithelial neoplasia (CIN) was developed to emphasize on the spectrum of abnormalities in these lesions in an attempt to standardize treatment. It classifies mild dysplasia as CIN1, moderate dysplasia as CIN2, and severe dysplasia and CIS as CIN3. More recently, CIN2 and CIN3 have been combined into CIN2/3. On the other hand, the Bethesda system is based on the Papanicolaou (Pap) test results and classifies lesions into Low-grade Squamous intraepithelial lesion (LSIL) and High-grade intraepithelial lesion (HSIL). Although they are results of different tests, an LSIL Pap may correspond to CIN1, and HSIL may correspond to CIN2 and CIN3. Despite the high frequency of HPV infections, malignant transformation of infected tissues is not a common phenomenon suggesting that the immune system plays a central role in the control of these infections. This is substantiated by the increased frequency of HPV-associated cervical dysplasia observed in immunosuppressed patients and further supported by the effective immune response induced by anti-HPV vaccines. On the other hand, HPVs have evolved to infect the host and to some extend propagate almost inadvertently avoiding confrontation with the human immune system. As mentioned above, the synchronization of the viral cycle to keratinocyte differentiation process allows the virus to propagate without inducing a major inflammatory response and since HPV replication occurs in the epithelium, which is a poorly vascularized area, circulating immune cells cannot approach the virus easily. The lack of inflammatory cytokine release from infected keratinocytes limits the activation of resident skin Langerhans’s cells and dendritic cells, which are required for the effective induction of an adaptive immune response. In addition, accumulating evidence indicate that constitutively expressed cellular factors (restriction factors), including various intrinsic nuclear factors such as ND10 factor (sp100), IFIT1 (p56), IFI16, APOBEC, and others (reviewed in ) can dirInnate immunity is the first line of defense against pathogens, including HPVs, and is composed of nonspecific mechanisms including physical barriers, such as skin, soluble factors (cytokines and other inflammatory factors) and cellular components such as granulocytes  and non-granulocyte cells such as natural killer cells (NK cells), monocytes, macrophages and dendritic cells, which constitute an efficient bridge that connects the innate immune response with the adaptive immune system. HPVs have developed strategies that efficiently evade or inhibit the innate immune response. For example, E7 oncogene, and to a lesser extend E6, reduce NFκB activation , which sMost cells of the innate immune system express pathogen recognition receptors (PRRs) such as cytosolic retinoic acid-inducible gene I (RIG-I)–like helicases for the recognition of viral RNA and several members of the toll-like receptors (TLR) family that also play a crucial role in innate immunity. Within the TLR family, TLR9 is implicated in the recognition of double-stranded DNA CpG motifs present in the genome of viruses like HPVs. TLR9 activates cellular signals that lead to the production of type I IFN and proinflammatory cytokines during viral infections . HoweverProfessional antigen presenting cells (APCs), such as dendritic cells (DC), macrophages, and subsets of B cells are immune cells that constitutively express Major Histocompatibility Complex class II (MHC class II) molecules on their surface and specialize in presenting antigens to T-cells, thus playing a critical role in the activation of the T cell-mediated immunity. Conversely, non-professional APCs constitute a variety of immune and non-immune cells including fibroblasts, keratinocytes thymic epithelial cells, thyroid epithelial cells, glial cells, pancreatic beta cells, and vascular endothelial cells, which do not constitutively express MHC class II proteins, can express these molecules upon cytokine stimulation, especially IFN-γ, and may thus interact with and present antigens to naïve T cells . DCs plaKeratinocytes, the predominant cell type in the epidermis, play a critical role during HPV infection not only because they are the cells were HPVs proliferate but also because they have innate immune-like functions. For example, keratinocytes express various TLRs, including TLR9 and upon stimulation they secrete various cytokines like TNF-α, IL-8, CCL2, CCL20, CXCL9 and type 1 IFNs . As mentMacrophages contribute to the clearance of HPV infections by eliminating HPV infected cells  and by sNKG2D-Ls are molecules expressed on cells exposed to stress stimuli such as virus infection, radiation or malignant transformation and these ligands can be recognized and killed by immune cells expressing the immunoreceptor NKG2D, including NK cells, γδT cells, and other T cells subsets. Interestingly, NKG2D-Ls  have been detected in cervical cancer cell lines ,48 and tE7 oncoprotein directly interacts and blocks the major histocompatibility complex class I (MHC-I) heavy chain promoter leading to reduced MHC-I expression on infected cells . SimilarIndoleamine 2,3-dioxygenase 1 (IDO1) is a naturally immunosuppressive molecule that regulates immune function by controlling tryptophan levels. In normal conditions, IDO1 controls excessive immune activation preventing autoimmunity, however, tumor cells upregulate IDO1 activity by depleting tryptophan in the tumor microenvironment . Dermal Recently, using patient tissue specimens and cultured keratinocytes, Cicchini and colleagues found that oncoprotein E7 induces hypermethylation of CXCL14 promoter leading to the downregulation of the chemokine CXCL14. In murine models of head/neck and cervical cancers, restoring Cxcl14 expression in HPV-positive cancer cells dramatically increased NK and T cells in the tumor-draining lymph nodes, leading to potent anti-tumor responses in vivo . CXCL14 β1 promoter activity [Oncoproteins E6 and E7 of high risk HPVs also impair immune response by promoting the production of transforming growth factor β (TGFβ) by increasing TGF-activity . As a reactivity .Cell mediated immunity mediated by both, T-helper cells  and cytotoxic T-cells , plays a pivotal role in the elimination of HPV infections and in the regression of HPV-induced premalignant lesions . For exaFunctionally, Th cells are phenotypically divided into Th1 and Th2 subsets. Th1 cells primarily produce proinflammatory cytokines  and generate responses against intracellular pathogens such as bacteria and viruses. Conversely, Th2 cells produce IL-4, IL-5, IL-6, IL-10, and IL-13 and generate immune responses against extracellular pathogens. Interestingly, HR-HPVs have the ability to shift the immune response from Th1 to Th2 during early HPV-induced lesions, which harness antiviral responses, thus promoting disease progression .The immunoregulatory molecules programmed death-1  and its ligand PD-L1 (CD274) have been linked to HPV carcinogenesis. Ligation of PD-1  by its ligand PD-L1  induces an inhibitory signal in activated T cells leading to T-cell apoptosis, anergy and functional exhaustion. Increased PD-1 and PD-L1 expression has been found on HR-HPV infected cervical tissues and its expression was correlated with increasing CIN grade ,69. In aMoreover, in preclinical studies, microparticles shed from HPV16 E7-expressing keratinocytes suppressed the cytotoxicity of CD8+ T cells, which was attributed to the downregulation of CD40 and IL-12 in Langerhans cells , indicatNatural HPV infection generates a specific humoral immune response against various viral antigens, as demonstrated by a high prevalence of naturally acquired antibodies to specific HPV types in individuals without cancer, in both sexes and all age groups . These aHumoral immunity against HPVs may predict the risk of malignant transformation in HPV infected individuals, although this may depend on the type of cancer. For example, high levels of anti-HPV16 E6 antibodies were detectable in most patients with oropharyngeal carcinomas (OPCs) more than 10 years before diagnosis  and in oRegulatory T cells (Treg) are a subset of T cells expressing the transcription factor Foxp3 that are required for the induction of immune tolerance and auto-immunity prevention. Given the immunosuppressive functions of Treg cells, one may expect that HPVs promote the expansion of these cells, as a viral strategy to evade immune surveillance. This assumption is supported by the observations that HPV-induced lesions have elevated numbers of infiltrating FOXP3+ Tregs cells and Foxp3 expression is associated with increased odds of progression to high-grade lesions ,79. FurtIn tissue specimens from laryngeal squamous cell carcinoma (LSCC), accumulation of activated Treg cells and M2 macrophages (a subset of macrophages with immunosuppressive activities that are associated with increased tumor progression) was associated with adverse prognosis and a positive-feedback loop between activated Treg cells and M2 macrophages is crucial to the establishment of the immunosuppressive tumor microenvironment that is typically observed in these malignancies .Interestingly, a recent study showed that in OPSCC specimens, tumor infiltration by Treg cells expressing Tbet+ (Foxp3+ Tbet+ Tregs) was strongly correlated with a concomitant tumor-specific and type 1-oriented intratumoral T cell infiltrate where an increased infiltration Tbet+ Tregs was associated with improved clinical outcomes .In summary, important mechanisms implicated in the evasion of immune responses by HPVs include perturbing antiviral activities of innate immune system, hampering antigen processing and presentation, inducing a shifted Th1 to Th2 immune response, promoting immune anergy via the recruitment of regulatory T cells and likely by hijacking regulatory mechanisms of T cells apoptosis .Single nucleotide polymorphisms (SNPs) constitute the most common form of genetic variation in the human genome and depending on their location within a given gene, SNPs can alter gene expression or protein function. Numerous SNPs are associated with disease predisposition and many have been reported to associate with individual’s susceptibility to cancer, including HPV-induced cancers. DRB1*1301 variant appear to have lower risk of developing cervical cancer [DRB1*1302 allele are less likely to progress from CIN1 to CIN 2/3 [Human leukocyte antigen (HLA) genes, which are highly polymorphic are among the most studied factors since certain HLA variants are consistently associated with numerous inflammatory disorders and cancer. For example, women harboring the l cancer ,92. Simi CIN 2/3 .SMUG1 gene were associated increased susceptibility of CIN III and CSCCs [Genetic variants in non-HLA genes have been also reported in association with HPV-related cancers. For example, rs3087404 and rs2029167 SNPs in nd CSCCs .TGF-β1 has been reported to confer individual susceptibility to HPV+ OPC [The immunosuppressive cytokine TGFβ1 impairs NK cells cytotoxicity by promoting the downregulation of NK cells activating receptors including NKp30 and NKG2D  and highHPV+ OPC . ImportaHPV+ OPC . AlthougKLRK1 gene (which encodes NKG2D receptor), with increased risk of HPV-induced cancers, including cervical cancer, anal cancer and vaginal cancer [SNPs in various genes encoding immunoreceptors have been also associated with increased risk to HPV-induced cancers. Our group reported an association between the variant LNK, determined by the SNP rs1049174 in the 3′UTR region of l cancer . Mechanil cancer ,100,101.l cancer . Notablyl cancer . The facl cancer  suggest KIR genes 3DS1 and 2DS1 are more likely to develop a more severe form of RRP (caused by HPV-6/11) than those harboring these receptors [KIR genes may be implicated in the development of HPV-induced cancers or in the risk to develop persistent infections and considering the fact that various KIR inhibitory antibodies are been tested in the clinical setting [Killer-cell immunoglobulin-like receptors (KIRs), constitute a large family of transmembrane proteins expressed on the surface of NK cells and a minor subset of T cells that interact with specific allelic variants of MHC class I molecules expressed on all nucleated cell types, thus regulating the cytotoxic functions of NK and T cells. Most KIRs are inhibitory, meaning that after recognizing their corresponding MHC molecules, they suppress the killing activity of effector cells (NK or T cells), however, a limited number of KIRs are indeed activating, implying that their recognition of MHC molecules activates the cytotoxic activity of effector cells . Interes setting , furtherCYP1A1 to be significantly associated with cervical cancer [It is plausible that certain generic variants are relevant for some populations while have little or no impact in other populations. A recent meta-analysis that included 98 publications reporting genetic association studies of SNPs with risk to cervical cancer in Indian women revealed that only rs1048943 of l cancer .In summary several studies have shown that genetic variants in certain genes, especially functional SNPs capable of affecting gene expression or altering the function of the encoded protein, may contribute to HPV-induced carcinogenesis, however, as occurs with many gene association studies, some findings are still inconclusive with some studies report contradictory associations across different populations and even in subjects of the same ethnicity . Given tImmunodeficiency associated with HIV infection is associated with impaired immune response of HPV infections. As a result, HIV+ women have increased risks of persistent HPV infection and higher risk to develop pre-malignant lesions and HPV-induced cancers and unfortunately, highly active antiretroviral therapy (HAART) appears to have limited ability to clear HPV infection and induce regression of premalignant lesions ,107.p = 001) greater CIN-3+ risk than HPV-16 negative women. Similarly, HIV-infected women with LSIL had 9-fold (p < 0001) greater risk, indicating that HIV-infected women with a normal Pap result who test HPV16 positive have high pre-cancer risk [In a large study that included a total of 2791 HIV-infected and 975 HIV-uninfected women, followed up with Pap tests and colposcopy, women with HIV that were positive for HPV16 had 13-fold  [Lactobacillus crispatus inversely associated with symptomatic viral infections, including HIV, HPV, or herpesvirus infections [Lactobacillus colonization promotes the growing of anaerobic bacterial species, thus altering the microbial diversity [Preclinical studies have documented that changes in microbiota composition can alter local innate immunity. For example, infection with d TNF-α) . These fd TNF-α) ,114.Thisfections ,116 and iversity .Mycobacterium, aeromonas, Sphingomonas, Bordetella and E.coli) fungi and parasites in these OCSCC samples [Changes in the oropharyngeal microbiome composition may be also associated with HPV-induced cancers like OCSCC/OPSCC, as a recent study found integration hotspots for HPV16 along with other identified integration sites for a number of viruses, including the JC polyomavirus, as well as other pathogenic bacteria ( samples .It is plausible that HPV can directly or indirectly induce changes in the infected tissues that affect the local microbiota composition and vice versa, dysbiosis of the local microbiota may in turn create conditions favorable for the colonization and the development of persistent infections with HPV or ultimately promote the development of precancerous lesions in a similar fashion as dysbiosis of gastric microbiota may promote the development of helicobacter pylori induced gastric malignancies . Since its introduction by George Papanicolaou in 1940’s, the conventional Pap test for the cytological evaluation of cervical cells has emerged as the most extensively utilized cancer screening technique worldwide and has contributed to saving millions of women’s lives. However, despite the tremendous success of the Pap test in cervical cancer screening, this cancer still represents a major public health issue, especially in the developing world, where more than 85% of deaths attributed to cervical cancer occur, hence developing new screening strategies is urgently needed. In this regard, new screening methods are under active investigation. For example, the analysis of biomarkers of virus persistence or cellular transformation is an area of active interest and has potential utility for the screening of other HPV-induced cancers ,121. In A recent Cochrane Systematic Review concluded that there is “high-certainty evidence” that HPV vaccines protect against cervical pre-cancer in adolescent girls and young women aged 15 to 26 and this effect is greater in those who are negative for HPV16/18 or other high risk HPVs at enrolment than those unselected for HPV DNA status . While tThe immune system plays a crucial role in HPV clearance; however, the virus has also developed various immune evasion strategies that can eventually promote persistent infection in the host, which may ultimately result in the development of cancer. A significant increase in circulating NK cells expressing the activator receptors NKG2D, NKp30, NKp46 and ILT2 has been observed following immunization with Gardasil HPV vaccines , which mUnderscoring the precise role that these factors play in HPV-carcinogenesis, as well as the identification of other relevant factors is crucial for developing new preventive strategies or therapeutic approaches.Alphapapillomavirus, betapapillomavirus, Gammapapillomavirus, Mupapillomavirus and Nupapillomavirus [Alphapapillomavirus, which is the most abundant and best characterized HPV genera, however, in recent years, multiple HPV types belonging to lesser characterized genera are being increasingly isolated from human specimens. Among them, betapapillomavirus types are linked to the development of skin flat lesions in epidermodysplasia verruciformis patients [betapapillomavirus with non-melanoma skin cancers [Based on their L1 ORF phylogenetic relationships, HPVs are classified into omavirus . This arpatients  and sero cancers ,133,134.


Since most cervical cancer cases are caused by persistent high-risk human papillomavirus (HR-HPV) infection, knowledge of HPV among women is essential for the prevent of cervical cancer. This study was aimed to assess knowledge among women in western China about HPV and its association with cervical cancer, and to assess their acceptance of HPV vaccination.A sample of healthy women undergoing routine physical examinations in the Health Management Center of West China Hospital, Sichuan University between January and December 2014 completed a questionnaire.n = 320) had heard of HPV; among this subgroup, only half (53.44%) knew that it causes cervical cancer, only 26 (8.13%) correctly answered all questions about HPV. Multivariate analysis showed that respondents who had heard of HPV were more likely than other respondents to have a family history of any cancer, to undergo regular Pap tests and to have completed at least secondary education. Half of all respondents (51.22%) reported that they would be willing to be vaccinated against HPV.A total of 1300 questionnaires were distributed, and 1109 were completed and analyzed. Only 28.85% of respondents  contains supplementary material, which is available to authorized users. Cervical cancer is the fourth most common cancer among women worldwide, with an estimated 528, 000 new cases and 266, 000 deaths in 2012 [Most cervical cancer cases are caused by persistent high-risk human papillomavirus (HR-HPV) infection –5. HPV iTwo HPV vaccinations have proven effective at protecting against infection with HPV strains 16 and 18 which might lead to cervical intraepithelial neoplasia (CIN) lesions and cervical cancer , and theHowever, levels of HPV awareness among the general population are poor throughout the world. Systematic review of studies from several countries suggests that only 15–31% of respondents have even heard of HPV, regardless of age, while only 0.6–11% know that HPV is a risk factor for cervical cancer . SurveysIt is unclear how much knowledge the general population in western China has about HPV, such as how the virus is transmitted, how infection can be detected, and whether it is linked to cervical cancer. Therefore, we wanted to examine in detail the HPV knowledge of women and their attitude to HPV vaccination. The results of this study may help guide the design and improvement of interventions and campaigns to sensitize women to HPV infection and vaccination, thereby helping reduce incidence of cervical cancer.All participants provide informed written consent to take part in our study. Healthy women undergoing routine physical examinations in the Health Management Center of West China Hospital of Sichuan University between January and December 2013 were invited to take a single survey. To be enrolled in the study, women had to be able to read the questionnaire on their own and understand it on their own or with the help of staff, they were given the paper-based questionnaire to complete by themselves and then return to staff. Women were not eligible if they were older than 75 or had a history of cervical cancer.The target study population was 1300 women, which assuming a 75% response rate should provide a sample adequate for estimating proportions for which the 95% confidence interval would have a maximum width of 10%. This corresponds to maximum error of ±5% for all estimates of proportions.A self-administered questionnaire was designed specifically for the present study and based on the literature –19. The Question 1 (Q1) asked respondents whether they had heard of HPV. Surveys on which the response to Q1 was “no” were excluded from analysis of responses to Q2–6. These five questions asked whether respondents knew various details about HPV infection. Q1–6 were as follows:options: yes, no).Q1. Have you heard of HPV? .Q2. Do you think HPV causes cervical cancer?.Q3. Is HPV infection asymptomatic? .Q4. Is HPV infection a sexually transmitted disease (STD)? .Q5. Can HPV infection cause an abnormal Pap test? .Q6. Did you know that human papillomavirus and human immunodeficiency virus (HIV) are different? .Q7. Are you willing to receive the HPV vaccine which can protect against HPV infection? . Descriptive statistics were used to evaluate knowledge about HPV infection and vaccine acceptance. Responses of “don’t know” were re-coded as “no” in the statistical analyses. Associations between respondent characteristics and level of knowledge or vaccine acceptance were explored in univariate analysis using the chi-squared test. Factors in the univariate analysis for which the associated A total of 1300 questionnaires were distributed and 1260 questionnaires were recovered, corresponding to a recovery rate of 96.92%. Of the returned questionnaires, 151 were excluded because they were incomplete. The remaining 1109 questionnaires were accepted as valid and used in the final analysis, corresponding to an effective response rate of 88.02% (1109/1260).The median age among all respondents was 40 range, 18–76), and the mean age was 40.42 ± 9.02  had ever heard of it. Among this subgroup of 320 women who had heard of the virus, 23.75% (76/320) knew that HPV infection is asymptomatic, 49.38% (158/320) knew that HPV infection is an STD, 47.81% (153/320) knew that HPV causes abnormal Pap test results, and 28.75% (92/320) knew that HPV is different from HIV. In this subgroup, 8.13% (26/320) correctly answered Q2–6  knew that HPV causes cervical cancer. Univariate analysis showed that these women were more likely than other women to be able to read, to undergo Pap testing regularly and to have a family history of cervical cancer  reported being willing to take the HPV vaccine. These results suggest the urgent need to disseminate knowledge about HPV and its association with cervical cancer, and they further suggest that such campaigns are likely to be effective at increasing willingness to be vaccinated.The level of HPV knowledge among our respondents is lower than that reported in a 2010 survey of wealthier women in Australia , suggestOur results identified several factors that positively correlated with HPV knowledge: family history of any cancer, regular Pap testing and an educational background of high school or above. HPV knowledge was not, however, associated with age, ethnicity, health status, family annual income, relationship status, frequency of sexual relations, history of STDs, or condom use. Our results suggest that HPV awareness depends more on education and exposure to HPV or cancer than on family income. One implication of our findings is that HPV awareness campaigns should be targeted at groups that normally receive little or no education about health in general or cancer in particular, including poorer, less educated groups without access to advanced healthcare facilities.Such education campaigns should aim not merely to raise awareness of HPV but also to emphasize its link with cervical cancer. Most women in our sample who had heard of HPV did not know that the virus causes cervical cancer. Women who had a higher educational background or a family history of cervical cancer or who regularly underwent Pap testing were more likely to know about this link. Education programs about HPV have been shown to be effective at improving awareness of cervical cancer, the association between HPV and cervical cancer and perception of HPV vaccines , 23. StuStrikingly, our study found that despite quite low rates of knowledge about HPV and its association with cervical cancer, half of respondents were willing to take the HPV vaccine once they knew that the HPV vaccine can protect against HPV infection. The reason for this high willingness may be the rising incidence of cancer in China . InteresThe results of this survey should be interpreted with caution given that the format, wording of questions and sequence of questions can bias responses , 33. In Despite these limitations, our results may be more reliable than some other studies because we did not analyze the responses of several questions for women who had not heard of HPV; as a result, our analysis of those questions should more accurately reflect knowledge among women previously exposed to HPV information. In addition, we added a statement that HPV had no relationship with breast cancer, a misconception that occurred in the pilot study because the Chinese name of HPV includes the word “papilla”.Our results point to an urgent need for education and intervention to raise awareness of HPV and its link to cervical cancer. At the same time, they provide encouragement that such education can increase willingness to take the HPV vaccine, since even in the absence of prior knowledge about HPV. Future work should examine the effects of such education on the psychology and health-seeking behaviors of women, and ultimately on the incidence of cervical cancer.Additional file 1:Questionnaire (Chinese). (DOCX 17 kb)Additional file 2:Questionnaire (English). (DOCX 18 kb)


Stratified mucin-producing intraepithelial lesion (SMILE) is a rare precursor lesion in the uterine cervix that is considered a variant of adenocarcinoma in situ (AIS). Although human papillomavirus (HPV) is thought to be related to the development of SMILE, there is little information available on the detection of HPV integrated into the lesion.A 30-year-old female underwent a routine uterine cervical cancer screening, and her Pap smear indicated the possible existence of atypical glandular cells. A cervical biopsy with endocervical curettage was performed. The histopathological analysis showed that she had SMILE and high-grade squamous intraepithelial lesion (HSIL) on her cervix. The lesion was found to be positive for HPV genotypes 52 and 68 by multiplex PCR. In situ hybridization with HPV RNA probes revealed that these HPV types were involved in the onset of HSIL and SMILE, respectively.Rare, high-risk HPV genotypes may contribute to the development of SMILE, and their detection can be useful for preventing the progression to carcinoma and ensuring adequate patient management. Uterine cervical cancer is the second most commonly diagnosed cancer and the third leading cause of cancer death among women in developed countries . AlthougStratified mucin-producing intraepithelial lesion (SMILE) is an uncommon premalignant lesion of the uterine cervix . It is tAlphapapillomavirus 9  or Alphapapillomavirus 7  [Studies over the last two decades have shown that persistent HPV infection is the main cause of cervical cancer development. Clinically validated HPV tests are recommended by the U.S. Preventive Services Task Force (USPSTF) and the Japan Society of Obstetrics and Gynecology for cervical pre-cancer screening, triage, and treatment follow-up in clinical practice , 14. Abo and 68) . However and 68)  with the and 68) . HPV RNA and 68) , 23. In A 30-year-old female  was referred to our hospital for routine uterine cervical cancer screening, and her Pap smear indicated the possible existence of atypical glandular cells. A colposcopic examination revealed dense white lesions in the 1 and 11 o’clock directions Fig. a as wellWe next examined lesions by single-molecule RNA fluorescent ISH using the RNAscope system   and specHistologically, SMILE is characterized by a multilayered atypical epithelium composed of cells with intracytoplasmic mucin in all cell layers. SMILEs are p16 positive and have a high MIB-1 proliferation index. However, SIL and AIS may coexist with SMILE, which is not surprising given their association with HPV infection. Moreover, a minority of invasive cervical carcinomas also exhibit both squamous and glandular features (referred to as adenosquamous carcinoma). Apart from the original description, there is little information on SMILEs, especially in relation to high-risk HPV infection . Most SMPrevious reports have suggested the preferential physical interaction of the virus with squamocolumnar junctions in the transformation zone of the basal epithelium that forms reserve cells with stem cell (SC) properties , 25. TheExamination of additional cases would be helpful in confirming the existence of CSCs in SMILEs. Therefore, we consider that most high-risk HPV types in cervical cancer are easily detectable, given their diffuse presence in the epithelium compared to that of high-risk HPV types, such as Alpha-7, that are predominant in the basal layer. Furthermore, the case implies that different HPV-infected cells individually define their disease phenotype as HSIL or SMILE. We speculate that SIL, AIS, and SMILE differ in terms of cellular origin with different HPV life cycles. Alpha-7 HPV types such as HPV18 and HPV68 may preferentially remain in the basal epithelium, unlike HPV16-related Alpha-9 HPV types, such as HPV52. In addition, complex cases of multiple HPV infection may exhibit distinct histopathology. Notably, our observations had certain limitations because of the study being a case report. Hence, further studies are needed to explore these possibilities. Nonetheless, our findings provide a basis for investigating multiple infection by Alpha-7 and -9 HPV types, the carcinogenicity of the rare HPV genotypes, and the outcome of SMILE in the cervix.


Cervical colposcopy biopsy was performed if any of the three results was positive. HC2, FISH, and pathology were compared. A total of 1200 women underwent screening, 150 patients underwent cervical biopsy: 32 in the normal group, 38 in the CIN1 group, 66 in the CIN2/3 group, and 14 in the invasive cervical cancer group. More patients had HR-HPV infection in the CIN2/3 group and ICC group compared with the CIN1 group. hTERC increased with increasing histological dysplasia. There was significant difference in hTERC positive rate between each of the three groups. More patients with hTERC gene amplification were observed in the positive HR-HPV group than in the HR-HPV negative group. In conclusion, hTERC is a potential marker for precancerous cervical cancer lesions. hTERC might be correlated with HR-HPV infection in cervical diseases.This study aimed to examine hTERC gene in different grades of cervical intraepithelial neoplasia (CIN) and cervical cancer, and the association between hTERC and high risk-human papillomavirus (HR-HPV) infection. Patients who underwent cervical cancer screening at the Second Affiliated Hospital of Kunming Medical University between October 2010 and December 2011 were enrolled. All patients underwent liquid-based cytology test and hybrid capture 2 (HC2) for HPV detection. hTERC was examined using fluorescence  Cervical cancer in China accounts for 12% of the worldwide cervical cancer incidence and for 11% of the cervical cancer deaths4. In countries that develop and implement cervical cancer screening, the incidence of cervical cancer has been significantly reduced by regular follow-up and monitoring screening quality5. But in many areas there are no standardized procedures or conditions to perform cervical cancer screening. In China, screening varies according to region, and barely reaches 19% of eligible rural women, resulting in increasing numbers of patients with cervical cancer4.Approximately 525,000 people worldwide are being diagnosed with cervical cancer each year6. The precancerous lesions maintain for a long time and can be reversed, so identification of cervical intraepithelial neoplasia (CIN) together with reasonable and effective early treatment are the keys to reducing the incidence and mortality of cervical cancer7. Human papillomavirus (HPV) infection is closely related to cervical cancer8. High risk-HPV (HR-HPV) can be detected in cervical cells of 99.8% of patients with cervical cancer9. Although the sensitivity of HPV-DNA detection is high, the specificity is not high, especially for young women with an active sexual life. Most HR-HPV infections are transient and disappear after 8–10 months, with only a small percentage of HR-HPV being persistent and eventually developing to cervical cancer10. Only long-term or repeated infection of HR-HPV may lead to unlimited proliferation of cells, greatly increasing the incidence of cervical cancer11. The low specificity of HPV detection limits its clinical application, and more reliable screening methods are needed.The occurrence and development of cervical cancer takes about 8–12 years from high-grade precancerous lesions to invasive cervical cancer12. The human telomerase RNA component (hTERC) acts as a template during telomere elongation but is also a vital part of the telomerase enzyme function13. Mutation, activation, and amplification of the hTERC gene may lead to increased telomerase activity14. Telomerase reactivation or upregulation is a critical feature in the vast majority of cancers15. Expression of hTERC gene is consistent with telomerase activity. hTERC is expressed in many normal tissues and benign tissues, but its relative up-regulation can reflect the degree of malignant progression and it is highly expressed in malignant tumors16. In cervical cancer and precancerous lesions, hTERC also has different degrees of amplification, and it has been considered to be a potential tumor marker for cervical cancer17.Telomeres are repetitive nucleotide sequences at the end of the chromosomes that maintain the stability of chromosomes and protect genes. These sequences are added by telomerase which contains a reverse transcriptase consisting of an RNA component, a reverse transcriptase and telomerase-related proteins18. HR-HPV infection can lead to hTERC gene amplification19. There is no activation of telomerase in the early stage of cancer induced by HR-HPV infection, but it tends to be consistent with the increasing histological dysplasia. Therefore, it is believed that there are sequential differences in the process of cervical carcinogenesis. So, HR-HPV infection is the initiation factor leading to reactivation of hTERC, and the combined action of the two result in cervical cells gaining unlimited proliferation ability20. There is a close correlation between HR-HPV infection and activation of the hTERC gene in the development of cervical cancer23. Increased HR-HPV and hTERC are related to more aggressive disease and may be important parts of future screening tests22.The HPV proto-oncoprotein encoded by its oncogene E6/E7 can up-regulate telomerase activity in cervical cancer and cervical intraepithelial neoplasia after HR-HPV infection, so that undifferentiated epithelial cells continue to proliferateThe aim of this study was to explore the role of the hTERC gene as a biomarker in cervical cancer. Therefore, hTERC gene expression and HR-HPV infection in exfoliated cervical cells of normal, CIN and ICC patients were detected, and the relationship between the two was analyzed.During the study period (October 2010 to December 2011), 1200 women participated in the cervical cancer screening program and 150 (12.5%) underwent cervical biopsy, including 32 (21.3%) in the normal group, 38 (25.3%) in the CIN1 group, 66 (44.0%) in the CIN2/3 group, and 14 (9.3%) in the ICC group. The HR-HPV infection rate was significantly different among the four groups (P < 0.001). There were more patients with HR-HPV infection in the CIN2/3 (90.9%) and ICC groups (92.9%) compared with the CIN1 group  and normal group . There was no difference in HR-HPV infection between the CIN2/3 and ICC groups (P = 0.864) . Patients with normal cervical histology had a rate of 12.5% of hTERC amplification. This rate increased to 34.2% for CIN1, 53.0% for CIN2/3, and 85.7% for ICC. There was significant difference in the hTERC positive rate between the CIN1 and CIN2/3 groups, between the CIN1 and ICC groups, as well as between the CIN2/3 and ICC groups (P < 0.05)  Table .Table 2hThis study investigated hTERC in different grades of CIN and cervical cancer, and the association between hTERC and HPV infection. The results showed that hTERC increased with increasing histological dysplasia and there was a significant difference in the hTERC positive rate between each of the three groups. 90.9% patients had HR-HPV infection in the CIN2/3 group, 92.9% in the ICC group and 65.8% in the CIN1 group. More patients with hTERC gene amplification were observed in the positive HR-HPV group than in the HR-HPV negative group. These results suggest that hTERC is a potential marker for precancerous cervical cancer lesions and that hTERC might be correlated with HR-HPV infection in cervical diseases. Nevertheless, some patients had lesions that were negative for both markers. This highlights that even if HR-HPV+ and hTERC amplification are involved in the pathogenesis of ICC, they are not the only factors involved and additional studies are necessary to identify what is driving the increasing histological dysplasia in those patients.24. More than 100 genetic types have been identified, including more than 40 types that can invade the female reproductive tract. These are divided into high-risk and low-risk types based on their carcinogenicity. Many laboratory studies and epidemiological investigations have shown that the main cause of cervical cancer is HR-HPV infection, and there is a chronological sequence for HR-HPV infection and cervical cancer25. The most sensitive method for diagnosing cervical lesions is HPV-DNA. Some studies have shown that 95% of patients with cervical cancer are infected with HPV26. PCR was used to detect HPV in 1918 cervical cancer specimens from nine countries, and the results demonstrated that the detection rate of HPV is as high as 96.6%27. The results of this study showed that HR-HPV infection rates in the normal, CIN1, CIN2/3, and ICC groups were 31.3%, 65.8%, 90.9%, and 92.9%, respectively. The differences among the groups were statistically significant (except between the CIN2/3 and ICC groups). The results showed that the HR-HPV infection rate increased with the increasing histological dysplasia of the cervical lesions, which is consistent with previous results21. Moreover, the results demonstrated that HR-HPV infection was not statistically significant between CIN2/3 and ICC groups.HPV somatotype is performed based on homology of nucleotide sequence28. Most studies have shown that the development of cervical cells from CIN1, CIN2, and CIN3 to cervical cancer is basically accompanied by the amplification of the long arm of chromosome 329. The gene most closely related to cervical cancer may be the hTERC gene. hTERC gene amplification can prevent cell apoptosis, leading to unlimited cell proliferation. Therefore, abnormal amplification of the hTERC gene may be an early event in tumor formation12. Heselmeyer-Haddad et al.30 used FISH trichrome probes in cells with cervical lesions; the 3q26 hTERC gene was amplified at different degrees in CIN2, CIN3, and cervical cancer. Moreover, normal cells, ASC-US, CIN1, and CIN2/3 were distinguished according to the degree of amplification of the hTERC gene. Tu et al.31 revealed that the amplification rate  of hTERC increases gradually in negative for intraepithelial lesions or malignancy (NILM), ASC-US, atypical squamous cell-cannot exclude high grade intraepithelial lesion (ASC-H), low grade squamous intraepithelial lesion (LSIL), and high grade squamous intraepithelial lesions (HSIL). Except for ASC-H and LSIL, pair-wise comparison was statistically significant. From these two studies, Tu et al.31 have concluded that the hTERC gene is a better method for differentiating between high- and low-grade lesions both cytologically and histologically. The results of this study showed that the positive rates of the hTERC gene in the normal, CIN1, CIN2/3, and ICC groups were 12.5%, 31.21%, 53.03%, and 85.71%, respectively. Compared to the normal group, there were significant differences in the CIN1, CIN2/3, and ICC groups (P < 0.05). There were significant differences between the CIN1 and CIN2/3 groups, between the CIN1 and ICC groups, and between the CIN2/3 and ICC groups. With the increasing histological dysplasia, the positive expression of hTERC gene showed a significantly increasing trend. The results were consistent with Tu et al.31 and later studies that also suggested that hTERC gene detection can detect cervical cancer in an early stage and may predict lesion progression23.The chromosomes of most solid tumor cells in humans often exhibit certain instability such as changes in the number of chromosomes and structural abnormalitieset al.32 used HR-HPV 16 to transfect normal human cervical epithelial cells. These transfected cells were passaged for more than 100 generations and had strong telomerase expression, suggesting that HPV infection is correlated with activation of telomerase. HR-HPV infection may lead to amplification of the hTERC gene33. HR-HPV infection is not accompanied by the activation of telomerase in the early stage of cervical cancer, but with the increasing histological dysplasia, the two become more consistent. There is a sequence in the process of cervical carcinogenesis, which means that HR-HPV infection causes the re-activation of the telomerase gene, resulting in unlimited proliferation of cervical cells12. Virus oncogene protein E6 of HPV may have a certain correlation with telomerase activation, and up-regulation of hTERC gene expression during telomerase activation may be the main pathway.HPV infection is closely related to the activation of telomerase. Studies on the carcinogenic mechanism of HPV show that in cervical cancer and cervical intraepithelial neoplasia, the proto-oncoprotein encoded by the oncogene E6/E7 can up-regulate telomerase activity after HR-HPV infection, leading to continuous proliferation of epithelial cells. Ohta 22.In this study, the HR-HPV rates in the normal, CIN1, CIN2/3, and ICC groups were 31.25%, 65.79%, 90.91%, and 92.86%, respectively. With increasing histological dysplasia of the cervical lesions, HR-HPV infection was aggravated, and hTERC gene expression was also increased, thus the two tended to be consistent. In this study the positive rate of hTERC was 54.63% (59/108), while the negative rate was 45.37% (49/108) in the HPV positive group. Whereas, the positive rate of hTERC was 11.90% (5/42) and the negative rate was 88.10% (37/42) in the HPV negative group. The difference was statistically significant, suggesting that that there was a correlation between hTERC gene amplification and HR-HPV infection, which is consistent with previous studiesThese results add to the growing evidence for the usefulness of hTERC as a biomarker in cervical cancer and precancerous lesions. But, because only long-term or repeated infection of HR-HPV may lead to hTERC gene amplification and the infinite proliferation of cervical cells, there are patients in this study without hTERC amplification in the HR-HPV infection group. hTERC is expressed in many normal and benign tissues, but up-regulation can reflect the degree of malignant progression of tissues and it is highly expressed in malignant tumors. Telomerase activity is closely related to the development of tumors. The hTERC gene plays an important role in telomerase activity. Therefore, inhibiting telomerase activity through hTERC might be used as a new method for treating tumors.The activation and regulation of a proto-oncogene is a very complex and continuous process. The occurrence and development of cervical lesions can be induced and activated by one gene, so the study of hTERC and its relationship with HPV infection needs to be further verified in more detailed studies.The strength of the study lies in the 1200 women who underwent screening and the inclusion of different grades of CIN and cervical cancer. The study is limited because it was a single center and the groups were relatively small. In addition, the potential role of HPV types in hTERC positivity was not explored. Larger studies from multiple centers are needed to fully evaluate the role of hTERC as a biomarker for cervical cancer.Significant hTERC gene amplification rates were found in precancerous cervical lesions and these rates increased with the increasing histological dysplasia of the cervical lesions through to ICC. This suggests that hTERC can be used as a biomarker for precancerous lesions of cervical cancer and might be expected to become an early screening method for cervical cancer. The cervical lesions were also associated with HR-HPV infection and there was a certain correlation between hTERC gene and HR-HPV infection.Outpatients and inpatients who voluntarily participated in cervical cancer screening at the Department of Gynecology of the Second Affiliated Hospital of Kunming Medical University between October 2010 and December 2011 were enrolled. All women had a history of sexual activity; were not pregnant; had not undergone colposcopy biopsy before; and were cervical treatment-naive. Each woman underwent a liquid-based cytology test (LCT), hybrid capture 2 (HC2)-HPV test and hTERC gene test. Cervical colposcopy biopsy was performed if LCT ≥ atypical squamous cell of undetermined significance (ASC-US) or positive HPV (≥1.0 pg/ml) or hTERC gene ≥ threshold determined from women with normal samples as described later. Two pathologists participated in the assessment. This study was approved by the Ethics Committee of the Second Affiliated Hospital of Kunming Medical University. Informed consent was waived due to the nature of retrospective study.A brush for LCT cell collection was inserted into the junction of the squamous epithelium and columnar epithelium of the external cervical orifice. With the external cervical orifice as the center, the brush was removed after completely rotating evenly 5–7 times. Then, the brush head was placed in the LCT cell preservation solution and shaken well, letting the collected cells disperse into the preservation solution and sent to the pathology department for detection. Cells remaining in the preservation solution were investigated for hTERC gene expression.A cervical sampling brush for HC2 was inserted into the cervical canal, which was rotated in the same direction 3–5 time to collect exfoliated cervical cells. Then, the specimens along with the brush were placed in the preservation solution to be sent for gynecological examination at our hospital using the HC2 HR-HPV Test Kit .Cervical cells were isolated from 5–10 ml of LCT cell preservation solution and treated with 3 ml of collagenase B solution at 37 °C for 20–30 min. The cells were pelleted and mixed with 5 ml of deionized water preheated to 37 °C and incubated at 37 °C for 20 min with occasional mixing. Stationary liquid (2 ml) was slowly mixed into the hypotonic mixture. The pellet was harvested and washed twice with 5 ml of stationary liquid for 10 min. The final pellet was resuspended in fixative solution and spread on a glass slide. After the slides had dried they were washed twice with 2× saline sodium citrate (SSC) solution (5 min each), then soaked in 0.1 mol/L hydrochloric acid solution for 10 min, 0.02 mg/ml pepsin working solution at 37 °C for 10 min, and then washed twice with 2× SSC solution (pH 7.0) (5 min each). The samples were dehydrated sequentially in 70% ethanol, 85% ethanol, and 100% ethanol, dried and then heated to 56 °C. The GLP hTERC/chromosome 3 centromere (CSP3) DNA probe mixture (10 μl)  was added, then immediately covered with a coverslip and sealed with rubber. Co-denaturation was performed at 75 °C for 8 min, then hybridization at 42 °C for 16 h. The coverslips were removed, and the glass slides were placed in 0.3% NP-40/0.4× SSC solution at 67 °C with shaking for 1–3 s, then rinsed for 2 min. The slides were then in 0.1% NP-40/2× SSC solution and shaken for 1–3 s, and rinsed for 30 s. The slides were placed in 70% ethanol, and rinsed for 3 min. DAPI re-staining agent (15 μl) was added to the target area, then covered with coverslips and kept in the dark for 10–20 min. Before observation with light microscopy. The whole hybridization area was scanned under a 40× objective lens and the quality of the specimens was observed. Satisfactory specimens had over 75% of the nuclei with hybridization signals. hTERC gene amplification in cervical cells was then observed under a 100× objective lens, and the signals were counted. The GLP hTERC specific probe was labeled with red fluorescence. The CSP3 probe labeled with green fluorescence was used as the control.Normal cells were those that gave two red and two green signals in a single interphase nucleus , the hTERC gene threshold in this study was 6.2%. Then, 100 cells from each study participant were randomly selected. If the percentage of cells was greater than the threshold, then the sample was positive. If the percentage of cells was less than the threshold, then the sample was negative. If the percentage of abnormal cells was equal to the threshold, then another 100 cells were randomly selected for detection.Statistical analysis was performed using SPSS 17.0 . Categorical variables are expressed as frequency (percentage) and analyzed using chi-square test. P < 0.05 was considered statistically significant.


Recent studies have shown a 13-fold increase of oropharyngeal cancer in the presence of HPV, while α-HPV detection seems to be rare in oral cavity in comparison to anal or cervical district, many novel β and γ types have been isolated in this anatomical site suggesting a wide tropism range. Currently, there are no guidelines recommending HPV oral cavity screening as a mandatory test, and it remains unknown which HPV types should be included in HPV screening programs. Our goal was to assess HPV prevalence in oropharyngeal, anal, and cervical swabs using different sets of primers,which are able to amplify α, β, γ HPV types.n = 124), anal (n = 186), cervical specimens (n = 43) from HIV positive and negative patients using FAP59/64 and MY09/11 primers. All untyped strains were genetically characterized through PCR amplification and direct sequencing of partial L1 region, and the resulting sequences were classified through phylogenetic analysis.We analysed the presence of HPV DNA in oropharyngeal (p < 0.05).HPV prevalence was 20.9% in 124 oropharyngeal swab samples, including infections with multiple HPV types (5.6%). HPV prevalence in this anatomical site was significantly associated with serostatus: 63.3%in HIV positive and 36.3% in HIV negative patients (1 species (2.4%) using FAP primers. Six of the untyped strains clustered with sequences from species 7, 9, 10, 8,12 of γ genus. Four sequences remained unclassified. Finally, β and γ HPV prevalence was significantly lower than their respective HPV prevalence as identified by the Luminex system in all anatomical sites that were analyzed in previous studies.Unclassified types were detected in 6 specimens. In our analysis, we did not observe any difference in HPV  prevalence between men and women. Overall, β species were the most frequently detected 69.7%. When using anal swabs, for HIV positive patients, β genus prevalence was 1% and γ genus was 3.7% including 6 unclassified types. In cervical samples from 43 HIV positive women (18 HPV negative and 25 positive by MY09/11 PCR), only one sample was positivite for βThis study provides new information about viral isolates present in oropharyngeal site and it will contribute to improve the monitoring of HPV infection.The online version of this article (10.1186/s12985-019-1132-x) contains supplementary material, which is available to authorized users. Human papillomavirus (HPV) persistent infections are considered the primary cause of ano-genital cancer, where greater than 99% of cervical cancer and more than 60% of anal cancer contain HPV DNA , 2. ThroTo date, there are no guidelines recommending HPV oral cavity screening as a mandatory test, and no HPV DNA test has been approved for HPV detection in oral cavity. In addition, it remains unknown which HPV types should be included in oral cavity screening. Agalliou  highlighIn this study, we analyzed the presence of HPV DNA in oral, anal, and cervical specimens collected from HIV positive and HIV negative individuals, living in the same geographic area (regione Lazio) by using MY09/11 , 21 FAP5This is a retrospective study carried out at Laboratory of Virology INMI L Spallanzani on residual oropharyngeal samples collected for respiratory virus detection, anal and cervical swabs collected for HPV testing in diagnostic routine. The Hospital ethics committee approved the protocol. Date of birth, date of swabs sampling, HIV serostatus were recorded from Institutional database. In HIV positive patients, the most recent CD4 T cell count (± 1 month from the date of sample collection) and HIV RNA viral load (± 1 month from the date of sample) with a detection limit of 40cp/ml  were used to correlate clinical features and HPV positivity.Oropharyngeal swabs -Oropharyngeal samples were collected as following: the nylon-flocked tip was rotated 3–4-times against right and left buccal mucosa, palatine, tonsils, upper and lower pharynx area. The swab was then plunged and stirred in 1.5 mL DMEM medium with streptomycin and ampicillin. Specimens were refrigerated within 3–5 h after collection until processing.Anal swabs -We retrospectively analysed 186 anal swabs previously tested by MY09/11 primers and typed by CLART HPV2 Clinical array or Sanger sequencing . One hundred samples belonged to HIV positive women (50 HPV positive by MY09/11 PCR and 50 negative by MY09/11 PCR), 86 anal swabs were collected from men who have sex with men (MSM). All MSM specimen resulted positive in MY09/11 PCR.- A total of 43 cervical swabs  were considered in this study. All samples belonged to HIV positive women.Cervical swabs General characteristics of HIV infected patients are presented in Additional file Oropharyngeal swabs were removed from the medium which we divided in two parts: half part was used for detecting respiratory viruses panel  and the other half was employed in testing HPV DNA, having been stored at − 80 °C until use. Before nucleic acid extraction, all specimens were pre-treated. Briefly, 600 μL of clinical material was digested with 20 μL proteinase K solution  and lysed with AL lysis buffer  at 56 °C for 10 min. Nucleic acid extraction was done with a magnetic bead-based automated platform  in accordance with the manufacturer’s instructions. Nucleic acids were eluted in 60 μL of AVE elution buffer . Nucleic acid from anal and cervical swabs were extracted as described elsewhere , 31.Samples that were β-globin negative were excluded from the study , 33.Ten μL of eluted nucleic acids were employed for evaluating the presence of HPV types by using MY09(5’CGTCCMARRGGAWACTGATC3’) and MY11(5’GCMCAGGGWCATAAYAATGG3’) , 21 PCR ’) and FAP 64 (5’CCWATATCWVHCATITCICCATC3’) primers [All adequate samples were retested using FAP 59 , including infections with multiple HPV types  (Table 2 species (Table 1 were the most represented (n = 13), while one sample was identified as γ type . Additionally, one mixed infection (Q1017) and 5 untyped HPV strains were also detected. Among β1 species, the most frequent types were HPV5  and HPV19 . Three multiple infections harboured β1 and β2 types, whereas β2 types and untyped strains were observed in 4 other multiple infections. No multiple infection harboured α with β or γ species . HPV distribution differed significantly when the number of HPV β, γ-types were considered in HIV positive and HIV negative subjects: 21  vs 12  respectively . HIV positive people with HPV in oropharyngeal swabs showed a mean CD4 T cell count of 532.3 ± 275.6. Among HIV positive patients with detectable HPV, 6/16 had no detectable HIV RNA in plasma and the other patients (n = 10) showed a mean RNA copies/mL169.0 ± 130.0.Out of 13, three samples were identified as α and ten as β types. Alpha-types were detected in two HIV positive patients , and in 1 HIV negative patient. HPV α-types, 16 and 70, were detected in HIV positive subjects  after being typed by CLART HPV2 clinical array, while HPV13 was amplified in HIV negative subject ; it was typed by Sanger sequencing. All β HPV types belonged to βp > 0.05).HPV prevalence rates were compared for statistical significance, using a Chi squared test, in men and women both HIV positive and HIV negative. However, no difference was observed  were retested with FAP primers.n = 0; AIN I, n = 5); atypical squamous cells of undetermined significance (ASC-US), n = 0; normal, n = 13) and 31 samples showed HPV multiple infection, all with at least 1HR type (62%) . HPV type was undetermined in one sample. Among single infection with AIN I, one specimen was infected with a low risk (LR)Type (HPV81). Nineteen anal swab specimens were identified as HPV positive by FAP primers detection  (Table 2 types (HPV9 and HPV37) were found in two specimens co-infected with α HPV types; 2 specimens harboured γ10 HPV types (HPV121 and HPV180), both associated with type α HPV multiple infections. All FAP untyped strains (n = 6) were found in anal specimens infected by α types with at least one high HR type. Considering cytological aspects, 12/19 samples, which resulted positive with FAP primer PCR, had AIN  lesions, while 7/19 samples showed normal cytology  type. In 61.2% of samples we observed a clinically evident anal pathology, including 7 subjects with anal intraepithelial neoplasia (AIN) II, 46 patients with AIN I. Among HPV positive women, 18 harboured a single infection , only 3 samples (3.0%), previously tested MY09/11 HPV positive, resulted HPV positive with FAP primer . No untyped HPV strains were observed among female samples.Among anal swabs from females .Mean of CD4 T cell count was 690.7 ± 343.9 cells/μL. Overall 170/186 patients (91.4%) were under antiretroviral therapy (ART). All patients without ART had a CD4 T cell count > 400/ μL. Current ART use was not associated with risk of β and ɣ infection  showed normal cytological findings. Eleven/43 cervical samples resulted FAP positive (25.6%). However, only 6 HPV types were not previously detected by MY09/11 primers. Five samples harboured α types , and only one specimen gave positivity for β1 (HPV14) (2.4%). The mean CD4 T cell count was 539 ± 230 cells/μL. Ninety-six percent of women were receiving ART per WHO guideline of starting ART at CD4T cell count < 350 cell/mm3. The woman harboring β HPV strain was under ART. She had a CD4 T cell count of 540 cells/μL and HIV RNA viral load was not detected.Cytology findings were available for 20 specimens already resulted positive by CLART HPV2 clinical array: 10 had normal cytology, 6 were ASC-US or low grade squamous intraepithelial lesion (LSIL), 3 harboured HPV single infections, 3 showed HPV multiple infections, and 4 were high grade squamous intraepithelial lesion (HSIL). All MY09/11 HPV negative women , while other authors found higher prevalence when they used a platform able to detect HPV types belonging to genera other than α genus  both in nd study , was absnd study , no diff3, β4, β5 HPV types. This may be due to the types of biological samples that were used or to differences in the efficiency of extraction methods that were applied, which could have influenced the outcome of PCR [1 genotypes, HPV12 and HPV5 were the prevalent genotypes. While, among β2 strains, HPV38 was the most represented,and it was never detected in our samples. Interestingly HR HPV38 was also the main β2 type observed also in Moscicki study, whereas HPV21 represented the main β1 type [2 types, HPV9 and HPV37, that had not been reported among oropharyngeal swabs. Βeta types frequency was sensibly lower to that found by Luminex system reported by Torres et al. [1 species, while HPV38 and HPV120 were the most frequently observed β2 types. Among γ genus, the γ10 species was the most prevalent in both MSM HIV positive and -negative groups, while we observed only γ untyped strains. Donà reported higher prevalence of β (27.6%) and γ types (29.3%) in a group of MSM similar to ours using the Luminex system [Unlike Forslund and Hampras , 43, we e of PCR . Finally β1 type . A diffe β1 type , where s β1 type . This mas et al.  which fox system . Some hyx system . To obta14; HPV32, α1) which were not detected by CLART HPV2 clinical array/MY09-11primers and 1 β1 type (HPV14). This low prevalence of β types confirms the data reported by Bottalico et al. [In cervical swabs the FAP primers mainly showed the presence of α genotypes  surpassing α and β genera, and that 77% of the new types deposited in the HPV center within 2015 belonged to γ genus –56, our Additional file 1Table S1 Study population characteristics of HIV infected patients. (DOCX 14 kb)


Many countries are implementing human papillomavirus (HPV)‐based cervical screening due to the higher sensitivity of the test compared with cytology. As HPV is sexually transmitted, there may be psychosexual consequences of testing positive for the virus. We aimed to review the literature exploring the psychosexual impact of testing positive for high‐risk cervical HPV.MEDLINE, PsycINFO, CINAHL Plus, Web of Science, and EMBASE were searched with no date limits. We also searched the grey literature, reference lists of included articles and carried out forward citation searching. Eligible studies reported at least one psychosexual outcome among HPV‐positive women. Qualitative and quantitative papers were included. We extracted data using a standardised form and carried out a quality assessment for each article. We conducted a narrative synthesis for quantitative studies and a thematic synthesis for qualitative studies.Twenty‐five articles were included. Quantitative study designs were diverse making it difficult to determine the impact that an HPV positive result would have in the context of routine screening. The qualitative literature suggested that psychosexual concerns cover a broad range of aspects relating to women's current and past relationships, both interpersonal and sexual.The psychosexual impact of testing positive for high‐risk cervical HPV is unclear. This review highlights the need for further research in the context of HPV‐based cervical screening. As primary HPV testing is introduced more widely, it is important to understand women's responses to testing HPV positive in the cancer screening context to minimise any adverse psychosexual impact. In the primary HPV testing pilot in England, approximately 13% of the screened population received an HPV positive result.An essential criterion for any screening programme is that the overall benefits should outweigh the harms2This review was registered with PROSPERO (CRD42018083969) and followed the Preferred Reporting Items for Systematic Review and Meta‐Analysis (PRISMA) guidelines.2.1http://www.opengrey.eu), PsycEXTRA, the reference lists of included articles, and forward citation searching.The search included terms relating to (a) high‐risk cervical HPV and (b) a psychosexual or disclosure‐related outcome  and were linked using Boolean operators (see 2.2Studies were included if they mentioned (a) HPV and (b) a psychosexual or disclosure‐related outcome. Reviews, conference abstracts, commentaries, opinion pieces, and editorials were excluded. Studies were also excluded if they were not in English, explicitly focused only on low‐risk HPV or focused on the psychosexual impact of cervical cancer, treatment for cervical cancer, or colposcopy.Titles were screened by K.B. Two reviewers (K.B. and M.R.) screened the abstracts of the remaining papers (agreement rate = 85%). Where a paper could not be assessed using the abstract, the fulltext was obtained. Disagreements were resolved by discussion.2.3Using a standardised data extraction form see  2, one r2.4The quality of studies was assessed using modified versions of the National Institute for Health and Care Excellence (NICE) quality appraisal checklists for quantitative and qualitative studies see  3 and 4.2.5Quantitative and qualitative findings were analysed separately. For quantitative studies, a narrative synthesis was conducted and the results reported descriptively. We used Popay et al'sFor qualitative studies, we conducted a thematic synthesis, following the three stages outlined by Thomas and HardenA coding frame was developed and applied to the data (by K.B.). A second reviewer (M.R.) independently coded 20% of these papers, and any inconsistencies were resolved through discussion.33.1The search yielded 4801 articles after the removal of duplicates. Following exclusions, 40 fulltexts were reviewed. Twelve articles were excluded during the full‐text review, and two were included following backward/forward citation searches, resulting in 30 papers , United States (n = 5), Taiwan (n = 4), Australia n = 2), Greece, Hong Kong, Italy, China, Brazil, Sweden, and Belgium all n = 1) and were published between 1988 and 2018. Studies were quantitative . The quality of external validity was mixed. Most qualitative studies were judged to be well conducted n = 12)  found that, in the control group (who only received counselling), women who were HPV+ had slightly greater satisfaction with intimate relationships than women who were HSV+; however, in the experimental group women with HPV had slightly lower satisfaction with intimate relationships than women with HSV. In this study, the HPV and HSV groups were not statistically directly compared, and the range of potential scores was not reported.In a descriptive study of 51 women who had recently been informed that they were HPV+,3.3.3Four studies assessed frequency of sex following an HPV+ result.Two studies reported no difference in frequency of sex between women who were HPV+ and women who were HPV−.3.3.4Four studies assessed interest in sex, thoughts about sex, and sexual arousal following HPV diagnosis.3.3.5Four studies assessed feelings about partners and relationships.Two studies found no evidence that an HPV+ result affected feelings about partners or relationships.3.4A thematic synthesis of 13 studies identified three major themes relating to psychosexual impact: (a) source of HPV infection, (b) transmission of HPV, and (c) impact of HPV on sex and relationships. 3.4.1A common response from women with HPV was to question which partner (current or previous) the infection had come from.Some women expressed concerns that their partner had been unfaithful.3.4.2Concern about passing HPV on to a partner was common.Worry about re‐infection and recurrence was common.3.4.3Whilst some women were concerned HPV might negatively impact their relationshipSeveral studies identified a reduced interest in and frequency of sex,dirty,” “contaminated,” and unworthy of sexual attention.HPV had a negative impact on some women's sexual self‐image.The risks associated with oral sex were mentioned by a few women4This review synthesises the existing literature on the psychosexual impact of testing positive for high‐risk cervical HPV. The diversity of quantitative study designs and inclusion of study populations with abnormal cytology or other conditions makes it difficult to determine the impact that an HPV+ result would have in the context of routine primary HPV testing; however, some studies suggested that testing HPV+ can have a psychosexual impact. The qualitative literature suggested that psychosexual concerns are raised by some women who test HPV+ and that these concerns cover a broad range of aspects relating to their current and past relationships, both interpersonal and sexual.Including quantitative and qualitative articles in the review allowed us to highlight the range of psychosexual concerns that women testing HPV+ have. Traditional psychosexual measures used in the quantitative studies assessed specific aspects of sexual behaviour in line with medical classifications of psychosexual disorders . The heterogeneity in study design and time from receipt of HPV test results to when data were collected could provide an explanation for the mixed findings, and this makes it difficult to form conclusions about the prevalence and severity of the psychosexual impact of an HPV+ diagnosis. Whilst some studies included in the review did use validated measures, a validated measure specific to HPV testing that assesses aspects of psychosexual and interpersonal relationships  would help to ensure contextually valid items are included and provide a tool that can allow comparisons between studies. Only two papers included in the review measured psychosexual impact longitudinally. Future studies should measure the psychosexual impact of testing HPV+ over time to ascertain if psychosexual impact changes. Knowledge of when psychosexual impact is greatest could help to determine when interventions are most appropriate.4.1Since the quantitative papers included a range of psychosexual outcomes, it was not possible to conduct a meta‐analysis. Whilst we excluded any articles that explicitly focused on low‐risk types of HPV, some of the papers included in the review did not describe the type of HPV participants had and it is possible that some articles included participants with low‐risk HPV.4.2It is important to understand, and minimise, any psychosexual impact of testing HPV+ in the context of primary HPV testing. In line with previous studies , the qualitative synthesis highlights that women who test HPV+ have a number of questions about HPV such as the source of the infection, whether partners can re‐infect each other and how to prevent the transmission of HPV. Information materials could increase knowledge and address some of these concerns. Additionally, health care professionals carrying out cervical screening could be trained to give brief information during screening to ensure that women understand their results when they receive them. Whilst HPV is classified as an STI, it differs from other STI's as it is normally asymptomatic, does not need treatment, and does not usually cause any long‐term problems. Communicating this information to women is important and may help to reduce psychosexual impact.5This review synthesises the literature on the psychosexual impact of testing HPV+. The qualitative studies included in the review provide rich information about the source and nature of psychosexual distress experienced by some women. In particular, women were concerned about transmitting HPV to a partner and where the HPV infection came from. The diversity of quantitative study designs and samples makes it difficult to draw conclusions about the magnitude of psychosexual impact in the context of primary HPV testing. Whilst this review draws together what is currently known, it also highlights the need for further quantitative and qualitative research in the context of primary HPV testing. It is important to understand the psychosexual impact of testing HPV+ in a routine context to minimise undue concern among women, and to avoid compromising future screening re‐attendance.KB is funded by a Medical Research Council (MRC) studentship (grant reference: MR/N013867/1). JW, MR and LM are funded by a Cancer Research UK career development fellowship awarded to JW (grant reference: C7492/A17219).No conflicts of interest to declare.Table S1. Search strategyClick here for additional data file.Table S2. Data extraction formClick here for additional data file.Table S3. Quality appraisal checklist—quantitative studiesClick here for additional data file.Table S4. Quality appraisal checklist—qualitative studiesClick here for additional data file.Table S5. A brief description of themes relating to the psychosexual impact of testing positive for high‐risk cervical HPV and the studies associated with themClick here for additional data file.


Current guidelines suggest annual co-test follow-up in non-16/18 high-risk HPV positive patients without abnormal cytologic findings. Considering the relatively high false-negative rates of cervical cytology, a follow-up period of one year might constitute an additional risk for development of cervical malignancies in these patients. The current study aims to evaluate colposcopic biopsy results of cytologically normal patients detected to carry high risk HPV by screening tests. A total of 160 patients with normal cervical cytology and high-risk HPV subtypes who underwent colposcopic examination were included in the study. Patients were divided into two groups according to their HPV results: HPV 16-18 and other types (non-16/18 high-risk subtypes). ECC, cervical biopsy, LEEP/conization results were analyzed for both groups. Histopathological results of the groups were compared in terms of ≤LSIL, HSIL and cervical cancer rates. Cervical biopsy results in the “16-18” group were assessed as HSIL in 40 (48.2%) patients, LSIL in 6 patients (7.2%) and normal in 37 (44.6%) patients. ECC results revealed HSIL in 9 (10.8%) patients and LSIL in 1 (1.2%) patient. Nineteen out of 42 patients who underwent LEEP/conization had HSIL , while 3 patients had LSIL. None of the cases had cervical carcinoma.The present study detected that 15.6% of women infected with non-16/18 high-risk HPV subgroups developed ≥HSIL lesions. Although this rate seems lower than HPV 16-18 group, it is still too high to be overlooked. In conclusion, we suggest further clinical trials with larger number of patients to be conducted on this topic. National Cancer Institute, Centers for Disease Control and Prevention, Jemal et al., (2013) and Viens et al., (2016) report that HPV infection is associated with nearly all (more than 90%) of cervical cancers . HPV 16 alone is responsible from half of all cervical cancers, while HPV 18 is responsible from 20% . Fifteen HPV subtypes  are defined as high-risk (hrHPV) for the development of cervical cancer by Centers for Disease Control and Prevention, 2011; Jemal et al., 2013)The ASCCP guidelines recommended immediate colposcopy in HPV 16/18 positive cases, regardless of cytology results, while co-testing after 1 year is suggested in the presence of other hrHPV subtypes with a negative cytology . Two studies conducted in Turkey reported 20% of women with normal cervical cytology to be HPV-DNA positive and 32.5% of hrHPV positive women to have normal cervical cytology . Various studies have reported the false negative rate of cervical cytology to be between 15% and 63% . Additionally, patients might interrupt their annual follow-ups for various reasons. The aim of this study is to compare colposcopic biopsy results of HPV 16/18 positive patients and patients carrying at least one of the hrHPV subtypes both with normal cervical cytology.® HPV kit (Roche) for HPV genotyping were utilized. A total of 160 patients who were examined for at least one of the high-risk DNA subtypes and were found to have no abnormal findings on cervical cytology were included in the final statistical evaluation. All patients were evaluated by a gynecologist experienced in colposcopy (HA). Patients were divided into two groups according to HPV results as HPV 16/18 positive  and as non-16/18 hrHPV subtypes . Endocervical curettage (ECC), cervical biopsy, LEEP/conization results of both groups were analyzed. Histopathologic results of the groups were compared in terms of ≤LSIL, HSIL and cervical squamous cancer rates. Eighty-three patients in the HPV 16/18 group and 77 patients in the ‘OTHERS’ group were evaluated.This retrospective cohort study has been conducted between January 2015 and December 2017 in a tertiary center located in western Turkey at the Clinic of Obstetrics and Gynecology, Izmir Katip Çelebi University Ataturk Training and Research Hospital. A total of 167 women referred to our hospital due to HPV DNA positivity from primary screening centers were included in the study. The study protocol was approved by the İzmir Katip Çelebi University Ethics Committee (2017-308). Patients who did not accept colposcopy and biopsy, or had previous history of cervical dysplasia were excluded from the study. Informed consent was obtained from all participants and the study was in agreement with the Declaration of Helsinki for Medical Research Involving Human Subjects. Cervical cytologic evaluation was performed by conventional or liquid-based cytology. Abnormal histopathologic findings were classified as low-grade squamous intraepithelial lesion (LSIL), high-grade squamous intraepithelial lesion (HSIL), and squamous cell carcinoma (SCC). Hybrid Capture2 (Qiagen) kit for HPV-DNA scans and CLART kit (Genomica) or CobasThe median age of the “16-18” and the “others” group was 44 (28-66) and 45 (30-66), respectively. While 72.3% of the patients were premenopausal and 27.7% were postmenopausal in the “16-18” group, 68.8% were premenopausal and 31.2% were postmenopausal in the “others” group. Median parity of the both groups was 2 (0-6 in the “16-18” group and 0-5 in the “others” group) . Cervical biopsy results in the “16-18” group were assessed as HSIL in 40 (48.2%) patients, LSIL in 6 patients (7.2%) and normal in 37 (44.6%) patients. ECC results revealed HSIL in 9 (10.8%) patients and LSIL in 1 (1.2%) patient. Nineteen out of 42 patients who underwent LEEP/conization had HSIL , while 3 patients had LSIL. None of the cases had cervical carcinoma.The most common subtype in the “Others” group was found to be HPV 52 (15.5%). Cervical biopsy results were assessed as HSIL in 12 (15.6%) patients, LSIL in 6 (7.8%) patients and normal in 59 (76.6%) patients. ECC results revealed 1 HSIL (1.3%) and 1 LSIL. Eight out of 12 patients who underwent LEEP/conization had HSIL , while none had LSIL nor cervical carcinoma.Evaluation of the groups regarding histopathological findings showed that 48.2% of the cases in the “16-18” group had ≥HSIL lesions while only 15.6% of the “others” group had ≥HSIL lesions (p˂0.01). Presence of HPV 16-18 resulted in 5-fold increased risk of developing ≥ HSIL lesions compared to other high-risk HPV subtypes (OR: 5.03% 95% CI 2.3-10.6).We found that the detection rate of ≥HSIL lesions was significantly higher in HPV 16/18 positive cases compared to other hrHPV subtypes . The most frequent subtype except HPV 16/18 was HPV 52. Current guidelines recommend annual co-test follow-up in non-16/18 hrHPV infected women with negative pap-smears, since the risk of cervical preinvasive lesion or cancer development is directly proportional with persisting HPV infection and passing time . Delayed diagnosis might lead to patients encountering severe epithelial changes . However, it has been reported that false negative rate of pap-smear test is about 15–65% . In this case, non-invasive follow-up of non-16/18 hrHPV cases for 1 year should be questioned due to the probability of increased risk for cervical dysplasia. A study investigating detectability of biopsy-confirmed high-grade cervical lesions by pap-smear reported that false low-grade cytomorphology rates were higher in non-16/18 hrHPV and mixed hrHPV infected women than HPV 16/18 infected women. In light of this the authors suggested that diagnosis-treatment approach might be delayed due to increased pap test underdiagnosis rates . Another problem is the lack of annual co-test follow-up compliance by patients. In a study nearly half of the patients were lost to follow-up . Despite the widespread incidence of hrHPV infections especially among young females, majority of infections are transient and spontaneously eliminated . Moreover, slow progressive nature of the infection prolongs the development time of an invasive lesion, thereby increasing chances of recognition in the precancerous stage . A previous study reported that detection rates of CIN3+ cervical lesions by primary HPV screening were 50% higher than scanning by cytology, however the number of colposcopies were doubled simultaneously . In our study, comparison of initially negative cytology HPV 16/18 positive patients with non-16/18 hrHPV positive patients regarding development of ≥ HSIL lesions yielded a 5-fold increased risk in the HPV 16/18 group. Although that risk is high enough to be ignored, given the slow natural course of the disease, potentially treatable preinvasive lesions, and increased demand for personnel and equipment due to colposcopic examination of cytology negative hrHPV positive patients, recommending annual co-test follow-up to smear negative non-16/18 hrHPV positive cases sounds reasonable. In conclusion, the risk of cervical preinvasive lesions among hrHPV infected cases without cervical cytomorphological abnormalities is 5-fold higher in HPV 16/18 positive cases. Although follow-up at one year is theoretically risky in non-16/18 hrHPV positive but cytology negative women, annual follow-up is a reasonable approach due to slow natural course of the disease. Authors declare no conflict of interest or any funding.