EGFR, TP53 and HSPA1A/B overexpressed in HPV(+) cells and IL6, FAS and DPP4 in HPV(−) cells. The mRNA profiles of HPV(+) or HPV(−) exosomes resembled the cargo of their parent cells. miR expression profiles in cell lysates identified 8 miRs expressed in HPV(−) cells vs. 14 miRs in HPV(+) cells. miR-205-5p was exclusively expressed in HPV(+) exosomes, and miR-1972 was only detected in HPV(−) exosomes. We showed that HPV(+) and HPV(−) exosomes recapitulated the mRNA expression profiles of their parent cells. Expression of miRs was dependent on the HPV status, and miR-205-5p in HPV(+) and miR-1972 in HPV(−) exosomes emerge as potential discriminating HPV-associated biomarkers.Human papillomavirus (HPV)(+) and HPV(−) head and neck cancer (HNC) cells’ interactions with the host immune system are poorly understood. Recently, we identified molecular and functional differences in exosomes produced by HPV(+) vs. HPV(−) cells, suggesting that genetic cargos of exosomes might identify novel biomarkers in HPV-related HNCs. Exosomes were isolated by size exclusion chromatography from supernatants of three HPV(+) and two HPV(−) HNC cell lines. Paired cell lysates and exosomes were analyzed for messenger RNA (mRNA) by qRT-PCR and microRNA (miR) contents by nanostring analysis. The mRNA profiles of HPV(+) vs. HPV(−) cells were distinct, with  Head and neck cancers (HNCs) belong to the ten most common malignant diseases worldwide . Due to TP53 is mutated in 84% of HPV(−) and only 3% of HPV(+) HNCs [To date, it remains unclear why HPV(+) patients respond better to therapy and have a better prognosis than patients with HPV(−) cancers. The viral hypothesis associating the HPV positivity with a more robust anti-tumor immune response of cytotoxic T cells is probably most widely considered ,12. It s(+) HNCs . HoweverWe have recently reported that exosomes produced by HPV(+) HNC tumor cell lines or derived from plasma of HPV(+) patients are molecularly and functionally distinct from HPV(−) exosomes . These dExosomes are a subset of small extracellular vesicles (EVs) sized at 30–150 nm, which are produced by all cells and circulate freely in all body fluids. Exosomes have a unique biogenesis: they are formed in the endocytic compartment of parent cells in multivesicular bodies (MVBs) and upon MVB fusion with the cellular membrane are released into tissue spaces . The exoHere, we used the same exosomes isolated from supernatants of the HPV(+) and HPV(−) HNC cell lines to study genomic mRNA and microRNA (miR) profiles of these vesicles. We report that the mRNA profiles of HNC cells and exosomes these cells produced were similar, although HPV(+) cells and exosomes were enriched in HPV-related transcripts. Analysis of miR profiles in cells and exosomes also indicated similarity and showed that fewer miRs were present in exosomes compared to cells. Surprisingly, only a single miR (miR 205-5p) was uniquely expressed in HPV(+) exosomes, while another miR (miR 1972) was only present in HPV(−) exosomes.Exosomes isolated from supernatants of all cell lines showed typical sizes, concentrations and characteristics as previously reported by us . The HPVEGFR, TP53 and HSPA1A/B. The mRNA transcripts for IL6, FAS, DPP4 and CCND1 were predominantly expressed in HPV(−) cell lines and their exosomes.The heatmaps in HPV16E6 and HPV16E7, were exclusively expressed in HPV(+) cell lines as well as in HPV(+) exosomes. CDKN2A which codes for p16 protein, commonly used as surrogate marker for HPV in clinical practice, was overexpressed in cell lines and exosomes from SCC-47 and exosomes from SCC-90. Overall, the immunostimulatory genes, including HSP90, PIK3CA and SMAD4, as well as immunosuppressive genes, such as STAT3 and TGFB/TGFBR, were expressed at comparable levels and independently of the HPV status of exosomes or cells.The supervised heatmap analysis grouped the mRNA profiles according to their functional impact B. As expWaterfall plots  were useHPV16E6 and HPV16E7 mRNAs compared to cell lines, which could be due to the concentration of these mRNAs within the exosomes.The horizontal (x) axis represents the baseline, on which both measures are equally expressed. The vertical (y) axis is used to visualize the relative amount of mRNA, which is indicated by the different vertical bars. FASLG, PDCD-1 (=PD-1) and CD70 were higher in the cell lines than in exosomes  samples. Moreover, the immunoregulatory genes TGFBR2, TNFSF10, DPP4 and FAS show higher values for HPV(−) than HPV(+) samples  exosomes. Overexpression of CDKN2A, EGFR, TNFSF4 and CXCR4 was detected in HPV(+) exosomes (CD70 in HPV(+) cells (In  samples A. The diexosomes B. Simila+) cells C.Nanostring analysis identified 798 endogenous miRs in all the cell lines and respective exosomes. False discovery rate (FDR) estimation was used to identify significant differences in miR levels in cell lysates and exosomes. The data were analyzed using volcano plots, which indicated the fold differences of miRs expression levels in exosomes vs. cell lysates or the HPV status: HPV(+) vs. HPV(−).The list of specific miRs differentially expressed with the highest significance in the analyzed subgroups is presented in The subgroup analysis of HPV(+) cell lines revealed 69 significantly overexpressed miRs and 55 overexpressed miRs in exosomes, respectively , while tCombining lysates and exosomes of HPV(+) or HPV(−) cell lines, we observed statistically significant overexpression of 8 miRs in the HPV(−) subgroup and 14 miRs in the HPV(+) subgroup A. FocusiThe interaction analysis revealed two miRs: miR-1972, which are most likely found in HPV(−) exosomes, and let-7i-5p for HPV(+) cell lysates .These results indicate that the miR cargo of HNC cell-derived exosomes differs from the miRNome of the cell that produced these exosomes. Moreover, our data indicate that HPV(+) and HPV(−) HNC cells express different signatures of miRs and release exosomes with a different miR content, suggesting that miRs in exosomes may be predictive of the HPV status of the cell that produced these exosomes.We and others have reported that exosomes produced by human tumor cells, TEX, exert suppressive effects on immune cells and re-program their functions ,26,27. RHPV16E6 and HPV16E7 were exclusively expressed in HPV(+) lines as well as in HPV(+) exosomes, confirming the ability of exosomes to discriminate the HPV(+) from HPV(−) tumors, although mRNA levels for p16 were reported to be elevated only by some HPV(+) cell lines and exosomes  vs. HPV(−) cells. The supervised and unsupervised analyses of HPV(+) and HPV(−) heatmaps indicated that mRNA transcripts in cells were comparable to those carried in exosomes. However, considerable differences emerged between mRNA profiles of cells and exosomes based on the HPV status. Importantly, the HPV-associated markers, exosomes . HoweverPIK3CA and SMAD4, TGFB/TGFBR and STAT3, were expressed at comparative levels in cells and exosomes independently of their HPV status. These data are in agreement with the protein and functional data reported by us previously, which did not discriminate between HPV(+) and HPV(−) exosomes with respect to their immunosuppressive effects on T lymphocytes [CCND1, CDH1 and MET were found to be overexpressed in HPV(−) exosomes as were the immunoregulatory genes TGFBR2, TNFSF10, DPP4 and FAS relative to their expression in HPV(+) exosomes. This suggests that HPV(−) exosomes might play a role in promoting tumor growth and in tumor protection from immune elimination, as we have previously suggested [CDKN2A, EGFR, TNFSF4 and CXCR4 was detected in HPV(+) exosomes.Further, mRNA transcripts for various immunoregulatory genes, including phocytes . On the uggested . In contuggested . OverexpTAP1 [The miR contents of the HNC cell lysates used here were previously characterized by Wald et al. . We haveTAP1 , suggestOverall, our genomic analysis of exosomes produced by HNC cell lines indicates that exosomes can discriminate the HPV(+) from HPV(−) cells. These results are in agreement with the proteomic and functional data we previously reported using the same HPV(+) and HPV(−) cell lines . Furtherv/v) exosome-depleted and heat-inactivated fetal bovine serum  and 1% (v/v) penicillin/streptomycin in 150cm2 flasks. After 48–72 h, cells were expanded from 40% to 80% confluency and supernatants were collected for further exosome isolation.HPV(+) cell lines  were obtained from Dr. Robert L. Ferris, HCC), and HPV(−) cell lines (PCI-13 and PCI-30) were established in our laboratories as previously reported . The tumg for 10 min at room temperature and at 14,000× g for 30 min at 4 °C, and ultrafiltrated by 0.22 µm syringe-filters . Pre-clarified supernatants were concentrated 50-fold using Vivacell 100 filters . Aliquots (1 mL) of clarified, concentrated culture supernatant were loaded on mini-SEC columns [Freshly harvested culture supernatants were purified by differential centrifugation at 2000×  columns  and elut columns .g for 2 h, and the pelleted exosomes or HNC cells were resuspended in 350 µL of lysis buffer from the Exiqon RNA isolation kit. Total RNA was extracted following the manufacturer’s instructions. Quantity and quality of total RNA content was assessed using Nanodrop. RNA samples were aliquoted and sent to Children’s Hospital Los Angeles (CHLA) for miR analysis and to the genomics facility at the University of Pittsburgh for mRNA assessment.The mini SEC fractions #3 and #4 were ultracentrifuged at 100,000× http://mev.tm4.org). Relative dCt values were calculated for all exosome specimens in relation to their individual cell line or in relation to the HPV-status.The exact amount of total RNA was determined using Qubit Fluorometric quantitation . Reverse transcription was performed on 100 ng of total RNA in triplicate using QuantiTect Reverse Transcription Kit . PCR was performed with cDNA of 1 ng RNA equivalence per well. All real-time PCRs were run on Applied Biosystems 7900HT . Newly designed primers for E6 and E7  were valThe data comprised results of miR count values from nanostring assays for exosomes and cell lysates from five cell lines, two of which were HPV(−) (PCI-13 and PCI-30) and three HPV(+) . Three replicate assays were performed for each cell line for both lysate and exosomes.The nanostring assay identified 828 miRs. Of these, 798 were classified as endogenous, 5 housekeeping, 6 ligations, 8 negative controls, 6 positive controls, and 5 spike-ins. This analysis was restricted to endogenous miRs.p-value . Note that “hsa” has been removed from the miR names in the plots and p < 0.05) is highlighted by a red line in the graphs.Endogenous miR count values were transformed to the log10 scale and then quantile-normalized along with data from six assays from an unrelated HUVEX experiment . For eac


VE against oropharyngeal HPV‐16/18, HPV‐31/45 and HPV‐31/33/45 infections were 82.4% , 75.3% (95%CI: 12.7–93.0) and 69.9% (95% CI: 29.6–87.1), respectively. In conclusion, the AS04‐HPV‐16/18 vaccine showed effectiveness against vaccine and nonvaccine HPV‐types oropharyngeal infections in adolescent females up to 6 years postvaccination.We studied effectiveness of the AS04‐adjuvanted HPV‐16/18 (AS04‐HPV‐16/18) vaccine against human papillomavirus (HPV) oropharyngeal infections associated with the increase of head/neck cancers in western countries. All 38,631 resident adolescents from 1994 to 1995 birth cohorts of 33 Finnish communities were invited in this community‐randomized trial (NCT00534638). During 2008–2009, 11,275 girls and 6,129 boys were enrolled in three arms of 11 communities each. In Arm A, 90% of vaccinated girls/boys, and in Arm B, 90% of vaccinated girls received AS04‐HPV‐16/18 vaccine. Other Arm A/B and all Arm C vaccinated participants received control vaccine. All Arm A participants and Arm B female participants were blinded to vaccine allocation. Oropharyngeal samples were analyzed from 4,871 18.5‐year‐old females who attended follow‐up visit 3–6 years postvaccination. HPV DNA prevalence was determined by SPF‐10 LiPA and Multiplex type‐specific PCR. Total vaccine effectiveness (VE) was defined as relative reduction of oropharyngeal HPV prevalence in pooled Arms A/B HPV‐vaccinated females  What's new?Cervarix vaccine against oropharyngeal HPV‐16/18 infection and moderate effectiveness against non‐vaccine types HPV‐31/33/45 infection in female adolescents. The protective effects were observed up to 6 years after vaccination. These results provide further evidence that HPV vaccination holds the potential to reduce oral HPV infection, thereby offering protection against HPV‐related head and neck cancers.The incidence of oropharyngeal cancers due to human papillomavirus (HPV) infection is increasing in Western countries. However, the efficacy of HPV vaccines against oropharyngeal infection is not well documented. Based on a large community‐randomized trial in Finland, the authors report high effectiveness (82.4%) of  AlaluminumAS04‐HPV‐16/18AS04‐adjuvanted HPV‐16/18 vaccineCIconfidence intervalGCPgood clinical practiceHBVhepatitis B virusHPVhuman papillomavirushrHPVhigh‐risk human papillomavirusesIARCInternational Agency for Research on CancerMPLmonophosphoryl lipid An (%)number (percentage) of subjects reporting an eventNnumber of subjects with available resultsOPSCCoropharyngeal squamous cell cancerVEvaccine effectivenessOncogenic, high‐risk human papillomaviruses (hrHPV) cause worldwide 9% of cancers in females and 1% in males.In western countries, the incidence of HPV‐positive OPSCC has increased very rapidly, virtually doubling in a decade since the 1970s.In the adolescent and young adult populations, the prevalence of oral HPV infections varies widely from 0.4% to 10% mostly depending on the method used for viral DNA detection.All the 33 randomized Finnish communities had more than 35,000 inhabitants.clinicaltrialgov.com) and its amendment for the collection of the oral gargle samples were approved by the ethical committee of the Pirkanmaa hospital district in 2007 and 2011. The trial was conducted in accordance with good clinical practice (GCP) and all applicable regulatory requirements including the Declaration of Helsinki. Participants aged <15 years provided written informed assent; written informed consent was obtained from the participants’ parents or legal representatives. Written informed consent was obtained from study participants aged 15 years in line with local regulations.The primary HPV‐040 trial  vaccine, and 10% received Engerix B vaccine .3+). In Arm B, 90% of vaccinated females received the AS04‐HPV‐16/18 vaccine, and 10% of vaccinated females and all vaccinated males received the HBV vaccine. The 90 and 10% proportions were receiver‐blinded proportions to assess the herd effect. The blinding was maintained for girls and boys in Arm A and for girls in Arm B. In Arm C communities, all vaccinated participants received the HBV vaccine. Virtually all (99%) vaccinated participants received all three vaccine doses.The vaccination (first three study visits) took place at the healthcare facilities of 250 municipal junior high schools of the study site communities. In Arm A, 90% of vaccinated participants received Both vaccinated and nonvaccinated females of the 1994–1995 birth cohorts from the study site communities were invited to attend follow‐up visit at the age of 18.5 years during 2012–2014. Cervical and oropharyngeal samples for HPV DNA testing were obtained on each visit. Setting the age for attending the follow‐up visit at 18.5 years was per protocol to allow a minimum of 3 years between vaccination and cervical sampling.During the follow‐up visit, the attendees filled‐in a questionnaire on living conditions, life‐habits and sexual behavior. At age 18.5 years, a cross‐vaccination with either the HBV vaccine or the AS04‐HPV‐16/18 vaccine was offered to the attendees. Female attendees (139), who had moved between Arm C and Arm A or B communities during the follow‐up by age 18.5 years were removed from the final analyses.All samples were analyzed by PCR for HPV DNA. HPV typing was performed by a broad‐spectrum PCR SPF10‐LiPA25  using HPV‐specific hybridization probes enabling detection of 14 oncogenic HPV types  and 11 nononcogenic HPV types . To ensure maximum sensitivity in the detection of HPV‐16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68, 6 and 11, samples initially considered to be SPF‐10/DEIA positive for HPV were re‐evaluated by the multiplex type‐specific PCR.The number of subjects invited to participate in the study and the number of subjects enrolled was tabulated by gender, for birth cohorts 1994, 1995 and overall.The analysis of VE was done on female study participants and was based on the total enrolled cohort which included all study participants from all communities, including subjects who only completed the behavioral questionnaire at 18.5 years of age. For the analysis of a specific endpoint, only subjects with measured endpoints were considered.The statistical analysis of VE of the AS04‐HPV‐16/18 vaccine against oropharyngeal infection was done by comparing the prevalence in females vaccinated with the AS04‐HPV‐16/18 vaccine from pooled Arms A and B over the prevalence in all females from Arm C for birth cohorts 1994–1995, for the following HPV types: HPV‐16, HPV‐18, HPV‐16/18, HPV‐6/11, HPV‐31/45 and HPV‐31/33/45.The VE was computed as 1 minus the odd ratio of prevalence rates between the investigated arms and the control arm, estimated using the Mantel–Haenszel test adjusted for clustering and stratified by the historical seroprevalence used in the randomization . The 95% confidence interval (CI) on VE was computed using the general inverse variance approach.As a post hoc analysis, the VE of the AS04‐HPV‐16/18 vaccine against oropharyngeal infection for birth cohorts 1994–1995 was computed for the following HPV types: HPV‐31, HPV‐33, HPV‐35, HPV‐45 and HPV‐52. For this post hoc analysis, only point estimated of the VE was provided due to the low number of events. It was computed as 1 minus the prevalence , and indistinguishable between the two types . The analyses related to the present objective are descriptive only.We hereby disclose results of our trial's secondary objective that was to evaluate VE of the AS04‐HPV‐16/18 vaccine against oropharyngeal infections in females of the 1994 and 1995 birth cohorts at the age of 18.5 years. Other objectives will be described in a separate publication and are available on the Clinical Trial Register .Click here for additional data file.


The northeastern region of Argentina has the highest age-adjusted cervical cancer mortality rates. Given the strong link between HPV infections and cervical cancer, one of the main interventions is the population-based use of HPV vaccines. However, the acceptability is not very clear in low- and middle-income countries. The purpose of this study was to estimate the level of HPV vaccine acceptance and associated determinants among caregivers of girls in a northeastern city of Argentina.A school-based survey was conducted in 2015 using a multistage sampling method. The primary sample unit were schools stratified by socioeconomic status selected at random, and caregivers of school girls were interviewed. The acceptability was determined using the adapted Theory of Planned Behavior. We performed logistic regression models to assess associated determinants.The study included 347 caregivers. The intention to vaccinate was 59.88%. A positive attitude of caregivers  and positive influence of social norms  were the main predictors independently associated to the intention to vaccinate against HPV. In contrast, practicing a Christian non-Catholic religion decreased the intention to vaccinate against HPV . All other factors evaluated were not significantly associated with intention to vaccinate against HPV.This study shows that evaluating attitudes, normative social beliefs, and perceived self-efficacy regarding HPV vaccination can be of utmost importance for mapping and planning of health-related strategies in developing countries. Cervical cancer (CC) is the fourth most common cancer in women worldwide . The burCervarix (which covers HPV types 16 and 18) and Gardasil , were approved in 2006 and 2007, respectively by the National Ministry of Health in Argentina [Given the strong link between HPV infections and cervical cancer, several new interventions have been introduced to reduce the burden of disease. The main one is the population-based use of HPV vaccines. In Argentina, two HPV vaccines, rgentina , 5. The Although the World Health Organization has urged countries to achieve at least an 80% of vaccine coverage in every district for all vaccines included in the national program of immunization .People´s intention to vaccinate against HPV is a factor that contributes to the acceptance of the vaccine. Thus, many studies have been carried out to research the determinants of HPV vaccine acceptance –13. In tThere is a lack of knowledge regarding how caregivers´ beliefs and sociodemographic factors could affect the individual behavior of vaccinating girls against HPV in Argentina. In accordance with the Theory of Planned Behavior , 20, whiThe area of study was the city of Resistencia, capital city of the Chaco province, in Northwestern Argentina. Resistencia is the most populated city in the province and is the main economic and cultural center; it is also one of the most productive cities in the Northwestern region of Argentina . It is lParents or caregivers of 11-year-old girls attending 5th or 6th school grade were eligible to participate and recruited from June to December 2015 in the city of Resistencia, Chaco. A multistage sampling method was used considering an expected prevalence of intention to vaccine of 46.6% (based on a prior pilot study) for a universe of 3,190 11-year-old girls . ConsideSchools within geographical segments of 300-houses (radius) from the 2010 Argentinean National Census were classified into five strata according to the proportion of unsatisfied basic needs (UBN) of each radius . The UBNThe selected schools were contacted by an investigator and invited to participate in the study. An informed consent form was sent to the parents or caregivers of eligible girls. Those who accepted it were sent a self-administered questionnaire by the school teachers. All teachers involved in the study were trained to supervise the completeness of the questionnaire by experienced researchers.The survey was based on the theoretical model of Adapted Theory of Planned Behavior (TPB) , 20. TheThis well-established model used in other preventive strategies provided the basis for questionnaire development regarding healthy behaviors. For this survey, we examined caregivers´ intention to have 11-year-old girls vaccinated against HPV and various factors that predicted vaccine uptake.Questionnaire development adhered to the steps needed to construct a TPB questionnaire and was based on a pilot study carried out to explore the intention to vaccinate, and to test and validate the questionnaire . The tarI have the intention to have my daughter vaccinated”. Parental intention was measured using a 5-point Likert scale and dichotomized using the mean score as a cut-off value. This method has been reported in other studies [The outcome being evaluated was caregivers´ intention to vaccinate their 11-year-old daughters against HPV at the moment of filing the questionnaire. Intention was measured by parents or caregivers self-report on their daughters´ HPV vaccination under the statement: “ studies ,22.Caregivers were asked to agree or disagree with several statements about HPV virus, related diseases and vaccination, using a 5-point Likert scale where 1 meant to strongly disagree; 3 meant neutral; and 5 meant to strongly agree. Afterwards, these statements were classified into TPB´s Attitude, Subjective Norm and Perceived control domains, which were the main predictors under assessment.Attitudes domain refers to the positive or negative judge a person makes when performing or not a behavior. In this study, it explored HPV virus awareness and knowledge as a necessary factor for cervical cancer development, as well as of the HPV vaccine effectiveness for cervical cancer prevention. The attitude domain scale of 3 items used a cut-off point of > = 4.31 (mean) to classify attitudes towards HPV vaccination as “positive”. Subjective norms domain refers to the beliefs about the perspective of significant people regarding weather or not a behavior is recommended. In this case, it explored recommendations to vaccinate against HPV by family members, close friends, teachers, physicians, nurses or television, and religion practices. Subjective norm scale consisted of 4 items, and a cut-off point of > = 3.73 (mean) was used to classify it as positive (meaning that Subjective norm supported HPV vaccination). Finally, Perceived control domain, also known as “self-efficacy” to vaccinate consisted of a 6 items scale, and a cut-off point of > = 3.65 (mean) was used to consider it positive.Because the wording of statements was sometimes positive and sometimes negative, we reversed the scoring where needed so that a higher score indicated always a strong agreement with HPV vaccination. Using the mean score as a cut-off value is an accepted method and has been used in other studies , 22.As explanatory variables, we considered socio-demographic characteristics including age (in years), gender , family role , educational attainment , main occupation , religion practice (yes/no), and type of religion (Catholic and other Christian-Non Catholic religions). Other covariates assessed were “knowledge about the HPV virus” (yes/ no), “family history of cervical cancer” (yes/ no), “advice to vaccinate by a teacher (yes/ no), pediatrician or nurse” (yes/ no), and “overall adherence to the national vaccination schedule” (yes/ no) .svyset command to account for sample stratification and weighting to provide unbiased estimates of the population parameters. Statistical significance was defined as a two-sided p-value <0.05 for all tests.We summarized continuous variables using means with standard deviations and categorical variables with percentages and 95% Wald Confidence Intervals (CI). To test the statistical significance of the difference between the intention to vaccinate of each categorical variable, the χ2 test was applied. T-test was used when testing continuous variables. Backward stepwise logistic regression analysis was used to study the effect of independent variables on the intention to the HPV vaccine. A p-value <0.2 was required for retention of each variable in the final model. Multicollinearity of independent variables was checked by the variance inflation factor (VIF) statistic. Results are presented as crude (ORc) or adjusted odds ratio (aOR) and 95% confidence intervals (CI). The Area Under the Curve (AUC) and Hosmer-Lemeshow goodness of fit tests were considered as discrimination and calibration parameters for the model. All statistical analyses were performed using the statistical software Stata IC, version 14.2 . Due to the complex sampling design, we used the Universidad Nacional del Nordeste, Argentina . Written informed consent was provided by all participants.The protocol was independently reviewed and approved by the Bioethics Committee of the Faculty of Medicine of the The flow chart for the participants’ inclusion is shown in Table 1. The response rate was 79%.The socio-demographic characteristics of the 347 parents/caregivers are summarized in The reported intention to vaccinate was 59.88% (95% CI: 57.57–62.14). The mean age of the caregivers surveyed was 39 years (SD 0.51). According to baseline socio-demographic characteristics, most of them were mothers (83.7%), had a tertiary/university (33.9%) level of education, and full-time employment (43.7%). The majority of the participants were Catholics. Recommendation to vaccinate by pediatrician and nurse were similar but higher than the recommendation by teacher.Table 2. All domains presented an acceptable Cronbach Alpha. The items with higher variability were found in subjective norm and perceived behavioral control domains.Mean scores within TPB domains and items used as predictors are shown in Table 3presents the results of the regression models with their crude (ORc) and adjusted (aOR) odds ratios. Attitudes towards the HPV vaccine in particular was the most important TPB domain related to the intention to vaccinate reported by the caregivers surveyed . It was followed by subjective norm  and perceived behavioral control . A positive attitude of caregivers  and positive influence of social norms  were the main predictors independently associated to the intention to vaccinate against HPV. In contrast, practicing a Christian non-Catholic religion decreased the intention to vaccinate against HPV . All of the other factors evaluated were not significantly associated with intention to vaccinate against HPV.This study sought to explore factors influencing caregivers’ intention to vaccinate their 11-year-old daughters in Resistencia, province of Chaco (Argentina) using the Theory of Planned Behavior. Our results showed an intention to vaccinate against HPV of 59%. Similar findings were also reported from Italy, with the intention to vaccinate with HPV was 61.8% . A positIn accordance with the results of similar studies, most of the caregivers surveyed were women of around 40 years of age (90%) , 32. ThiA qualitative systematic review published in 2015 assessed the use of various theories applied to the understanding of HPV vaccination as a behavior . Fifteenet al. studied parental intention to vaccinate young girls in Taiwan, and found significant association for all three domains, including PBC. However, the study was based on a convenience sample which could limit its generalizability to other contexts [et al. evaluated mothers’ intention to vaccinate their children in South Korea, and found controversial results regarding PBC associations with the intention to vaccinate against HPV (PBC was significantly associated in boys but not in girls) [More recent studies also support both attitudes and subject norms domains as the main predictors of HPV vaccination intention. Wang contexts . Kim et n girls) . Furtheret al. found that non-Baptists were 3.6 times more likely to accept the HPV vaccine for their daughters compared to Baptists [et al. found that Catholic parents were more likely than non-religious parents to have their daughters vaccinated [et al., our study showed that Catholic caregivers were more likely to accept the HPV vaccine. The low rate of acceptance among non-Catholic parents could be due to sexually-related concerns (22).Other researchers have studied different social and demographic factors, such as educational attainment, socioeconomic status and religion practices of caregivers. Our findings only support a negative association between practicing a Christian non-Catholic religion and the intention to vaccinate. Galbraith Baptists . Sheltonccinated . Similaret al. found that in British Columbia, Canada, the main reason for having girls vaccinated was the vaccine’s effectiveness, while the main reasons for not having girls receive the vaccine were concerns about its safety, preference to wait until girls were older, and not enough information to make informed decisions [et al. assessed how changes in parents’ attitudes were associated with improvements in HPV vaccine uptake in the United States [Contrary to our findings, other studies in high income countries have found that HPV knowledge and fear of adverse events could be important barriers to HPV vaccine uptake , 39–41. ecisions . VanWormd States . Despiteet al. showed that neither physicians’ nor nurses’ recommendations to get the HPV vaccine in the previous year were associated with the actual receipt of the next HPV vaccine dose [Despite the positive association between subjective norms and intention to vaccinate, our findings showed that neither the recommendation to vaccinate made by a physician, nor a nurse or teacher were relevant to caregivers’ intention to have their daughters vaccinated against HPV. This is supported by other studies which have found similar results. For example, VanWormer ine dose . It is wine dose –44, indiWe believe that these results could be relevant when putting the issues that are the object of our research into the Latin American context, particularly in South America. It is very important to understand the factors that may interfere with or enhance the success of HPV immunization programs worldwide, and especially in developing countries such as Argentina. Our findings suggest that government efforts towards improving HPV vaccination uptake among girls in Resistencia, Chaco should focus primarily in the implementation of strategies to shape a positive attitude towards HPV vaccination. Subjective norms also increased the intention to vaccinate, but only when family members, influential or close people were considered.Our study may have limited generalizability beyond the caregivers living in Resistencia, Chaco given that the individual constructs under evaluation could potentially vary by different cultures and social context. However, efforts were made to improve generalizability by means of random sampling schools. A cognitive bias could happen to understand negative statements under TPB although the instrument was pretested with a small sample of similar population. Another important limitation is the fact that our study did not include a qualitative assessment of the reasons why caregivers and adolescents didn’t have the intention to vaccinate, remaining a topic for further investigation in the Argentinean context.Evaluating attitudes, normative social beliefs and perceived self-efficacy regarding HPV vaccination can be of utmost importance for mapping and planning of health-related strategies in developing countries.S1 Table(XLS)Click here for additional data file.S2 Table(XLS)Click here for additional data file.


Although, there is a variable burden of human papillomavirus (HPV) in women infected with HIV in developing countries, there are few studies that attempted to surmise such variable evidences. This review aimed to estimate the pooled prevalence of HPV genotype distribution and risk factors contributing to HPV infection among women infected with HIV in low- and middle-income countries.We conducted a systematic review and meta-analysis of studies conducted in developing countries and reported HPV prevalence. We searched electronic databases: PubMed/Medline, SCOPUS, ScienceDirect, Excerpta Medical Database from Elsevier, Web of science, Cumulative Index of Nursing and allied Health Sciences and Google scholar databases to retrieve primary studies published in English language till 11th August 2019. We used random-effects model to estimate the pooled prevalence of HPV genotypes, and funnel plot to assess publication bias. The registration number of this review study protocol is CRD42019123549.2= 3782.80, p value < 0.001, I2 = 99.6%). The estimated pooled prevalence of all HPV genotypes was 63.0% (95% CI: 48.0–78.0) while the pooled prevalence of high risk and low risk HPV genotypes were 51.0% (95% CI: 38.0–63.0) and 28.0% (95% CI: 12.0–43.0), respectively. The pooled prevalence of HPV genotype 16 was 20%, while genotype 18 and 52 were 15% and 13%, respectively. Different risk factors reported for HPV infection and the frequently reported were low CD4 count below 200 cells/mm3 and high HIV viral load.We included nineteen studies with a total of 8,175 participants in this review. The prevalence of HPV was extremely heterogeneous across the studies (χThe pooled prevalence of HPV among HIV infected women in low- and middle-income countries was considerable and the proportion of high risk HPV genotypes were high when compared with low risk genotypes. Therefore, it is essential for the HPV prevention program to prevent the double burden of HPV and HIV in women. The papillomavirus is a heterogeneous group of DNA virus with circular, non-enveloped, double-stranded DNA genomes , 2. ThisThe prevalence of HPV is variable across the world. The study reported from developed countries indicate that the prevalence of HPV was 11 to 12% . The recThe highest prevalence was reported in sub-Saharan Africa (24%), Eastern Europe (21%) and Latin America (16%) .The burden of HPV infection is higher in HIV infected women 50.8%) than un-infected (22.6%)  and 78.8% than unSeveral risk factors are reported to be associated with HPV infection and these include HIV infection, other STIs , and multiple sexual partners , 21. TheHIV infection increases the risk of cervical infection due to high-risk HPV genotypes that induces high-grade cervical squamous intraepithelial lesions (HSILs), which in turn leads to the development of pre-invasive cervical lesions and invasive cervical cancer (ICC) –25. HIV We conducted a systematic review and meta-analysis of published articles to estimate the pooled prevalence of HPV in LMICs. We systematically searched the papers published in the following electronic databases; PubMed/MEDLINE, SCOPUS, Science Direct, Excerpta Medical Database from Elsevier (EMBASE), Web of science, Cumulative Index of Nursing and Allied Health Sciences (CINAHL) and Google scholar. The review was conducted in accordance with Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) standard . We usedPubMed database search strategy was:((((molecular[tiab] OR "Molecular Epidemiology"[Mesh]) AND ((Human papillomavirus[tiab] OR HPV[tiab]) OR "papillomaviridae"[MeSH Terms])) AND (((Human immunodeficiency virus[tiab] OR HIV[tiab]) OR "hiv"[MeSH Terms]) OR "hiv"[MeSH Terms])) AND (infected[tiab] OR positive[tiab])) AND  OR "women"[MeSH Terms]) OR "female"[MeSH Terms]) AND (("1966/01/01"[PDAT]: "2019/08/11"[PDAT]) AND "humans"[MeSH Terms] AND English[lang] AND "female"[MeSH Terms]).The search was repeated to identify the consistency of search terms and results. Two authors independently reviewed the titles, abstracts and full articles of the retrieved studies.We included a cross sectional and cohort studies conducted in LMICs based on World Bank Country Classifications, 2018  and thatWe assessed the quality of included studies by using the 14 items Quality Assessment Tool for Observational Cohort and Cross-Sectional studies NHLBI, NIH . This asData from eligible abstract and/or full text of the articles were extracted by considering the outcome variables  HPV genotypes and low-risk HPV types), and factors that could potentially be associated with these outcomes. The characteristics of study participants of an eligible paper such as age range, mean or median age, sex, HIV sero-status, the prevalence of HPV genotype were also extracted. Study characteristics such as first author, year of publication, study duration, study setting, study location or country, study design, sample size were also extracted Table . Other eThe majority of the studies included in our review had more than eighty percent and the lowest score observed was 62.5% in terms of quality. There was however one abstract included in the review, which was difficult to assess the quality of the article Table .2) based Q test with significant level of p value < 0.1 and I2. The I2 value of25% indicates low heterogeneity while 50% moderate and 75% high [We estimated the pooled prevalence of HPV with its 95% Confidence Interval (CI) using random effects meta-analysis model assuming the true effect size varies between studies . The pro75% high . The pot75% high . The potWe included 19 studies in our review ; p value < 0.001 and I2 = 99.6%). The pooled prevalence of high risk HPV was 51.0% ; p value < 0.001 and I2 = 98.88%. Similarly, the estimated pooled prevalence of low risk HPV was 28.0% ; p value < 0.001 and I2 = 98.32%.We pooled data from 8175 HIV infected women to estimate the pooled prevalence of HPV infection using meta-analysis. The overall pooled prevalence of all types of HPV infection was 63.0% Fig.  with higI2 = 98.53%) followed by 18 (I2 = 97.18%) and 31 (I2 = 96.17%) . The highest prevalence was observed for genotype 16 (20%) followed by 18 (15%) and 52 (13%). Almost all genotypes indicated heterogeneity and the highest heterogeneity was observed in genotype 16  with heterogeneity of I2 = 95.93% and p value < 0.001)  with heterogeneity of We assessed the effects of sample size and year of the study on heterogeneity between the studies using meta-regression model. Both sample size and publication years significantly predicted the heterogeneity of the effect sizes Table . In the The funnel plot  for the pooled prevalence of all genotypes HPV, high risk HPV and low risk HPV indicated that there is a publication bias Fig. a–c. FiguMolecular genotyping and HPV detection techniques applied for selected studies were Linear Array HPV Genotyping Test (LA), careHPV, genotyping using PCR-restriction fragment length polymorphism analysis, Reverse line-blot hybridization, INNO-LiPA HPV genotyping Extra® assay Table .3, history of three or more sexual partners were reported as the factors associated with HPV infection [High HIV viral load and low CD4 count were the most frequently reported factors that associated with high-risk HPV infection , 48, 50.nfection , 38  followed by 18 (15%) and 52 (13%), respectively. Low CD4count and high HIV viral load were the risk factors that most frequently reported in this review.This finding was lower than the findings in Kenyan which reported 68.0% overall pooled prevalence of high risk HPV among HIV positive women . GenotypLower CD4 count most frequently reported in this review is concordant with previous reviews in which low CD4 was strongly associated with HPV infection –55. PrevThe review conducted to estimate prevalence of HPV genotype among African women, including Ethiopia revealed that HPV16, 52, 35, 18, 58, 51, 45, 31, 53, and 56 were the ten most common genotypes in the normal cervical cytology while HPV 16, 18, 35, and 52 were the four common types . CorrespThe original research article conducted in Korea reported prevalence of 16.7% with the high risk HPV type of 12.5%  which isOur finding indicated heterogeneity on the outcome variable which is the effect size equivalent to the prevalence of HPV genotypes. Therefore, careful interpretation of the heterogeneity chi-square test (variation in effect estimates beyond chance) is required, since it has low power in the situation of a meta-analysis when studies have a small sample size or are few in number. It is worth noting at this junction that while a statistically significant result may indicate a problem with heterogeneity, a non-significant result must not be taken as evidence of no heterogeneity.This review is conducted by searching more than five biomedical databases and a large number of pooled participants are involved in the study. Another strength is that this review assessed HPV prevalence studies among HIV infected women in developing countries at large and reported pooled estimates of all HPV genotypes, high risk HPV genotype and low risk HPV. The limitation of this study was inclusion of studies published only in English language. This could be one of the possible causes for observed publication bias and heterogeneity of the estimated effects.This review indicated that the pooled prevalence of all genotypes HPV and high risk HPV among HIV infected women in LMICs were considerable. To enhance the well-being of HPV/HIV co-infected women it is necessary to strengthen programs for diagnosis, treatment, and provide HPV vaccination based on common high-risk genotypes.


The role of human papillomavirus (HPV) as single or multiple infections in pregnant women would be relevant to determine the time to progression and/or the time to regression of cervical lesions. In this preliminary study, we determined the prevalence of HPV as single or multiple infections in pregnant women from Northeastern Mexico.Samples from 31 pregnant and 62 nonpregnant women were examined between January 2015 and November 2015 at UMAE-23 of the Instituto Mexicano del Seguro Social (IMSS). The samples of cervicovaginal exudate were obtained for HPV DNA detection using the INNO-LiPA test, and HPV infections were analyzed as single or multiple infections. Participants completed a questionnaire on sociodemographic, gynecological, obstetric, and sexual behavior characteristics.The mean age of the pregnant women was 25.7 A higher prevalence of HPV as a single infection was found in this sample of pregnant Mexican women. Follow-up is necessary to evaluate the persistence or regression of the infection. In Mexico, cervical carcinoma is the second most common neoplasia and the second most frequent cause of death by cancer in women (1). Human papillomavirus (HPV) types 16, 18, 31, 45, and 58 are the most frequent in cervical samples from Mexican patients (2). Many factors can increase the risk of HPV infection; for example, sexual behavior, number of sexual partners, vaginal infection, and early onset of sexual activity. Diet, smoking, genetic factors, and immune suppression are also related to HPV infection (3-6). Studies have reported that pregnancy affects the immune system of the host and alters seroreactivity to HPV infection . Another important factor that should be considered in HPV-infected patients is the genotype distribution between single and multiple infections. Recently, Aguilar- Lemarroy and co-workers reported a higher prevalence of the HPV genotype as a single infection in Mexican patients with cervical cancer (2). Few studies have addressed the role of HPV as single or multiple infections in pregnant women. However, this information would be relevant to determining the time to progression and/or the time to regression of cervical lesions.Thus, this case-control study was conducted to investigate the presence of HPV as single or multiple infections in cervical samples from pregnant and nonpregnant women using the INNO-LiPA test.Thirty-one pregnant women between age 19 and 27 years in the third trimester of pregnancy and 62 non-pregnant women (as controls) within a similar age range (18-28 yr) were prospectively enrolled in this cross-sectional study from January 2015 to November 2015.All pregnant women who visited the Gynecology and Obstetrics Department of UMAE-23, Instituto Mexicano del Seguro Social (IMSS), Mexico, for the first time during pregnancy were invited to join the study. A gynecologist explained the aim of the study, the procedures, and the complications involved. Control samples were obtained from women attending early cancer detection programs at the same hospital. This was followed by an interview conducted by research assistants, in which all the participants responded to a questionnaire respecting reproductive history and sexual behavior.None of the women included in the study presented cervical lesions, as assessed by clinical examination of the cervix, cytology, and colposcopy. Women with immunosuppressive diseases, as hepatitis C, hepatitis B, HIV, and diabetes mellitus, were eliminated from the study.Cervical samples were obtained with a cytobrush in the course of gynecological exploration. The cytobrush was inserted into the endocervical canal, rotated 3 to 5 times, placed into the transport medium , and immediately sent to the Laboratory of Molecular Cytogenetics of Centro de Investigación Biomédica del Noreste, IMSS, Mexico. Cervical scraping samples were frozen at -20°C until DNA extraction. DNA was extracted using a Wizard Genomic DNA Purification Kit . The DNA quantity and quality were verified using a NanoDropⓇ Jenway kit .The INNO-LiPA HPV genotyping test  directs the amplification of a 65 bp region of the HPV L1 gene and allows the identification of 28 anogenital HPV genotypes: twelve high-risk (HR) HPVs, one probable HR HPV, seven possible HR HPVs, and eight low-risk HPVs (9). Two internal controls and two positive controls (both 65 bp products from the HPV L1 region) were included. Results from specimens where the controls failed to amplify were excluded from the study. The INNO-LiPA test permits the simultaneous detection of multiple genotypes in a single sample.Written informed consent was obtained from all participants and the study protocol was reviewed and approved by the Ethics and Scientific Committee of IMSS (Code: 293539).The risk of HPV infection associated with pregnancy [odds ratio (OR) and 95% confidence interval (CI)] was estimated by logistic regression, P No statistically significant differences were found between the two groups regarding age,  were HPV-positive. Of these, eight (73%) were infected with a single genotype and three (27%) were infected with multiple genotypes. HPV-16 was detected in six pregnant women . Among the 62 nonpregnant women, eight were HPV-positive (12%). Of these, three (36%) were infected with a single genotype and five (64%) were infected with multiple genotypes. The HPV-44 and HPV-52 genotypes were the most prevalent in all cases involving multiple infections (Table II).A multivariate analysis showed that pregnancy was associated with an increased risk for multiple HPV infection  and for single HPV infection .In this preliminary study, we determined the prevalence of HPV as single or multiple infections in pregnant women from Northeastern Mexico. Our findings revealed a significant increase in HPV prevalence among pregnant women (35%) compared with nonpregnant women (13%) (OR = 2.7).et al.  reported a higher prevalence of HR HPV infection in pregnant women compared with nonpregnant women, with detection rates of 37.2%  and 14.2% , respectively. Previous studies have reported similar findings (12-14).Studies comparing the distribution of HPV types in pregnant and nonpregnant women are scarce in the Mexican population. Hernández-Girón et al. (15) performed a meta-analysis of the risk of HPV infection in pregnant women from Asia, Europe, and North America and found a significantly increased risk among pregnant women , with a general OR of 1.42.Liu In contrast, Nobbenhuis (16) found no difference in HPV prevalence between pregnant and nonpregnant women. Such discrepancies may have arisen from the HPV detection method used and the number of detectable HPV genotypes. In our study, we used the INNO-LiPA test, which is able to identify 28 anogenital HPV genotypes and includes two internal controls and two positive controls. The INNO-LiPA test permits the simultaneous detection of multiple genotypes in a single sample.Previous study reported a dependence of HPV prevalence on age (11), parity, smoking history, oral contraceptive use, and history of sexually transmitted infections. In our study, the groups under study were homogeneous in terms of these factors (17-19).A possible explanation for the higher prevalence of HPV found in pregnant women respect to nonpregnant women is based that immunosuppression occurs during pregnancy because of the reduction of cellular immunity (7). It has been suggested that the high levels of estrogen and progesterone that occur during pregnancy decrease the cellular immunity, which is crucial for resolve the HPV infection. Our previous results showed that a high prevalence of HPV-52 and HPV-44 was involved in multiple infections in nonpregnant women (2). In contrast, a high prevalence of HPV-16 as a single infection was observed in pregnant women.Interestingly, our results revealed that 78% of the pregnant women who were HPV-positive presented with a single infection compared with 28% of the nonpregnant women.The transcriptional promoter of the E6/E7 oncogenic region of HPV-16 contains a steroid-hormone-receptor-binding element that activate HPV E6/E7 transcription, suggesting a hormonal stimulation effect on HPV replication. These observations indicate that pregnancy may have an effect on the patient's capacity to eradicate multiple HPV infections (20). Yamasaki (21) reported that accelerated clearance of HPV may be caused by the normalization of the host's immune system during the third trimester of pregnancy. Thus, the presence of HR HPV in the late period of pregnancy can be considered as being persistent and, therefore, represents a risk factor for neoplastic lesions.In conclusion, a higher prevalence of HPV as a single infection in pregnant women was found, despite the number of women included in the study was relatively small. Further investigations are necessary to establish the biological role of HPV as a single infection and its participation in the development of precancerous lesions or cervical carcinoma.The authors declare that there is no conflict of interest regarding this study.


Gardnerella vaginalis was associated with higher levels of inflammatory cytokines. Women with no history of HPV infection had similar cytokine profiles as those with HPV 16 post-clearance. This in vivo study documented an immune response shortly after HPV 16 clearance. G. vaginalis appeared to be involved in shaping this immune response. The appearance of G. vaginalis may have resulted from a shift from anti-microbial to anti-viral immune response with loss of bacterial control. The similar high levels of cytokines seen in women with no history of HPV suggest that a certain level of inflammatory surveillance is required to maintain an HPV negative state. This data may inform therapies such as probiotics or pro-inflammatory agents for treatment of persistent HPV.Persistent human papillomavirus (HPV) infections is necessary for the development of cervical cancers. Consequently, understanding the biologic mechanisms resulting in clearance is key in cancer prevention. Similar to other mucosal sites, it is expected that the local microbiome plays a significant role in shaping the immune response responsible for HPV clearance. Using cervical wash repository samples from a prospective study of HPV in women, this study investigates the microbiome and its associated inflammatory milieu during HPV 16 pre-acquisition, persistence and clearance states. For comparison, samples from women with no history of HPV ever during the study period were selected. We showed that 9 of 13 inflammatory cytokines were found to be significantly increased in the immediate post-clearance visit compared to the pre-acquisition or infection visits.  In vitro and in vivo studies suggest that clearance of HPV infections is directed by a host anti-viral immune response , the cause of cervical cancers, are well-known and include cell cycle deregulation, interference with telomerase activity and apoptosis Doorbar, . Yet, stLactobacilli species or L. iners were more likely to have hrHPV or dysplasia/cancer than VM dominated by L. crispatus   with 5 ml of normal saline were obtained at each visit. CW were immediately placed onto ice and frozen at −20° C within 4 h until transported on ice to the lab weekly. Two ml were removed for HPV DNA testing and the remaining 3 ml were divided into 3 cryovials for −80° C storage. HPV testing was performed based on PCR amplification using Roche Linear Array and an in-house Luminex platform  and then cleared with no subsequent positive tests for HPV 16 . The age range at first sampling ranged from 16 to 25 years, the number of visits with eligible samples ranged from 5 to 8 (see below) and the majority were white or Latina see . We seleSince women who acquire HPV 16 may be uniquely different, we identified 8 women who never acquired any HPV during their observation time in the study  . Visits In order to examine the integrity of the older samples, we first chose 10 samples from different women with a diagnoses of bacterial vaginosis (BV) by Nugent Score;  combined with bead beating. The hypervariable regions V1-V3 of the 16S ribosomal RNA (rRNA) gene were amplified using universal primers 27F and 534R according to the protocol developed by the Human Microbiome Project , microbiome dominated by L. iners (DLI), and those not dominated by any Lactobacillus species (NDL)  was measured by Shannon index, and was calculated using QIIME  post-clearance and each of the 3 pre-acquisition and 3 infection visits and 2nd post-clearance visit from women who acquired HPV 16. For women with no history of HPV infection, all the samples as a control group were compared to the samples in the immediate (1st) post-clearance from women who acquired HPV 16. The dynamics of the CW microbiome were analyzed based on the probability of transitions among the 3 states  using the Markov chain modeling , dominated by L. iners (DLI), and non-dominated Lactobacillus (NDL)] are shown in Gardnerella vaginalis, and fewer samples had DLI  (p = 0.02), indicating that the microbiome at post-clearance visits was at dysbiosis. Alpha diversity measured by Shannon index, which is an indicator of the evenness, and richness of the microbial community, found no statistical differences between the two groups with different HPV 16 status  . The sam6 status , howeverWe investigated the microbiome temporal stability by analyzing transition probabilities between microbiome community states in the subjects using Markov chain modeling. We found differences between WNHPV and those with HPV16. WNHPV had relatively stable microbiota states . If the L. iners, the relative abundance increased at the immediate pre-infection and then decreased thereafter. G. vaginalis increased in relative abundance at immediate pre-clearance and significantly increased at post-clearance, which then returned to a baseline months later. Enterococcus had an inverse relationship with G. vaginalis. On the other hand, in WNHPV, the relative abundance of L. iners was higher than in samples from the immediate post-clearance visits of HPV infected subjects. G. vaginalis in WNHPV was lower than immediate post-clearance but similar to pre- and post-infection levels.We next examined the CW microbiome changes at the species level . SeveralTo investigate the changes in host immune response relative to the HPV status and microbiome changes, we analyzed the expression profiles of 13 inflammatory cytokines listed in p ≤ 0.005; p < 0.01).At the immediate post-clearance visit, overall cytokine levels were highly elevated compared to the pre-acquisition and infection visits. The mean number of cytokines ranked in the top quartile based on the expression level was 8.6, compared to the 1.9 to 2.9 in the other visits including the 2nd post-clearance visit months later . When compared to the immediate post-clearance visit, WNHPV had similar levels of IFN-γ, IFN-α-2, and TNF-α, lower levels of IL-4 (p < 0.001), IL-5 (p < 0.001), IL-10 (p = 0.02), and IL-12 (p = 0.02) and higher levels of IL-13 (p = 0.02) and MIP-1α (p < 0.001). The median values are shown in When compared to the samples from the WNHPV, the expression levels of ILs-4, -5, -10, -12, and -13, IFN-γ, IFN-α2, MIP-1α and TNF-α were significantly elevated compared to samples from all the pre-acquisition and infection visits , IL-5 (p = 0.004), IL-10 (p = 0.02), IL-12 (p = 0.004), IL-13 (p = 007), TNF-α (p = 0.006), IFN-γ (p = 0.005), and MIP-1α (p = 0.02)  . Of noteine peak .As HPV 16 persistence is associated with the development of several anogenital cancers, it is important to understand mechanisms involved in the clearance of HPV. To our knowledge, ours is the first longitudinal study to examine associations among the cervical-vaginal microbiome, and local immune milieu at HPV 16 acquisition, persistence and clearance. Several findings were worthy of discussion. First, the striking increase in inflammatory cytokine expression was not witnessed until the immediate post-clearance visit. We had hypothesized that we would observe a time period when the response was mounting prior to clearance. Our findings imply that the several-fold increase in the majority of inflammatory markers observed after a documented clearance event was residual evidence of a successful anti-viral response. Since we only obtained samples every 4–6 months, this finding was not surprising in retrospect but emphasizes that the successful immune response is likely to occur suddenly and swiftly and these responses persist for a relatively short period as the expression of the inflammatory markers returned to the baseline months later. Of interest, WNHPV had inflammatory expression levels similar to the post-clearance visit in women with HPV infection. This may suggest that the subjects who acquired HPV 16 were more vulnerable to HPV because of an inadequate immune milieu or response. Another possible explanation is that WNHPV had a previous infection that cleared and now have a higher immune surveillance protecting them from HPV reinfection.To carry this notion of immune surveillance further, the Markov-chain model of the CW microbiome dynamics analyzed in our study supports the theory that women who acquire HPV 16 have a different local environment than those who appear protected from acquisition. Women who acquired HPV 16 had relatively unstable CW microbiome states in that they had high probabilities of switching from one state to another. In contrast, WNHPV had stable states throughout the visits, predominantly being in DLC and DLI, both of which are considered healthier states than NLD microbiota. The stability observed in the microbiome of this group and yet a higher inflammatory state similar to that observed in the post-clearance state suggest that there is a protective microbiome homeostasis against pathogens.G. vaginalis is often more abundant in NLD, the observed increase in G. vaginalis during persistence and post-clearance was not surprising. NLD microbiome of which G. vaginalis is one of the predominant species, is often considered a pathogenic microenvironment since it has been linked to several morbidities such as pelvic inflammatory disease, premature labor, and HIV acquisition  2, 3 were exited for treatment.in vivo study is the first to document that the clearance of HPV 16 is associated with a distinct inflammatory eruption witnessed immediately after clearance. The inflammatory burst was associated with microbial dysbiosis and an increase in NLD. It is plausible that HPV 16 clearance was associated with anti-viral immune response resulting in a microbiome shift that promoted the dominance of these bacteria in the community. The high stability of the microbiome state observed in WNHPV was associated with a higher level of cytokines as seen in post-clearance state, suggesting that the local microbiome plays a significant role in immune surveillance to maintain an HPV negative state. The use of probiotics to achieve vaginal microbiome stability may be a promising approach to promoting HPV clearance in women with persistent infections.In conclusion, this https://www.ncbi.nlm.nih.gov/, PRJNA588856. Cytokine dataset is available on request.The microbiome datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found at: The primary study was approved by the Committee on Human Research, University of California, San Francisco and patients consented to participate and to place their samples in repository. This study used samples from the repository and was approved by the Institutional Review Board of the University of California, Los Angeles.A-BM was the initial principal investigator of the prospective cohort study and designed this sub-study and oversaw the statistical analysis and prepared the manuscript. HL and BS collaborated in the design of the sub-study and contributed to the manuscript preparation. HL oversaw all the microbiome analysis and all the statistical analyses of the microbiome, cytokine, and metabolome data. BS performed all the microbiome, cytokine, and metabolome data analysis and statistical analyses. HH and EB were essential to the development of the assays for processing the biorepository samples. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Human papillomaviruses (HPVs) have been divided into mucosal and cutaneous types according to their primary epithelial tissue tropism. However, recent studies showed the presence of several cutaneous types in mucosal lesions and healthy mucosa from different anatomical sites.Gamma-PV PCR and CUT PCR.Here, the HPV prevalence and type-specific distribution were assessed in a variety of mucosal samples from 435 individuals using a combination of two established broad-spectrum primer systems: Alpha-PVs were most frequently found (59%), followed by Gamma- (37%) and Beta-PVs (4%). The prevalence and persistence of HPV infection in the anal canal of 226 individuals were further explored. Overall HPV, Gamma-PVs and multiple HPV infections were significantly higher in men vs. women ; multiple HPV infections were more common in individuals ≤40 years (p = 0.05), and significantly higher prevalence of Gamma-PVs and multiple HPV infections was observed in HIV-1-positive vs. HIV-1-negative individuals . Out of 21 patients with follow-up anal swabs, only one persistent infection with the same type (HPV58) was detected.Overall HPV prevalence in anal canal swabs, cervical cancer biopsies, genital warts and oral swabs was 85, 47, 62 and 4%, respectively. In anal canal swabs, Gamma-PVs (except species Gamma-6) are ubiquitous viruses with dual muco-cutaneous tissue tropism. Anal canal Gamma-PV infections may be associated with sexual behavior and the host immune status. This study expands the knowledge on Gamma-PVs’ tissue tropism, providing valuable data on the characteristics of HPV infection in the anal canal.Our findings suggest that  Papillomaviridae family  , and 94 oral swabs collected during routine paternity testing, were obtained from the archival collection of samples of the Instituto de Biología Molecular y Celular de Rosario  and the Institute of Microbiology and Immunology, Faculty of Medicine, University of Ljubljana . Before subsequent analyses, all samples were collected, processed, and stored at − 80 °C, as described previously  or Beta-PVs [HPV38 (Beta-2)] ] Fig. .Fig. 2DAlpha-, Beta- and Gamma-PVs were present in 59, 3 and 35% of the initial samples obtained from all study participants at the first visit, respectively. In addition, multiple infections were detected in 43% of included samples.The overall prevalence of HPV infection in the anal canal of the 226 patients included in the study was estimated at 79%  (Table p = 0.05). A significantly higher prevalence of Gamma-PVs and multiple HPV infections was observed in HIV-1-positive vs. HIV-1-negative individuals . All differences between gender, age and HIV-1 status observed during univariate analyses were additionally confirmed using multivariate analyses . Interestingly, none of them were identified in a variety of 653 cutaneous samples with the same testing approach [Thirty-six different approach . These rapproach , 40–43, approach .Beta- and Gamma-PVs) were relatively common in the mucosal epithelia, suggesting a possible dual tissue tropism of the majority of HPV species [Gamma-PV types were identified in a surface swab of a genital wart and a cervical cancer biopsy sample, possible as part of the normal mucosal microbiota, as reported previously [Our observations are not surprising since previous studies have demonstrated that typically cutaneous HPV types . Nevertheless that the prevalence of Alpha-PV infections was similar in anal canal of MSM and women (p = 0.515), interestingly, the prevalence of Gamma-PVs was significantly higher among MSM (p = 0.027). Although the mentioned differences may have originated from the use of different HPV detection methods in different studies, it is likely that receptive anal sexual intercourse might indeed have resulted from more frequent acquisition of Gamma-PVs, as suggested previously for Alpha- and Beta-PVs [Beta- and Gamma-PVs, may occur through self- or partner- inoculation [In line with previous epidemiological studies that have analyzed the prevalence of HPV infection in anal canal of MSM , 27, 28,Beta-PVs , 49. On culation , 51.p = 0.05). The age-specific HPV infection prevalence trends in the anogenital region may have resulted from younger subjects being more sexually active than older individuals, with up to 3-fold higher number of sexual partners [In line with previous observations that younger age could be associated with the higher risk for acquiring anal HPV infection in men , 52, in Beta- and Gamma-PV prevalence data among HIV-1-positive individuals are conflicting [Gamma-PVs and multiple HPV infections was significantly higher in HIV-1-positive subjects in comparison to HIV-1-negative individuals . As the contradictory results obtained in epidemiological studies could be attributed to differences in the immune status of HIV-1-infected individuals and to the heterogeneity of enrolled patients and HPV detection methods used, additional studies are warranted to further explore the association(s) between HIV-1 and anal HPV infection.Since flicting , 28, 41,Alpha-PV infection in the anal canal [Beta- and Gamma-PV infections were found to be transient, a persistent Alpha-PV infection was detected in one patient. Although transient Gamma-PV infections have previously been described in healthy skin samples [Alpha- and Beta-PVs can establish persistent infections of the anal canal [Gamma-PVs, followed by direct sequencing of PCR products, which may represent a limitation, as it could have led to the underestimation of some HPVs causing persistent infections and being present in lower viral loads. Therefore, it is possible that Gamma-PVs also cause transient infections of the anal canal, mostly having a commensal role and being transmitted through sexual and non-sexual routes.Due to the scarce knowledge on the natural history of non-al canal , 21 pati samples , in studal canal , 27. It Gamma-PVs are ubiquitous viruses with a wide tissue tropism, as they were detected in both mucosal and cutaneous sites, with the exception of members of the Gamma-6 species, which most probably only colonize mucosal ecological niches [Gamma-PV infections of the anal canal may be associated with sexual behavior and the immune status of infected individuals. In conclusion, the present study expands the knowledge on Gamma-PVs’ tissue tropism, providing valuable data on the characteristics of HPV infection in the anal canal.Based on results of our study and findings published previously, it could be concluded that l niches , 30, 53.Additional file 1: Table S1. HPV types/putative types detected by Gamma-PV and CUT PCR assays. Novel putative HPV types are depicted in bold. Un: species not included in the current HPV taxonomy. *: The same HPV/putative type was found in different sets of samples. Gamma-X refers to sequences grouped in the genus Gamma-PV from which the prevailing HPV type/s and species could not be determined.Additional file 2: Table S2. Distribution of HPV types in 21 initial and 23 follow-up anal canal swab samples from 21 individuals. Linear Array HPV Genotyping Test  is based on the reverse-line blot hybridization technique to detect the following 37 different mucosal HPV types distributed in 11 Alpha-PV species (high-risk HPV types depicted in bold): Alpha-1 (HPV42), Alpha-3 , Alpha-5 , Alpha-6 , Alpha-7 , Alpha-8 (HPV40), Alpha-9 , Alpha-10 , Alpha-11 , Alpha-13 (HPV54), Alpha-14 (HPV71). RHA Kit Skin (Beta) HPV assay  detects the following 25 different Beta-PV types by reverse-line blot hybridization technique: Beta-1 , Beta-2 , Beta-3 , Beta-4 (HPV92), Beta-5 (HPV96). HPV types (or negative results) identified simultaneously by more than one HPV test are depicted in bold. Samples containing possible unknown HPV type(s) are indicated as HPV-X. Pos: Positive; Neg: Negative; N/A: Not Analyzed.Additional file 3: Table S3. Alignments of the forward and reverse primer sequences of CUT PCR, Gamma-PV PCR and CP PCR with corresponding regions of L1 and E1 ORFs of 64 selected Gamma-PV types. Lines and characters represent identical and mismatched nucleotides, respectively. Conserved sequences at the 3′ regions of Gamma-PV primers are highlighted in grey. Degenerate nucleotides of primers: i = inosine, y = c/t, r = a/g, w = a/t, h = a/c/t, m = a/c, s = g/c.


Deceptive path-planning is the task of finding a path so as to minimize the probability of an observer (or a defender) identifying the observed agent’s final goal before the goal has been reached. It is one of the important approaches to solving real-world challenges, such as public security, strategic transportation, and logistics. Existing methods either cannot make full use of the entire environments’ information, or lack enough flexibility for balancing the path’s deceptivity and available moving resource. In this work, building on recent developments in probabilistic goal recognition, we formalized a single real goal magnitude-based deceptive path-planning problem followed by a mixed-integer programming based deceptive path maximization and generation method. The model helps to establish a computable foundation for any further imposition of different deception concepts or strategies, and broadens its applicability in many scenarios. Experimental results showed the effectiveness of our methods in deceptive path-planning compared to the existing one. Deceptive planning in an adversarial environment enables humans or AI agents to cover their real intentions or mislead an opponent’s situation awareness. This would be of great help to many real-world applications, such as deceptive network intrusion , roboticmagnitude metric to measure the path deceptivity. To compute its value, based on the classical model-based goal recognition approaches [G given observation O; further, This paper mainly uses the proaches , our mettated in , which htated in , that goWith the deception being defined upon each separate node or step, a deceptive path could be easily generated when we maximize deception along the path from the start to the end. Inspired by this intuition, the paper proposes a single real goal magnitude-based deceptive path-planning model, as well as a deceptive path generation model formed into a mixed-integer programming problem. The advantages of the model are generalized as follows: firstly, using magnitude to quantify deception at each node helps to establish a computable foundation for any further imposition of different deception strategies, and thus broadens the model’s applicability in other scenarios with different deception characteristics; secondly, differently from the previous method  where deThe paper is organized as follows. This work lies at the intersection of two well-established and much-studied disciplines within computer Ssience: path-planning, which is the problem of finding a path through a graph or grid from a given start to a given goal ,10,11; aGrounded on the proposition that the order of goals ranked by likelihood can surely be inverted to describe their unlikelihood, the paper uses the results of the probabilistic goal recognition to plan a deceptive path using optimization method. Thus in this section, we first introduce the basic concepts, background, and applications of deception, especially the deception in path-planning. After that, we talk about the related works in goal or plan recognition, and review the recent developments of problem formulations, models, and algorithms applied in probabilistic goal recognition.The deception problem is significant; it is common, and occurred frequently in human history . As for Defined by  and we qdenial and deception.As a more focused area, research on the deceptive path appears in the literature under various guises. Jian et al.  tried todenial strategy. Using the same strategy but in a more practical case, Root et al. [Hespanha et al. ,18 and Rt et al.  studied Deception strategy (presenting misleading information) arose in a path-planning-related experiment carried out by roboticists Shim and Arkin [nd Arkin , inspireRecent innovative work on goal recognition design (GRD) ,36,37,38bounded nonoptimal setting, where an agent was assumed to have a specified budget for diverting from an optimal path. Additionally, as they presented, it suits for situations where deceptive agents aim at achieving time-sensitive goals, with some flexibility in their schedule. This exactly is the condition for the deception strategy (presenting misleading information) to be applied. Though holding a different perspective, the GRD problem provides valuable insights to the study of deceptive path-planning.Interestingly, following the continued research on the GRD problem where the original assumptions are gradually relaxed, another form of deception appears in the literature as well. When considering suboptimal paths, the authors of  focused The most recent work by Masters et al.  presentsπ4 as in ) under tA closely related topic to deceptive path-planning is the deceptive or adversarial task planning. Braynov  presentsThe research on deception arises in many other studies which we mention here only briefly: in cyber security , privacyThe goal recognition problem has been formulated and addressed in many ways, as a graph covering problem upon a plan graph , a parsiAmong the approaches viewing the goal or plan recognition as an uncertainty problem, two formulations appear and solve the problem from different perspectives. One focuses on constructing a suitable library of plans or policies ,52, whilWithin that formulation, research continues to improve the model’s expressiveness and computational attractiveness, as in ,64,65,66Recently, the work ,56 showsSeveral works followed that approach ,72,73,74According to the relationships between the observer and the observed agent, the goal or plan recognition could be further divided into keyhole, intended, and adversarial types ,75,76,77We first begin by formalizing path planning and goal recognition. We propose a rather simple but general model suitable for different deceptive path planning domains. We assume that there are two players: one is the evader (the observed agent); the other one is the defender (the observer). The evader chooses one goal from a set of possible goals and attempts to reach it, whereas the defender must correctly recognize which goal has been chosen. We assume that the outcomes of the action are deterministic and the model is fully observable to the evader and the defender, meaning that the environment and each others’ actions are visible to both players. In addition, the evader has several assumptions about the defender. Firstly, the defender does not know the decision-making process of the evader. Secondly, the defender operates under a fixed recognition algorithm.The notion of a road network captures the intuition about all possible deceptive paths that can be planned against a set of goals.Definition 1.A road network is a triplet N is a non-empty set of nodes (or locations);i-th node in Define the path Definition 2.The single-goal path-planning problem is a triplet s and ends in one single goal g. Denote The solution to the single-goal path-planning problem is a path which starts in its source Definition 3.The probabilistic goal recognition problem is a tuple G given observation sequence O, with each two neighboring observations A solution to the probabilistic goal recognition is a conditional probability distribution costdif which was applied early in [costdif has an inverse correlation to goals’ posterior probabilities; that isIn this paper, we take a concept early in  to refleTheorem 1.costdifG could be computed using:The fined in  represenAs we have talked about in the last section, Masters and Sardina  proved tTheorem 2.Both Theorem 2 and Equation , including a single real goal s and moves towards G as well as the defender’s observation model P to plan a deceptive path. Corresponding to the single-goal path-planning problem, this work considers a single real goal situation and leaves the multiple-real-goals setting to the future research. We assume that the evader has different deception strategies on planning the path.Specifically, in this paper we could replace Definition 5.The evader’s deceptive strategy is a function Different strategies evaluate a different deceptive magnitude at each node. Later, we introduce strategies in detail. Further, we present a single, real goal, magnitude-based deceptive path planning model as follows:Definition 6.The single real goal, magnitude-based deceptive path-planning is a triplet R is the total amount of distance allowed for the deceiver traversing Apart from the elements in SRGDPP problem, the SRGMDPP adds a deceptive strategy function that maps the probabilistic goal recognition results into quantified path deception at each individual node. Also, a explicit presentation of total resource constraint enables the evader to flexibly adjust its deceptive path plan according to the time (or fuel) constraints in practice. We say that SRGMDPP provides a computable foundation for further impositions of different deception concepts or strategies, and broadens the model’s applicability in other scenarios.s–Inspired by the intuition that a deceptive path could be generated when maximizing deception along the i. Assign the same magnitude value i to values k in the set k; k or not, k is traversed, or else, s and reach i in the set Denote ′)·ck·xk∑k∈FS(i)xabout in ; one is Definition 7.The simulation strategy The simulation strategy measures the amount by which a false goal dominates the real goal. And the greatest deception occurs when there are the greatest dominance. Definition 8.The dissimulation strategy As a popular method, this type of deception (or we say the degree of ambiguity) is defined using the Shannon’s entropy, which is usually used to describe information uncertainty. By taking advantage of the defender’s observation model, the about in , though However, both concepts of deception reflect only a fraction of what it means to be deceptive. The difference lies in the extent to which people want to deceive the observer. According to the strength of intention protection desire, usually these two strategies could be balanced in human behaviors. Thus the paper proposes a more flexible weighted combination strategy.Definition 9.The combination strategy We quantify that aspect of deception as a linear combination of the simulation and dissimulation. If we deceive successfully using the combination strategy, the defender would either be led to the false goal  that direct to node i. This in turn means that node j cannot be reached from i and j must be in earlier positions than i of the whole path. Then, consider two cases: (1) when i itself locates in the first place before any other nodes; (2) for i is reached, a trajectory starting from s must have been drawn out already. Then, j is either on this trajectory, or it is not. When j is on, it cannot contribute one more edge into i. When it is not, two trajectories, j, coexist, and that contradicts the task of generating only one path from Assume to the contrary that all the ending nodes of edges in Equation , there mThe above theorem could be easily extend to more general situations.Theorem 4.For the node i, add these edges into the ordered edge set, and continuously do these two steps until no elements in i. Relying on Theorems 3 and 4, we present the cyclic-path-identify algorithm (whose pseudocode is given in Algorithm 1).Algorithm 1:Cyclic-path-identify algorithmThe above observations assure us that when confronted with nodes X, given the original road network s. We first retrieve the solution road network X, and initialize an ordered edge set j of i from the ending nodes of edges j and ending at Algorithm 1 shows how to identify the cyclic path from the optimization result G given O, as shown in combination strategy to In this section, we compare the performance of the model and strategies proposed by this paper with those (strategies o d4) by . First wo d4) by  generateLAB 2012 .Specifically, in ategy in , first ts to the last deceptive point (LDP), and then on to Path fined in , in whicIt might be more deceptive to a human observer until later in the path than s to the LDP, thereby requiring more time compared to the other three. However, upon this simple case study, there does not exist too much difference between Strategies S and C fully deceive the defender at each D strategy generates a cyclic path at the beginning (marked as red dashed line in D only try to make the defender not know the real goal, though it may still guess correctly. Compared to strongly-deceptive strategies S, “showing the false” is not their primary focus, and thus in most situations they are more efficient in terms of cost.In S, D, and C shown in As for the results of strategies The case study shows the effectiveness of our method in planning deceptive paths. Also, paths generated using SRGMDM do not rely on the so-called LDP as a transit point before moving towards the real goal, and the trajectories are more flexible, as shown in tated in ; our metIn order to have a more thorough comparison between different methods and strategies, in the following experiments, we conduct an empirical evaluation over a problem set which is generated upon the grid-based maps from the moving-AI benchmarks . The soutAs with the above cast study, in each case we generated one optimal path using the standard A* and seven deceptive paths, each corresponding to a different deceptive strategy. Using probabilistic goal recognition and under the assumption that priors for all goals are equal, we calculated probabilities at intervals to confirm/assess deceptive magnitude. As to evaluating traces with different lengths, the paper normalizes traces with different lengths into F-measure, which is frequently used to measure the overall accuracy of the recognizer [F-measure will be between 0 and 1, and a higher value means a better confidence of recognizing the right goal while a less deceptive strategy. Thus, the lower value, the more deceptive.We got the statistical results using the metric of cognizer  and is ccombination strategy were set the same as in the above case study. The resource constraints of paths following simulation, dissimulation, and combination were the same as those of paths using F-measure of A* increases greater than The results are shown in eir work , strategethod in  is not. simulation, dissimulation and combination. Firstly, we could find that in all situations, the simulation strategy always has a similar (when Now, we compare the performance of dissimulation and combination strategies, paths become more deceptive as the number of possible goals dissimulation is better than F-measure down from dissimulation fails Interestingly, for Combination is strictly better than simulation and dissimulation, two concepts of deception are both embedded in one strategy, and thus the merits of the two enable the combination strategy to outperform simulation when dissimulation when combination or a similar balanced strategy using different weight settings, along with our magnitude-based deception maximization model could be of great help in planning suitable paths.The experiments prove the effectiveness of our method in making full use of environment information, compared to the work in  that narIn this paper, we formalise deception as it applies to path-planning and present strategies for the generation of deceptive paths. The paper propose a single, real goal, magnitude-based deception maximization model which establishes a computable foundation for any further imposition of different deception strategies, and broadens the model’s applicability in many scenarios. Different from the previous method  where deThough this work is quite suggestive, there are still many open problems that could be researched in the future. Firstly, the paper assumes that the defender is naive and rational. It is naive because the defender does not expect to be deceived, and thus deploys no counter plans or tactics. Its rationality is inherited from the goal recognition paradigm. If it is not, on one hand it may use machine learning approaches, e.g., the work in , to adapSecondly, the current model assumes that the environment and deceiver’s actions are fully observable to the goal recognizer or the observer. However, for partially observable situations where its actions could be masked and behavioral patterns be covered, we would expect many more tactics could be available for the evader. Also as current formulation only considers one real goal setting, this work could be further extended to multiple real goal settings and thus transforming the problem into non-linear path-planning, which would be a much more valuable scenario where the evader have more than one targets.Lastly, we believe that the concepts of deception, along with its models, could also be applied in other domains, such as classical planning using a plan library, etc. Similarly, a probabilistic plan recognition algorithm could then be used to evaluate the magnitude of deception, and a newly proposed optimization model with resource constraint to maximize the plan deceptivity.


Introduction: Coenzyme Q10 (CoQ10) has a potential role in reducing the risk of myocardial infarction by slowing the progression of atherosclerosis and improving ischemia. In this study, we will assess the role of coenzyme Q10 in prophylaxis for reducing myocardial infarction and mortality related to myocardial infarction.https://www.randomizer.org/). Group A received 100 mg coenzyme Q10 daily (coenzyme Q10 group) in addition to standard therapy and group B received standard therapy only (placebo group).Methods: This open-label two open placebo-controlled randomized clinical trial was conducted in a tertiary care hospital in Sukkur, Pakistan from April 2016 to September 2019. Eight hundred nighty-two (892) patients with clinically diagnosed and documented evidence of hypertension were enrolled in the study from the outpatient department. Participants were randomized into two groups by 1:1 ratio using an online randomizer software, Research Randomizer  with relative risk reduction of 2.92 . The number needed to treat to prevent one non-fatal myocardial infarction was 34. Participants who received coenzyme Q10 had fewer incidence of fatal myocardial infarction over 12 months (1.5% vs 3.1%) with relative risk reduction of 1.65 . Number needed to treat to prevent one fatal myocardial infarction was 60.Conclusion: According to this study, coenzyme Q10 reduced the incidence of fatal and non-fatal myocardial infarctions. Clinicians should consider adding coenzyme Q10 to the treatment regimen of high-risk patients of myocardial infarction. We suggest coenzyme Q10 may be an effective prophylactic agent in patients at risk of myocardial infarction and it may help in reducing burden on the health care system. Coenzyme Q10 (CoQ10), which is synthesized in the inner mitochondrial membrane, is an essential compound of the human body . It alsoCoQ10 may have a plausible role in prevention and treatment of heart ailment, as it improves cardiac cellular bioenergetics ,4. CoQ10https://www.randomizer.org). Group A received 100 mg coenzyme Q10 daily twice a day (coenzyme Q10 group) in addition to standard therapy and group B received standard therapy only (placebo group).This open-label two open placebo-controlled randomized clinical trial was conducted in a tertiary care hospital in Sukkur Pakistan from April 2016 to September 2019. Eight hundred nighty-two (892) patients with clinically diagnosed and documented evidence of hypertension were enrolled in the study from the outpatient department. Ethical review board approval from Ghulam Muhammad Mahar Medical College was taken before enrollment of patients . Patients were enrolled using a consecutive convenient non-probability sampling technique. To nullify the effect of different treatment used in management of hypertension, patients only on angiotensin converting enzyme inhibitor (ACEi) in combination with hydrochlorothiazide for hypertension, rosuvastatin for hypercholesterolemia and aspirin for inhibiting platelet aggregation were enrolled. Participants were randomized into two groups by 1:1 ratio using an online randomizer, Research Randomizer  was diagnosed based on clinical symptoms, cardiac markers such as troponin, and electrocardiogram changes. Thirty-eight (38) and twenty-nine (29) participants in co-enzyme Q10 and placebo, respectively, were lost to follow up and were not included in the final analysis.https://www.medcalc.org) using 95% confidence interval. Survival probability will be plotted on Kaplan Meier graph. The log rank test was used to calculate the chi-square (X2) for each event time for each group. A p-value of less than 0.05 meant that there is a difference between the two groups and the null hypothesis is void.Statistical analysis was done using Statistical Package for Social Sciences (SPSS) version 23 . Continuous variables were analyzed via descriptive statistics and were presented as mean and standard deviation (SD) while categorical variables were presented as percentages and frequencies. Relative risk reduction (RRR) and number needed to treat were calculated via an online calculator  participants completed the study, 408 in the coenzyme Q10 group and 417 in the placebo group. There was no difference in demographics and risk factor profile between the two groups  with relative risk reduction of 2.92 . The number need to treat to prevent one non-fatal myocardial infarction was 34. Participants who received coenzyme Q10 had fewer incidence of fatal myocardial infarction over 12 months (1.5% vs 3.1%) with relative risk reduction of 1.65 . The number need to treat to prevent one fatal myocardial infarction was 60.As per Kaplan-Meier curve for non-fatal myocardial infarction, the log rank rest for the data in our study was p value = 0.09; thus the two curves were not statistically significantly different Figure .As per Kaplan-Meier curve for fatal myocardial infarction, the log rank rest for the data in our study was p value = 0.11; thus the two curves were not statistically significantly different Figure .According to this study, coenzyme Q10 reduces the risk of fatal and non-fatal myocardial infarction. Eleawa et al. also postulated that CoQ10 may be an effective prophylactic to prevent myocardial infarction (MI) and MI-induced cardiac changes. They found that coenzyme Q10 protects against the reinfarction in myocardial infarction. It also reduces infarct area, inflammation, and oxidative stress while normalizing left ventricular hemodynamics after MI . CoQ10 (Coenzyme Q10 also plays a role in various other cardiovascular disease and events. Coenzyme Q10 reduces admission in heart failure and reduces the episodes of pulmonary edema in heart failure . MunkholTo the best of our knowledge, this is the first study in a local setting that studies the role of prophylaxis against myocardial infarction. The study has its limitation as well. First, since it was single-center study, sample size diversity was reduced. Second, participants were not followed post-MI, hence the role of coenzyme Q10 in preventing MI induced could not be studied in detail.In this study, coenzyme Q10 reduced the incidence of fatal and non-fatal myocardial infarctions. Clinicians should consider adding coenzyme Q10 to the treatment regimen of high-risk patients of myocardial infarction. We suggest coenzyme Q10 may be an effective prophylactic agent in patients at risk of myocardial infarction and it may help in reducing the burden on health care systems. Using coenzyme Q10 as a supplement will improve the quality of life of such patients.