A classification accuracy of 91.3% was achieved in an independent blinded test set (n = 69), demonstrating the potential clinical utility of Raman spectroscopy.There is an unmet clinical need for new methods to aid clinicians in the early detection of cervical cancer and precancer. Spectroscopic methods such as Raman spectroscopy can provide a rapid, label-free and nondestructive measurement of the biochemical fingerprint of cells, tissues and biofluids. This study aims to demonstrate the clinical utility of Raman spectroscopy for the identification of cervical precancer. Raman spectra were recorded from cervical smear samples (n = 662) 1 and 136 CIN2+) were obtained as a training set. Raman spectra were recorded from single-cell nuclei and subjected to a partial least squares discriminant analysis (PLSDA). In addition, the PLSDA classification model was validated using a blinded independent test set (n = 69). A classification accuracy of 91.3% was achieved with only six of the blinded samples misclassified. This study showed the potential clinical utility of Raman spectroscopy with a good classification of negative, CIN1 and CIN2+ achieved in an independent test set.The mortality associated with cervical cancer can be reduced if detected at the precancer stage, but current methods are limited in terms of subjectivity, cost and time. Optical spectroscopic methods such as Raman spectroscopy can provide a rapid, label-free and nondestructive measurement of the biochemical fingerprint of a cell, tissue or biofluid. Previous studies have shown the potential of Raman spectroscopy for cervical cancer diagnosis, but most were pilot studies with small sample sizes. The aim of this study is to show the clinical utility of Raman spectroscopy for identifying cervical precancer in a large sample set with validation in an independent test set. Liquid-based cervical cytology samples ( Cervical cancer is the fourth most commonly diagnosed cancer, with 570,000 cases, and the fourth leading cause of cancer death, with 311,000 deaths, among women worldwide . CurrentRecently, human papillomavirus (HPV) testing has replaced cytology for primary screening in many countries due to a higher sensitivity than cytology for the detection of high-grade cervical precancer 2+) . HoweverRecently, optical spectroscopic techniques, such as Raman spectroscopy, have shown great promise for cancer diagnosis using tissues, cells and biofluids ,10. RamaOur group developed methods to record Raman spectra from clinical ThinPrep cytology samples on glass slides and to address blood contamination that can obscure spectral features and lead to variability ,15,27. Wn = 662) and by validating this model by means of an independent blinded dataset (n = 69).To date, however, the studies used small sample sizes and the results were validated using leave-one-out cross validation rather than using an independent test set. The aim of the present study is to show the clinical utility of Raman spectroscopy for identifying cervical precancer by developing a classification model based on spectral data from negative, CIN1 and CIN2+ ThinPrep cytology samples (n = 390). Ethical approval for use of pseudo-anonymised samples for this study was granted by the CWIUH Research Ethics Committee (study no. 28–2014).Cervical smear samples collected in PreservCyt solution were obtained with consent from patients attending the Colposcopy clinic at the Coombe Women and Infants University Hospital (CWIUH), Dublin, Ireland (n = 392) was provided through application to the Scottish HPV Archive, a research tissue biobank set up to facilitate HPV-associated research. The archive comes under the auspice of the National Research for Scotland tissue bioresource, 20/ES/0061. Our previous study demonstrated that samples stored at −25 °C could be analysed successfully by Raman spectroscopy and that fresh samples, biobanked (stored) samples and pooled fresh and biobanked samples achieved a similar sensitivity and specificity for detection of CIN 2+ [A further set of cervical smear samples  had insufficient cells remaining in the ThinPrep vial after cytology and HPV tests were performed or had excessive debris covering any remaining cells. These samples could not be used for Raman spectroscopic analysis and had to be excluded. The training set consisted of samples which were either confirmed as normal by histology with no abnormality detected (n = 115) or were negative on cytology and/or colposcopy but had no biopsy or histology (n = 211), samples which were confirmed as CIN1 by histology (n = 200) and samples which were confirmed as CIN2 or CIN3 (CIN2/3) by histology (n = 136)  . Samplesn = 69) was provided by the Scottish HPV Archive as above. These samples were used as an independent test set to validate the classification model. This set of samples was blinded to the researchers until after the spectral acquisition, data preprocessing and analysis steps were carried out. The samples were confirmed by histology as normal (n = 17), CIN1 (n = 32) and CIN2+ (n = 20)  .Cervical smear samples were tested for HPV using a variety of technologies. Cervical smear samples collected by the CERVIVA team at the Coombe Women and Infants University Hospital were tested for HPV DNA using the Cobas HPV DNA test (Roche) and for HPV mRNA using the Aptima HPV Assay (Hologic), as part of a series of CERVIVA research programmes underway at that time.Samples collected as part of the Scottish HPV archive were tested for HPV using a variety of tests, including: Hybrid Capture 2 (HC2) (Qiagen), Cobas HPV Assay (Roche), Cepheid Expert HPV assay, the Abbott Real-Time HPV Assay, the Aptima HPV Assay (Hologic) and Optiplex HPV Genotyping test .The Cobas HPV test (Roche) is a fully automated PCR-based HPV DNA test that detects in three separate channels: HPV16 individually, HPV18 individually and a pool of 12 other HPV genotypes (11 definite high-risk genotypes plus 1 possibly high-risk genotype) as follows: HPV 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68.The Aptima HPV Assay (Hologic) detects mRNA coding for the E6 and E7 viral proteins of HPV types 16, 18, 31, 33, 35, 38, 39, 45, 51, 52, 56, 58, 59, 66 and 68. The Aptima HPV Assay involves three main steps, which take place in a single tube: target capture, target amplification by transcription-mediated amplification (TMA) and detection of the amplification products (amplicon) by the hybridization protection assay (HPA). A 1ml aliquot of PreservCyt specimen was used.®2 technology which is a nucleic acid hybridization assay with signal amplification that utilizes microplate chemiluminescent detection. Specimens containing the target DNA hybridize with a specific HPV RNA probe cocktail. The HC2 High-Risk HPV DNA Test detects HPV DNA from 13 high-risk HPV types, 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59 and 68.The HC2 High-Risk HPV DNA test (Qiagen) uses Hybrid CaptureThe XPert HPV Assay (Cepheid) is a real-time polymerase chain reaction (PCR) assay for the detection of 14 high-risk types of HPV DNA through five separate channels . The assay is formulated in a single-use cartridge, provides a result within one hour, can be performed by non-laboratory-trained health-care workers and requires minimal hands-on time.The Abbott Real-Time HPV Assay (Abbott) detects 14 high-risk HPV genotypes with simultaneous identification of HPV 16 and HPV 18.The Optiplex HPV Genotyping test  is a multiplex in vitro test kit based on Luminex Technology for the qualitative determination of human papillomavirus (HPV) genotypes 6, 11, 16, 18, 26, 31, 33, 35, 39, 42, 43, 44, 45, 51, 52, 53, 56, 58, 59, 66, 68, 70, 73 and 82 in polymerase chain reaction (PCR)-amplified samples of genomic DNA isolated from cervical smears.The Roche Linear Array HPV Genotyping Test is a qualitative test that detects 37 high- and low-risk human papillomavirus genotypes.Samples were prepared for Raman spectroscopy using the ThinPrep 2000 processor . The ThinPrep processor homogenized the sample by spinning the filter, creating shear forces that break up any clumped material . The cells were then transferred onto the TransCyt filter and transferred onto a glass slide to produce a monolayer of cells approx. 20 mm in diameter. The slide was then ejected into a fixative bath of 95% ethanol.2O2 at room temperature for 3 min, followed by a 70% solution of industrial methylated spirits (IMS) for 3 min followed by multiple dips into 100% IMS to remove any remaining cellular debris and H2O2 and were air dried.The slides then underwent a pretreatment step to remove any molecular contamination by haemoglobin, which obscured several features of the cellular spectrum as described in our published protocol . Briefly−1, with an integration time of 30 s averaged over two accumulations. Spectra were recorded only from the cell nucleus as these were found to be more reproducible and consistent than spectra from the cell cytoplasm [Raman spectra were acquired as described in our published protocol  using a ytoplasm . Where pn = 662) and a blinded test set (n = 69) was employed to validate the model.Data were normalised and analysed using MATLAB software (MathWorks) and specific scripts developed and adapted for uploading of the spectra and their preprocessing, including smoothing , baseline correction (rubberband) and vector normalization. The spectra were corrected for the glass background using a linear least-squares method with non-negative constraints (NNLS) as described in our published protocol . The datn = 114), borderline (BNA)  or low-grade (LSIL)  on cytology, with low numbers of samples with atypical (ASCUS/ASC-H/ASC-BNA) , high-grade (HSIL)  or no cytology results . Almost half of the CIN1 samples were LSIL  and almost a third were BNA  on cytology. The remainder of the CIN1 samples were HSIL , negative , ASCUS/ASC-H/ASC-BNA  or had no cytology results . The CIN2+ samples were either LSIL , HSIL  or BNA  on cytology, with low numbers of samples with negative , ASCUS/ASC-H/ASC-BNA , high-grade/invasive  or no cytology results .Cytology results were available for over 95% of the training set samples and are shown in n = 301) and are shown in n = 125), and of these, 45.6% had an HPV DNA-negative result (n = 57), 24% had an HPV DNA-positive result (n = 30) according to an hc2 or Linear Array test and 30.4% had an HPV genotype result from a Cobas, Optiplex, Luminex, Abbott or Cepheid test (n = 38). The majority were either HPV16 and other high-risk HPV types (n = 10) or other high-risk HPV types (n = 21) (n = 93), and of these, 8.6% had a negative result (n = 8), 32.25% had a positive result (n = 30) according to an hc2 or Linear Array test and 59.14% had an HPV genotype test result , (n = 55). The majority were HPV16 and other high-risk HPV types (n = 11) or other high-risk HPV types (n = 28) (n = 83), and of these, no samples had a negative result, 21.68% had a positive result (n = 18) according to an hc2 or Linear Array test and 78.31% had an HPV genotype result from a Cobas, Optiplex, Luminex, Abbott or Cepheid test (n = 65). Again, the majority were HPV16 and other high-risk HPV types (n = 23) or other high-risk HPV types (n = 22)  were only available for less than half of the training set samples  . Less th(n = 28) . More th(n = 22) .n = 236) and are shown in n = 107), and of these, 55.14% had a negative result (n = 59) and 44.86% had a positive result (n = 48). Less than half of the CIN1 samples had an HPV mRNA test result , and of these, 27.96% had a negative result (n = 26) and 73.12% had a positive result (n = 68). Only 25.74% of the CIN2+ samples had an HPV mRNA test result (n = 35), and of these, 20% had a negative result (n = 7) and 80% had a positive result (n = 28).HPV mRNA test results (Aptima) were available for 35.65% of the training set samples (−1 (glycogen), 621 and 644 cm−1 (proteins), 728 and 784 cm−1 (DNA), 828 cm−1 (DNA/RNA), 853 and 936 cm−1 (glycogen and proteins), 957 cm−1 (DNA), 1004 cm−1 , 1035 cm−1 (proteins), 1092 cm−1 (DNA phosphate backbone), 1127 cm−1 (proteins), 1176 cm−1 (cytosine/guanine), 1210 cm−1 , 1245 cm−1 (amide III), 1320 cm−1 , 1338 cm−1 , 1422 cm−1 (DNA/RNA), 1450 cm−1 (proteins and lipids), 1578 cm−1 (nucleic acids), 1610 cm−1 , 1656 and 1669 cm−1 (amide I) (−1 (proteins) and negative peaks at 482, 853, 936 (glycogen), 1238 (proteins), 1400, 1422 and 1578 cm−1 (nucleic acids).The mean Raman spectra recorded from the negative, CIN1 and CIN2+ samples are shown in amide I) . Differe−1.A PLSDA classification model was developed from the spectral data of the training set. The PLSDA latent variables (LVs) showed similar features to those observed in the difference spectra at 482, 728, 784, 828, 852, 1092, 1238, 1334, 1380, 1450, 1485, 1578, 1656 and 1669 cmn = 17), CIN1 (n = 32) and CIN2+ (n = 20)  , Table 3n = 12) or borderline  on cytology. Similarly, the CIN1 samples were mainly low-grade  or borderline , with low numbers of high-grade  or no cytology result . The CIN2+ samples were a mixture of negative , borderline , low-grade  and high-grade  cytology.Cytology results were available for over 98% of the validation set samples and are shown in n = 68 samples and additional Abbott and BD tests for n = 8 samples) were available for over 98% of the validation set samples and are shown in n = 12), 17.65% HPV16 and other high-risk HPV types (n = 3), 5.88% low-risk HPV types (n = 1) and 5.88% no HPV types (n = 1). Similarly, the CIN1 samples were 68.75% other high-risk HPV types (n = 22), 18.75% HPV16 and other high-risk HPV types (n = 6), 3.13% low-risk HPV types (n = 1) and 3.13% no HPV types (n = 1) (n = 9), HPV16 and other high-risk HPV types , HPV16 , HPV16, 18 and other high-risk HPV types  and no HPV types .HPV DNA test results (Luminex test for  (n = 1) . The HPVn = 39) and are shown in n = 5). Of the CIN1 samples tested, 33.33% had a negative result (n = 5) and 66.66% had a positive result (n = 10). Of the CIN2+ samples tested, 31.57% had a negative result (n = 6) and 68.42% had a positive result (n = 13).HPV mRNA test results (Aptima) were available for 56.52% of the validation set samples (n = 69) are shown in the confusion matrix . The different spectra and latent variables (LV1 and LV2) showed that the discrimination was mostly based around increased nucleic acids , decreased glycogen  and changes in protein features , indicating an increased proliferation and altered protein expression as a result of HPV infection. These discriminating features were consistent with our previous studies [Initially, Raman spectra were recorded from a training set consisting of cervical ThinPrep samples . Raman spectra were recorded from this test set and used to test the classification model. In total, 63/69 samples (91.30%) were classified correctly. All 17 negative samples were classified correctly, but 2 CIN1 samples and 4 CIN2+ samples were incorrectly classified as negative. Five of the misclassified samples were low-grade or high-grade on cytology and were high-risk HPV DNA-positive and HPV mRNA-positive. Interestingly, one of the misclassified samples, which was CIN3 on histology, was negative on cytology, had no HPV types and was HPV mRNA-negative. Although misclassified according to histology results, interestingly, the negative classification was consistent with the cytology and HPV test results. Our study showed similar performance to a recently reported SERS approach using gold nanoparticle substrates to enhance the Raman signal of cervical exfoliated cells [A PLSDA classification model was developed from the training set for discrimination between negative, CIN1 and CIN2+ ThinPrep samples, and validated using an independent blinded dataset . For the samples with normal histology, these were comprised of samples with negative, borderline and low-grade cytology for the training set, but only borderline and low-grade cytology for the test set. As not all samples were tested for HPV DNA and mRNA, it was difficult to compare the training and test sets in terms of HPV test results. Another limitation of the study was that not all of the negative samples in the training set were confirmed by histology as normal, but this was difficult to achieve, as a biopsy was not performed for all patients undergoing a colposcopy. In addition, the sample set represented a disease-enriched population to increase the number of disease cases. This is standard practice when evaluating a new technology, particularly in the case of cervical cancer, where CIN2 prevalence is only around 1% in routinely screened high-income countries. Future work should involve a prospective study in a screening population. Although a technical validation exercise was performed demonstrating that stored samples are credible biospecimens for Raman analysis [In our study, for the samples with CIN1 histology, the test set corresponded reasonably well to the training set, as both sets of samples were mainly comprised of samples with borderline or low-grade cytology. For the samples with CIN2+ histology, the training set mostly consisted of samples with borderline, low-grade and high-grade cytology, while the test set also contained samples with negative cytology (analysis , the useRecently, HPV testing has been introduced for primary cervical screening, but the HPV DNA test cannot distinguish between a transient and a transforming HPV infection, and cytology-based triage is also needed. Our study aimed to evaluate Raman spectroscopy as a tool to support current cervical screening approaches, HPV testing and cytology. As HPV primary screening has been implemented widely, there is a need to improve the specificity through the use of a good triage test. Raman spectroscopy may be a good alternative to cytology triage, as it is less subjective with a better sensitivity. Our recent study has shown that Raman spectroscopy has potential as a triage test for HPV-positive women to identified transforming HPV infections , althougIn addition, in the future, with more widespread HPV vaccination, the performance of techniques relying on a subjective assessment, such as cytology, should decrease as a consequence of the reduction in the prevalence of the disease. Thus, new objective tools, such as Raman spectroscopy, would be needed for screening future vaccinated populations. Raman spectroscopy is a low-cost solution that has the potential to be integrated into existing services, as well being a potential solution for developing countries, where “see-and-treat” options are preferred to increase the effectiveness of screening hard-to-reach populations.In this study, a classification model for negative, CIN1 and CIN2+ ThinPrep samples was developed and validated. The model was developed from a training set of 662 samples and an accuracy of 91.3% was achieved in an independent test set of 69 samples.Raman spectroscopy is a label-free objective tool that may be a good alternative to cytology for the triage of HPV-positive cases. It may also be important in the future as HPV vaccination increases, and the prevalence of the disease decreases, because the reduced performance of current screening and triage tests should result in a clinical need for new objective tools for screening vaccinated populations.


One of the most frequent cancers that affects males globally is cervical cancer (CC) that kills hundreds or even thousands of women each year, particularly in underdeveloped nations. The study focuses on human papillomavirus (HPV) that contributes to cervical cancer (CC) development. In the majority of Arab nations, there seems to be no public education or vaccination programs. In research, methodological rigor is employed to find solutions to both theoretical and practical difficulties. This research aims to assess the knowledge and awareness of the HPV vaccination among Emirati men. Results of the research showed that Emirati males had a poor understanding of HPV and its vaccination. According to the findings of this research, Emirati males lack a basic understanding of HPV, which necessitates the implementation of national HPV education initiatives. We have identified several critical knowledge gaps that can be filled in the future regarding HPV infection and vaccination. The human papillomavirus (HPV) is widespread in the United States, and the majority of sexually active persons will get the virus over their lifetime . ChronicThe U.S. Food and Drug Administration has authorized and licensed three HPV vaccinations for use as preventative measures. According to current recommendations, the most undervaccinated group is males; nevertheless, a two-dose regimen is currently suggested; yet, vaccination coverage across all eligible categories is still low, with males constituting the most undervaccinated segment of the population. 39.7 percent of females and 21.6 percent of males aged 13 to 17 got all doses inside the series in 2014. As a reference, vaccination rates for females have been shown to range from 60.5% to 91.5% in a number of nations [Research in public health indicates that adoption and uptake of the vaccination are dependent on public awareness and understanding about HPV. A recent study indicated that only 68% of participants from a representative sample of UAE individuals were aware of HPV and the HPV vaccination , 7, 8.Contrary to popular belief, the bulk of current HPV insight and perception research focuses only on cervical cancer in both men and women. Most people have no idea of the link among HPV and noncervical malignancies, according to studies looking into people's understanding of noncervical HPV-related health consequences.There may be substantial consequences for vaccination uptake if more research looks into knowledge and awareness about noncervical HPV-related malignancies, both on its own and in connection to other forms of HPV awareness and knowledge . There a11-12-year-olds are now advised to get the HPV vaccine as a preventative measure, with an option for catch-up vaccinations until the age of 26. This was the recommended upper age limit for HPV immunization until recently. For persons aged 27–45, the FDA authorized HPV immunization in 2018 because of the vaccine's safety and effectiveness. According to an advisory committee in June 2019, the HPV vaccination may be given to persons aged 27 to 45 years. This advice is based not just on safety and effectiveness, but rather on vaccination effectiveness and cost-efficiency [In the United States, a large percentage of adults are still unvaccinated for HPV, putting them at risk of developing malignancies linked to HPV because of the country's persistently low HPV vaccination rates among the country's most important cohort, teenagers, and among catch-up young adults. 15.8 percent of females and 3.2 percent of males between the ages of 27 and 45 have gotten at least one dose of the HPV vaccination, according to the National Health Interview Survey 2017 . AccordiThe Health Information Nationwide Trends Survey (HINTS), sponsored by the National Cancer Institute (NCI), collects data on how the public uses cancer-related information on a national level. An adult sample in the US has been surveyed using this data source in the past to determine their knowledge and understanding of HPV. Researchers have utilized HINTS to examine discrepancies in HPV understanding and information across racial and gender groups, as well as between men and women and throughout the country. According to previous research based on this dataset, males and non-Hispanic black or Hispanic individuals were equally likely to know about HPV and the HPV vaccination, whereas women were much more likely . A 54-yeMiddle Eastern civilizations and religious conservatism may be seen across the region from Morocco to Saudi Arabia. As a result, this suggests more restrictive sexual practices than those seen in other civilizations . The freCervical cancer rates in this area are thought to be lower than those in the rest of the globe, despite the lack of cancer registries. One reason for the change in introducing countrywide cervical cancer prevention initiatives in this region is that despite erroneous incidence numbers, breast cancer was diagnosed as the most frequent disease in women in Algeria and Morocco and the third most prevalent cancer in Tunisia, Qatar, and the United Arab Emirates .There is still a high incidence of cervical cancer in the Arab nations, despite the low prevalence. The launch of a safe and efficient anti-HPV vaccination in the Arab MENA region and the rest of the globe is a fantastic chance to eliminate cervical cancer as well as many other HPV-related illnesses in the region and throughout the world. HPV vaccination was authorized in June 2006 by that of the Food and Drug Administration for primary prevention of the disease. Cervarix, a bivalent HPV vaccination, and Gardasil, a quadrivalent HPV vaccine, are now on the market . Anti-HPThe study contribution finds out whether or not a representative sample of Emirati males was aware of and knowledgeable about HPV. HPV vaccination, awareness and understanding of HPV-associated cervical cancer, knowledge and awareness of noncervical or “other” HPV-associated malignancies, and general HPV awareness and knowledge were all examined in the research. The influence of sociodemographic factors on research and understanding was also examined , 23.This study aims at identifying the level of awareness and knowledge of Emirati men about HPV, where they got that information, and what exactly they know about HPV.e) studies aspects that cannot be quantified and hence cannot be susceptible to estimate and quantitative interpretation. In contrast, quantitative research (f) necessitates extensive use of tools and methodologies [Consider research as a broad phrase that incorporates a diverse array of activities. Methodological rigor is used in research to find answers to both theoretical and practical problems. There are a range of approaches available depending on the goal and context of the study, including the following: descriptive analysis, which focuses on amassing data that establishes the existence of a phenomenon. For instance, applied analysis in product design, design engineering, and policy formulation aims to answer practical questions but also aid decision-making; in theoretical research, on the other hand, research is conducted to pique scientific curiosity rather than to immediately apply findings in real-world applications. For the study topic, qualitative research  for not applicable, while sexually transmitted disease as indicated in As shown in As shown in As shown in As shown in As shown in As shown in As shown in As shown in As shown in As shown in As shown in As shown in There is a wide range of variation in the general public's understanding of HPV. Gender, age, education, marital status, wealth, and race were all shown to be linked with previous authors' estimates of general HPV awareness when looking at sociodemographic variables . It is iAll participants had a good understanding of HPV's relationship with cervical cancer, but they had little idea of the link between HPV and other malignancies. Noncervical malignancies' relationship with HPV has not been the subject of comprehensive inquiry or reporting in the research literature to this point. The results reported by the few research that have been published are consistent , 30.Previous studies have consistently indicated that women had higher levels of vaccination understanding and information than men. The results on vaccine understanding and information and gender are at conflict with this. It was shown that being a woman was a significant and favorable predictor of actual vaccine knowledge, but that was only true for males who claimed to have heard of the immunization. Analytical approaches and knowledge categories may be to blame for these differences, according to our hypothesis. Men may have heard about HPV vaccination, but when other sociodemographic criteria are taken into account, it has little influence on real knowledge of HPV vaccine .Considering the importance of vaccine effectiveness, it was surprising to discover that the majority of the respondents did not think the vaccination prevented cervical cancer. Despite no significant differences in the percentage of men and women reporting talking to someone like a doctor about vaccination, women were far more likely than men to have received a doctor's advice to vaccinate. Vaccination recommendations are more likely to be given to female patients and parents of female children, according to previous studies. A combination of sexism and the fact that the vaccination has only been approved for boys since 2011 may explain the present results , 33.Health education researchers and practitioners may benefit from this study's results, which are summarized below. First and foremost, there was a lack of understanding and awareness of many elements of HPV, despite continued attempts to do so. Many intervention studies have shown the beneficial impact of receiving information on people's willingness and ability to become vaccinated. Informational interventions alone may not be adequate for long-term increases in vaccination uptake rates, according to a new comprehensive evaluation of the evidence. Our findings indicate that more HPV education efforts are required. Priority should be given to evaluating the efficacy of educational interventions on their own and in conjunction with other approaches , 35.Another problem was the lack of public awareness and knowledge about the relationship between HPV and other types of HPV-related malignancies. This discovery is especially significant in light of studies indicating that vaccination decision-making might be influenced by personal significance. Individuals and parents are more likely to accept and plan to vaccinate when they have a greater estimation of their own HPV risk and a higher estimation of the vaccine's advantages. When HPV-related health effects particular to males, such as disproportionate incidence of oropharyngeal cancer and penile cancer, are highlighted, men are more likely to support vaccination uptake. Men and parents of boys may benefit from educational activities that stress the danger of noncervical HPV-associated cancers, according to these data .Noncervical HPV-associated cancer awareness and knowledge are also important for methodological reasons. There has been a lack of emphasis paid to HPV's relationship with noncervical HPV-associated malignancies in the current literature. Increased precision in measurement will add to the already existing corpus of information. Noncervical cancer awareness and knowledge may be assessed by including questions on awareness and knowledge , 38.Last but not least, a lack of vaccination awareness and a widespread misconception that the vaccine is ineffective in preventing cervical cancer point to the need of public education on the subject. Interactions between healthcare providers and their patients give an opportunity to dispel common misunderstandings regarding risk, effectiveness, and other important decision-making variables . PatientInteresting about our results is the significant desire to learn more about the vaccination and the readiness to be vaccinated or to have their daughters vaccinated, despite the dearth of understanding about HPV, its causative association with cervical cancer, and the availability of the vaccine. These findings point to a major potential for HPV and cervical cancer education and awareness campaigns. There seems to be a lack of political will for national cervical cancer prevention due to a lack of cultural barriers. On the basis of our results, a vaccination of this kind might be more widely accepted if the general public was made more aware of HPV's importance, danger, and possible problems , 41. A cSeveral features have an effect on people's general awareness about HPV, knowledge of HPV-related malignancy, and knowledge of the HPV vaccination. More has to be done to increase HPV vaccination uptake, especially among those populations identified in this research as having poor HPV awareness. Improving our understanding of HPV might be crucial to increasing vaccination rates in the future. That relationship between sociodemographic variables, HPV knowledge, and vaccination uptake has to be explored further via research to be clear. Given the significance of personal relevance in vaccination decision-making, studies that investigate knowledge of noncervical HPV-associated malignancies should be prioritized. Health educators will be better able to influence uptake if they continue to study and record the effectiveness of their intervention tactics. Patient–provider relationships are crucial for vaccination acceptability and should be further studied and practiced in public health.


Psoriasis is a chronic inflammatory disease with characteristic skin manifestations. Several pathogens can cause flare‐ups of psoriasis. The risks of skin infections are increased in patients receiving immunomodulators. A patient with chronic psoriasis presented with human papillomavirus infection and anogenital warts and was treated surgically with acceptable results. Here we present a case with chronic psoriasis developed anogenital condyloma acuminata. The warts originated from normal skin, not from psoriatic plaque. We consider these reactivation of latent HPV unrelated to recent psoriasis therapy.  Clinical presentations include characteristic skin lesions, arthritis, and other extracutaneous manifestations.2Treponema pallidum particle agglutination tests) and an anti‐human immunodeficiency virus screening test (enzyme‐linked immunosorbent assay) were non‐reactive. We performed tumor excision and electrocauterization for the perianal lesions. Histopathological findings were compatible with condyloma acuminata  and HPV infection. However, we considered the relationship between chronic psoriasis and HPV activation in the presence of anogenital warts. To the best of our knowledge, no study has discussed HPV infection associated with other common skin manifestations besides psoriasis.Apart from some rare skin diseases, a high prevalence of HPV infections is also related to the treatment of psoriasis with psoralen‐ultraviolet‐A but not with tumor necrosis factor alpha inhibitors.4This case demonstrates a possible activation of latent HPV, which led to anogenital condyloma acuminate, in a patient with psoriasis that was unrelated to recent psoriasis treatments. This report also highlights the high incidence of HPV‐related diseases, including possible malignancies, in individuals with psoriasis. The underlying factor responsible for HPV activation, in this case, remains unclear. The interrelationship between psoriasis, cytokines, inflammatory cells, and HPV type should be further investigated.The authors declare no conflict of interests.Wu‐Feng Hsieh involved in literature review, study results interpretation, and manuscript editing. Ta‐Wei Pu involved in study design and final editing. Jung‐Cheng Kang and Chao‐Yang Chen involved in study conception. Je‐Ming Hu contributed to manuscript draft. Kuan‐Hsun Lin reviewed the literature.The project was approved by the Medical Ethics Committee of the Tri‐Service General Hospital.Written informed consent was obtained from the patient for the publication of this case report and accompanying clinical images.


Respiratory sinus arrhythmia (RSA) is a phenomenon in which the heart rate (HR) changes with respiration, increasing during inspiration and decreasing during expiration. RSA biofeedback training has an effect in relieving negative mental conditions, such as anxiety and stress. Respiration is an important indicator affecting the parasympathetic activation within the body during RSA biofeedback training. Although there are existing studies that consider individual differences when selecting optimized respiration using heart rate variability, the studies that use the high frequency components of HRV, which is an indicator of parasympathetic activation, are insufficient. For this reason, this paper proposes a process to identify optimized respiration for efficient RSA feedback, consisting of three steps: (1) application, (2) optimization, and (3) validation. In the application phase, we measured PPG data against various respiratory cycles based on the HF components of HRV and calculated the proposed heart stabilization indicator (HSI) from the data. Then, we determined the optimized respiration cycle based on the HSI in the optimization step. Finally, we analyzed seven stress-related indices against the optimized respiration cycle. The experimental results show that HSI is associated with the parasympathetic nervous system activation, and the proposed method could help to determine the optimal respiratory cycle for each individual. Lastly, we expect that the proposed design could be used as an alternative to improving the efficiency of RSA biofeedback training. Occupations that require much psychological attention over a long time  can cause emotional and psychological distress. Activities ,2,3 suchBiofeedback training enables individuals to maintain and restore autonomic nervous system balance by regulating the activation of their physiological activities ,16. Bio-In particular, RSA biofeedback training, one of the types of biofeedback training, allows the body to become more stable by increasing parasympathetic activity. It leads people to breathe at the rate of the heart’s rhythm and baroreflex activity is associated with breathing ,27,28. TResearchers suggest that high HRV power promotes autonomic homeostasis and the ability to regulate emotionally , whereasMost studies have observed that a training rhythm of about six breaths per minute increases the power of the HRV ,37,38,39Therefore, this study proposes an overall process consisting of three processes—(1) application, (2) optimization, and (3) validation—to find an optimized breathing cycle to enhance the RSA biofeedback effect. The first process includes the breathing guidelines, including a visualization program to extract the proposed heart stabilization indicator (HSI). The second process explains selecting the optimal breathing cycle for each individual based on the indicator. The last process validates the proposed optimization method using the stress-related frequency domain parameters of HRV obtained from the experimental results.This study is organized as follows. We proposed a process to find the optimized respiration cycle for an individual based on the HF components of HRV, consisting of three steps: (1) application, (2) optimization, and (3) validation, as shown in RSPt) to eliminate the effect of the stimulation sequence on the experiment. We configured the same inhalation and exhalation time  and gave the resting time (RESTt) after a breathing cycle for 2 min (CYCLEt) as Actions 1 and 2. We also developed a visual screen as the blue part under the LabVIEW environment for apparent execution of these actions and showed it to the participants. The experiment lasted for 10 min  in a space free from environmental noise.In order to secure the reliability of the collected PPG data, this paper defines specific respiration guidelines, as shown in To analyze meaningful results from PPG data, this paper defines the HSI indicator and HSI extraction process, and In order to verify the effect of the optimized breathing cycle on body stress, this paper defined seven stress-related indicators, and We recruited 100 participants who worked in telemarketing , aged between 20 and 50 years . The population ratio of participants was, respectively, 34%, 39%, and 27% for those in their 20s, 30s, and over the age of 40. The participants were physically healthy, and we gave them a small amount for active participation in the experiment. The average employment period was 11.8 months (SD = 14.7).The subjects worked a total of 8 h a day, 40 h per week. Additionally, when examining the characteristics of the subjects through the interview, the subjects did not usually have time to perform activities for a long time, and there were a total of four subjects who practiced breathing training such as yoga and abdominal breathing. The other subjects had no personal breathing training experience, and all subjects performed three training sessions in this experiment. We used the PPG sensor with a BIOPAC amplifier (Model PPG 100C) to record the PPG. We transferred the amplified signal to a LabVIEW platform via a National Instrument DAQ-6015 AC/DC converter. The measured location of the sensor was on the index finger of the left arm.We asked the participants to breathe using the presented visual guide of respiratory cycles. The respiratory cycles consisted of five stimuli with 9 s, 10 s, 11 s, 12 s, and 13 s, where each respiratory cycle consisted of inhaling and exhaling. For example, they repeated breathing in and out for 6.5 s each for 2 min when we asked subjects to perform the 13 s breathing cycle.We analyzed seven features, namely the percent and power of very low frequency (VLF), the percent and power of low frequency (LF), the percent and power of high frequency (HF), and the LF/HF ratio. The frequency band of VLF was 0.0033~0.04 Hz, the LF band was 0.04~0.15 Hz, and the HF band was 0.15~0.4 Hz.All dependent variables adhered to the within-subject design, and we performed non-parametric tests because they had non-normal distribution. The data analysis was performed based on the Wilcoxon signed-rank test between optimal RSP and none-optimal RSP.x-axis and y-axis represent the HSI and seven indicators, respectively. We confirmed the change in each indicator as the HSI value increased. To verify the effect on optimal respiration, we analyzed the seven features of HRV frequency, and p < 0.001, indicating a difference between optimal RSP  and non-optimal RSP . HF (%) also showed a significant result, p < 0.001, indicating a difference between optimal RSP  and non-optimal RSP .The LF (%) showed a significant main effect of optimal RSP, p < 0.05). The mean and SD of the optimal respiration of VLF power were 0.0004 (SD = 0.0004), and otherwise 0.0005 (SD = 0.0003). Those of HF power were 0.0043 (SD = 0.0038) for optimal breathing and 0.0035 (SD = 0.0029) for non-optimal breathing.The VLF and HF power showed a statistically significant difference (p < 0.001). The mean and SD of the optimal respiration of LF power were 0.0016 (SD = 0.0018), and otherwise 0.0029 (SD = 0.0018). Those of the LF/HF ratio were 1.1185 (SD = 3.021) for optimal breathing and 3.2043 (SD = 3.9283) for non-optimal breathing.The LF power and LF/HF ratio showed a significant difference  and acquired the PPG signals during this process. In the optimization process, we derived the HSI index after calculating the HF frequency through the collected data and selected the case with the highest HSI as the optimized RSP cycle per person. In the final validation step, we chose the seven stress-related indices as dependent variables by calculating the PPG signal, such as VLF (%), LF (%), HF (%), VLF power, LF power, HF power, and LF/HF ratio from the PPG signal. The experimental results show a statistical difference in the stress index in the optimal and non-optimal breathing case, and the tendency of the difference is consistent with the general tendency. Existing studies have shown that deep breathing has the effect of maintaining and changing a state of psychological comfort from negative states such as anxiety, stress, and anger, and its cycle affects the heart rhythm and activates the parasympathetic nervous system ,52,53,54In the analysis on the frequency domain, harmonics means a multiple of the fundamental frequency. For example, if the fundamental frequency is 2 Hz, high harmonies occur at 4 Hz, 6 Hz, etc. Because HRV is periodic information of the RR interval through frequency analysis, it has harmonic property. According to previous studies, the spectral amplitude within the HF band (0.15–0.4 Hz) was used as an index of RSA ,56,57. WThe pattern of HRV is used to analyze frequency data transformed from PPI fluctuations and has components such as fundamental, second, third, etc., due to frequency harmony. When the subjects performed optimal respiration, existing studies selected the subject’s optimal respiration using a dominant pattern in the LF region of HRV, which is the fundamental component ,60. HoweIn this context, we assumed that HF power could be used to check whether the subject’s parasympathetic nervous system is currently activated according to respiratory cycles, and we observed the HF power of the subjects according to the breathing cycle. As a result, each subject showed a dominant pattern at a specific frequency in the HF power range according to the respiratory cycle. The pattern of HF power showed a difference not only between individuals, but also the breathing cycle within individuals. For this reason, we used HSI, a quantification index based on HF components, to determine the optimal respiration for each individual.We tried to confirm the relationship between the HSI and the seven sympathetic and parasympathetic activation indices, which are linked to stress. This study conducted a validation on the effect of the parasympathetic activation between the optimal RSP and non-optimal RSP for the individual based on the HSI. This study proposed an overall process consisting of three steps—(1) application, (2) optimization, and (3) validation—to find an optimized respiratory cycle. We defined a performance index, or heart stabilization indicator (HSI), using data in the high-frequency (HF) range of heart rate variability (HRV) for selecting an optimized breathing cycle.In the first step, we measured the PPG data of the subjects based on the respiratory guidelines with random breathing cycles. Then, after calculating the HSI index using the proposed formulation and the PPG data, we selected the respiratory cycle showing the highest HSI index as the optimal respiratory cycle. In the last step, we verified the proposed method by comparatively analyzing the stress index related to the HRV frequency parameters in the case of optimal and non-optimal respiratory cycles.The experimental result in the two cases shows that there is a similar pattern compared with the previous studies from the perspective of the parasympathetic activation. Furthermore, there is a clear and significant statistical difference for the stress indices, except for VLF. In other words, according to the increase in the HSI, we confirmed that LF (%), LF power, and LF/HF ratio decreased, while HF (%) and HF power increased.Our findings clearly show that the HSI index can be a significant index for finding optimal respiration. The optimal respiration derived through the index can be applied as a reference value for the feedback, which is one of the main factors in biofeedback. Since the HSI indicator in this study is generated through resonance respiration, it is expected that there is the possibility for future research using it as a factor to check heart conditions such as arrhythmias as well as in the field of RSA biofeedback. Finally, we expect that the proposed design in this paper will used as an alternative for improving the effectiveness of RSA biofeedback.


The ESGO evidence-based guidelines cover the whole field of gynecologic oncology. The ESGO guidelines for the management of vulvar cancer were published in 2017 [When we proposed the topics for this Special Issue, our first thought was to integrate the results from the latest original research to practical reviews focusing on key questions. Both authors of this editorial are former presidents of international societies, Mario Preti from the International Society for the Study of Vulvovaginal Disease  [It is clear that the therapeutic approach has much improved, but we must not forget that in some countries there are different possibilities of access to treatment and prevention. The epidemiologic characteristics of vulvar squamous cell cancer (VSCC) represent an impediment to the adoption of screening strategies, so VSCC detection often occurs as an incidental finding during routine gynecologic visits. Indeed, VSCC prevention and early diagnosis also show a weak side in high-standard health services, even though scheduled cytologic or human papillomavirus (HPV) testing for cervical cancer screening could be an opportunity to perform a careful clinical inspection of the vulvar region, and to detect and treat multicentric HPV-related intraepithelial neoplasia. Even though only a small percentage of VSCCs are HPV-related , a recen< 0.001) . The ris< 0.001) ,11. ThisOn the other hand, lichen sclerosus is not a rare disease . There iMoving forward, what are our goals and the direction to dedicate expanded effort and resources ,17? We aThis Special Issue is composed of contributions from authors coming from diverse regions. All have extensive experience in their fields. They are international authorities and have submitted papers that provide cutting-edge evidence on multiple aspects of vulvar cancer. We wholeheartedly thank them for dedicating their time and sharing their expertise.


Introduction: Parents acquire information about human papillomavirus (HPV) vaccines online and encounter vaccine-critical content, especially on social media, which may depress vaccine uptake. Secondary analysis in a randomized trial of a Facebook-delivered adolescent health campaign targeting mothers with posts on HPV vaccination was undertaken with the aims of (a) determining whether the pre–post-change occurred in self-reports of the mothers on HPV vaccination of their adolescent daughters; (b) describing the comments and reactions to vaccine posts; (c) exploring the relationship of campaign engagement of the mothers assessed by their comments and reactions to posts to change in the self-reports of the mothers of HPV vaccination.Materials and Methods: Mothers of daughters aged 14–17 were recruited from 34 states of the US (n = 869). A social media campaign was delivered in two Facebook private groups that differed in that 16% of posts in one were focused on indoor tanning (IT) and 16% in the other, on prescription drug misuse, assigned by randomization. In both groups, posts promoted HPV vaccination  and vaccination for other disease . HPV and other vaccination posts covered the need for a vaccine, the number of adolescents vaccinated, how vaccines are decreasing the infection rates, and stories of positive benefits of being vaccinated or harms from not vaccinating. Guided by social cognitive theory and diffusion of innovations theory, posts were intended to increase knowledge, perceived risk, response efficacy , and norms for vaccination. Some vaccination posts linked to stories to capitalize on identification effects in narratives, as explained in transportation theory. All mothers received the posts on vaccination . Mothers completed surveys at baseline and 12- and 18-month follow-up to assess HPV vaccine uptake by self-report measures. Reactions  and comments to each HPV-related post were counted and coded.Results: Initiation of HPV vaccination (1 dose) was reported by 63.4% of mothers at baseline, 71.3% at 12-month posttest (pre/post p < 0.001), and 73.3% at 18-month posttest (pre/post p < 0.001). Completion of HPV vaccination (two or three doses) was conveyed by 50.2% of mothers at baseline, 62.5% at 12-month posttest (pre/post p < 0.001), and 65.9% at 18-month posttest (pre/post p < 0.001). For posts on HPV vaccines, 8.1% of mothers reacted , and 68.4% of posts received a reaction . In addition, 7.6% of mothers commented , and 50.0% of these posts received a comment. There were no differences in pre–post change in vaccine status by the count of reactions or comments to HPV vaccine posts (Ps > 0.05). Baseline vaccination was associated with the valence of comments to HPV vaccine posts (7.2% of mothers whose daughters had completed the HPV series at baseline made a favorable comment but 7.6% of mothers whose daughters were unvaccinated made an unfavorable comment).Conclusion: Effective strategies are needed in social media to promote HPV vaccines and counter misinformation about and resistance to them. Mothers whose daughters complete the HPV vaccine course might be recruited as influencers on HPV vaccines, as they may be predisposed to talk favorably about the vaccine. Comments from mothers who have not been vaccinated should be monitored to ensure that they do not spread vaccine-critical misinformation. Study limitations included lack of randomization and control group, relatively small number of messages on HPV vaccines, long measurement intervals, inability to measure views of vaccination posts, reduced generalizability related to ethnicity and social media use, and use of self-reported vaccine status.Clinical Trial Registration:www.clinicaltrials.gov, identifier NCT02835807. Despite the recommendation of the Advisory Committee on Immunization Practices (ACIP),  only 54%Parents frequently use the Internet as a reliable source of information on child health –11, and Social media, in particular, spread information on HPV vaccines and transmit vaccine-critical content , 32–34. Much of the past research on HPV vaccine content in social media has been descriptive and correlational , 26, 42,Mothers with teenage daughters were enrolled in a randomized controlled trial with the primary purpose of evaluating the effect of social media posts on their permissiveness toward indoor tanning (IT) by daughters. After a baseline survey, mothers were randomly assigned to one of two Facebook private groups. Randomization was accomplished by the project biostatistician, using permuted-block randomization (block size = 2). All mothers received a Facebook feed of posts on adolescent health topics, mother–daughter communication and current events, which differed on whether the feed included posts on IT (intervention) or prescription drug misuse (control). Both feeds contained posts on HPV vaccination and vaccination for other diseases  as there was no randomization on the presence of the HPV vaccination messages. A community manager added mothers to the groups, scheduled posts, and monitored and responded to comments during the 12-month campaign, after which mothers completed posttest surveys at 12- and 18-months post-randomization. To retain participants, any mothers who left the private groups were contacted and asked to re-join and mothers were alerted to upcoming posttests and compensated . Daughters were invited to complete the baseline survey and both posttests  but were not enrolled in the social media feed. Study staff other than the community manager and project coordinator were blinded. The Western Institutional Review Board (IRB) and the IRBs at East Tennessee State University and the University of Connecticut approved study protocols.Between May 2017 and June 2018, mothers were enrolled who met inclusion criteria: (1) having a daughter aged 14–17; (2) living in one of 34 U.S. states without a complete ban on IT by minors; (3) reading English; (4) having a Facebook account and logging in at least once per week; and (5) willing to “friend” the community manager of the project to join a private Facebook group. Mothers were excluded if they were unable to read English or did not consent and “friend” the community manager. Initially, mothers were recruited in Tennessee through community-based methods . When these methods did not yield a sufficient number of mothers, Qualtrics was contracted and recruited mothers from its survey panel in 33 other states. All mothers were blind to treatment because they received a social media feed whose purpose was described as providing information on adolescent health and mother-daughter communication. Statistical power calculations provided a target sample size of 860 that would achieve 80% power for small to moderate effects. Mothers provided email addresses for daughters and parental consent, and daughters were invited to complete assessments, providing informed assent. To avoid a major recruitment barrier, the participation of the daughter was not required. When mothers had more than one daughter, the one with the nearest birthday was selected.Health Chat, using principles of social cognitive theory (SCT)  , transpory (SCT) , and difry (SCT) . The camry (SCT) , 49. Posry (SCT) , 51. Posn = 90) not in the trial and changes were made to enhance aesthetics, message clarity, and engagement based on the results. Messages addressing current events were created during the intervention. Approximately 84% of posts addressed adolescent health topics and mother–daughter communication. Topics included vaccinations , mental health , substance use , healthy lifestyles , media literacy, and general parenting . They were selected based on formative research with mothers, engagement of the mothers during pilot testing, or emerging issues in comments of the mothers during the campaign. About 16% of posts focused on preventing IT (intervention group) or prescription drug misuse (control group). The two private Facebook groups received the same feed of posts except for the manipulated posts on IT or prescription drug misuse.Messages were created by investigators and reviewed by the entire team for acceptability and readability. Initial messages were pretested in a pilot feed with mothers  and general vaccine information, including influenza vaccination (n = 49 posts). Regarding HPV, posts covered the need for the vaccine (perceived risk), percent of adolescents vaccinated, the proportion of parents choosing to vaccinate children against HPV (descriptive norms), how HPV vaccines are decreasing infection rates (response efficacy), and stories of women who died from cervical cancer (perceived severity) or parents who decided to vaccinate their children (identification). Posts on vaccination for other diseases addressed misinformation surrounding vaccines, the need for annual influenza shots (risk), vaccine safety and efficacy (response efficacy), adolescent vaccine schedules (how-to knowledge), and reducing barriers to vaccination. Posts were in didactic  and narrative  format. Narratives were intended to influence through a process of identification with the characters in the stories. The primary focus on communicating with mothers was appropriate as they drive decisions about HPV vaccines . This rate was designed to be sufficient to influence but avoid message fatigue. A community manager scheduled posts, monitored reactions/comments, and replied to misinformation. In addition, mothers received a bi-weekly email newsletter highlighting the most popular recent posts.The primary outcome measures in the baseline and 12- and 18-month posttest surveys in this analysis were self-reports of HPV vaccination by mothers. Mothers were asked if the daughter had been vaccinated for HPV and if so, how many shots had she received. Initiation of vaccination was defined as receipt of one-shot of HPV vaccine and completion was defined as two or three shots (two shots are recommended for girls under age 15 while three shots are recommended for girls aged 15–17). Daughters were asked these same questions for themselves.Demographic characteristics were collected, namely, mother and daughter age and skin phenotype  . MothersThe engagement of the mothers with the HPV vaccination posts was measured by counting the number of reactions  and comments to each post. Comments and reactions to posts on HPV and other vaccines were extracted by a trained research assistant at the end of the social media campaign, using Grytics software. The content of reactions was recorded . Further, the content in the comments was coded by trained research staff. The comments were coded as favorable , unfavorable , or neutral . Each comment was coded by one research assistant and 60% were coded by a second coder to check inter-rater reliability . Emergent themes were then identified based on content codes.F-tests were utilized to determine if increases in HPV vaccination rates were statistically significant. Correlation coefficients were computed to compare mother and daughter reports. Multinomial logistic regression was fit to identify predictors of vaccine uptake from pre- to post-intervention. Alpha criterion level of 0.05 was set for all tests.Descriptive statistics were computed using SAS, Version 9.3. Demographic characteristics of the 869 mothers and 469 daughters enrolled in the study have been reported elsewhere . BrieflyF = 14.05, p < 0.001) and 62.5% reported that daughters had received two or three vaccine doses . Looking just at mothers whose daughters had not completed the HPV vaccine series at baseline , 18.5% of daughters with no shots at baseline and 53.0% of those who had received one shot at baseline had completed the series at the 12-month follow-up . At the 18-month posttest, 73.3% of mothers reported that daughters had received at least one dose of the HPV vaccine , and 65.9% reported that daughters had received two or three vaccine doses . Again, subsetting mothers whose daughters had not completed the HPV vaccine series at pretest , 22.2% of daughters with no shots at baseline, and 70.0% of those who had received one shot at baseline had completed the series at the 18-month follow-up . Reports of the mothers of HPV vaccine uptake were corroborated by daughters .At baseline, 63.4% of mothers reported that their daughters had received at least one dose of the HPV vaccine, with 50.2% saying they had received two or three doses . At the 12-month posttest, 71.3% of mothers reported that daughters had received at least one dose of the HPV vaccine (pre/post comparison n = 38), 8.1% of mothers reacted to a post , and 68.4% of all HPV vaccine posts received a reaction . In addition, 7.6% of mothers commented on an HPV vaccine post , and 50.0% of all HPV vaccine posts received a comment. Similarly, for posts on other vaccines, 5.4% of mothers reacted to a post  and 71.4% of the posts on other vaccines received a reaction . In addition, 4.6% of mothers commented on posts on other vaccines , and 42.9% of these posts on other vaccines received a comment. Looking at all posts in the feed, 55.8% of mothers reacted to a post and 68.2% of posts received a reaction. In addition, 58.5% of mothers commented on a post and 53.8% of posts received a comment.For social media posts on HPV vaccines  and other vaccine posts  did not differ by vaccine status at 18-month posttest.We explored the relationship between the engagement of the mothers with HPV and other vaccine posts and the status of the vaccine of their daughters. Specifically, we explored whether reactions and comments were associated with pre/post-change in vaccine status. We fit three multinomial logistic regressions, subsetting to include only mothers who reported that their daughters had not completed the HPV vaccine series at baseline. Counts of reactions and comments to HPV vaccine posts  of those who had not yet vaccinated their daughters at the beginning of the trial had completed the HPV vaccine series at either posttest than those whose daughters who had already had one dose at baseline (53% and 70%).A sizable group of mothers (27%) had daughters who remained unvaccinated throughout the trial and this group was more likely to post unfavorable comments, suggesting that some of these mothers with unvaccinated daughters were actively resistant to HPV posts. Vaccine-critical comments might be used to identify mothers who are resistant to HPV vaccines and tailor posts to respond to reasons for vaccine hesitancy expressed in their comments. By contrast, mothers who had initiated but not completed HPV vaccination did not comment much either favorably or unfavorably on posts about it. This latter group may have been uncertain about whether to complete it or had barriers to completion, but they were also not strongly resistant to the vaccine. The social media campaign may have nudged some of them to get the HPV vaccine series completed.Less than 10% of mothers engaged with the posts on HPV vaccination  but the number of posts that received a reaction or comment was similar in rate to all posts in the feed. The most common comment in response to HPV vaccine posts was a mother sharing that she had her daughter vaccinated. For this reason, mothers who complete the HPV vaccine series for their daughters might be recruited as influencers on HPV vaccines in future vaccine-promotion programs, as some appear predisposed to talk favorably about the vaccine. However, they may need to be instructed on how to make comments that are likely to influence other mothers because many just simply commented that they had vaccinated their daughters without providing other information that might be influential such as noting the benefits of disease prevention or that physicians recommended it. This same tendency, seen in an early analysis of comments to initial posts , continuThe comments that physicians did not support HPV vaccination need to be countered as medical professionals can be important, credible, and influential sources of information on HPV vaccination , 83–85. The study had several limitations. The lack of randomization and absence of a control group undermined conclusions about whether the social media campaign produced the observed change in HPV vaccination rates at posttest. The number of messages specifically on HPV vaccination was small  but they were supplemented by nearly 50 posts on other vaccinations  which should have improved vaccination intentions in general. The measurement times at 12- and 18-months post-randomization were long and risk history effects, where secular events  could have occurred after the HPV vaccination messages that influenced decisions of the parents to vaccinate daughters. Once again, the lack of randomization meant the design could not control for this threat to internal validity. The inability to collect data on views from the Facebook feed meant we could not determine whether mothers saw the HPV and other vaccination posts. The decision by Facebook to eliminate reports on views of posts in the private groups was out of our control and occurred after the study was launched. We contacted Facebook and requested access to the views data but they were unable to provide it. Instead, we decided to use counts of reactions and comments to the vaccination posts, which were still reported by Facebook, as indicators of viewership, but as noted above these likely under-estimated actual engagement with the vaccination posts. It is worth noting that the lack of an objective measure of exposure to campaign messages by individual respondents often is limited in community-based evaluations of public health campaigns that rely on other media such as television, radio, print, and billboards. While the sample was large and included mothers from 34 states of the US, it may have limited generalizability because mothers were predominately non-Hispanic white and had regular social media use, the Qualtrics survey panel tends toward participants with higher socioeconomic status, mothers who chose to participate were interested in their health of the daughters, mothers lived in states that may be less socially progressive as they did not have bans on indoor tanning by minors, and daughters were older than the first age at which HPV vaccination is recommended. Self-reports of vaccine status of the mothers also may be biased but these measures have shown good specificity and sensitivity in past surveys –91 and pHuman papillomavirus vaccination rates in the United States continue to lag national health goals. Social media is a major source of health information and supportive and critical information on HPV vaccination. Effective strategies are needed in social media to promote HPV vaccines and counter misinformation about them to move mothers who are resisting vaccination. Interspersing vaccine messages in a feed for parents may be effective, as it can expose them to a large number of messages. Even if social media messaging does not change the minds of highly resistant mothers who contribute unfavorable comments, carefully crafted messages may convince mothers who are uninformed, uncertain, or not currently taking action to vaccine their daughters to ignore or resist the false claims and misinformation about the HPV vaccine and complete the series for their daughters.The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.The studies involving human participants were reviewed and approved by The Western Institutional Review Board and the IRBs at East Tennessee State University and University of Connecticut. Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.DB: conceptualization, methodology, verification, investigation, writing—original draft, visualization, supervision, project administration, and funding acquisition. SP: conceptualization, methodology, verification, investigation, resources, writing—review and editing, supervision, project administration, and funding acquisition. KH: conceptualization, methodology, formal analysis, writing—review and editing, and funding acquisition. JBe: verification, validation, investigation, resources, data curation, writing—review and editing, and supervision. BW: conceptualization, methodology, investigation, writing—review and editing, resources, and funding acquisition. JBi: verification, validation, investigation, resources, data curation, writing—review and editing, and supervision. KB: conceptualization, methodology, investigation, writing—review and editing, supervision, project administration, and funding acquisition. JH: conceptualization, methodology, investigation, writing—review and editing, and funding acquisition. KA: verification, validation, investigation, data curation, and writing—review and editing. All authors contributed to the article and approved the submitted version.DB's spouse is an owner of Klein Buendel, Inc. and he receives a salary from Klein Buendel. JBe and BW receive a salary from Klein Buendel, Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Two control individuals whom had received three doses of the vaccine (HPV16/31-negative) were selected per case, matched on age at the first dose of vaccination and follow-up time in the study: a random control, and a high-risk control that was in the upper quartile of a sexual risk behavior score. We conducted an enzyme-linked immunosorbent assay (ELISA) for the detection of immunoglobulin G (IgG) antibodies specific to anti-HPV16 virus-like particles (VLPs). The results suggest that the average log antibody titers were higher among high-risk controls than the HPV16/31 incident cases and the randomly selected controls. We show a prospective association between anti-HPV16 VLP titers and the acquisition of an HPV16/31 incident infection post-receiving three doses of 4vHPV vaccine.The goal of this study was to investigate the serological titers of circulating antibodies against human papillomavirus (HPV) type 16 (anti-HPV16) prior to the detection of an incident HPV16 or HPV31 infection amongst vaccinated participants. Patients were selected from a prospective post-HPV vaccine longitudinal cohort at Mount Sinai Adolescent Health Center in Manhattan, NY. We performed a nested case–control study of 43 cases with incident detection of cervical HPV16 ( Cervicovaginal infection by human papillomavirus (HPV) is the most common sexually transmitted infection in young adults. Though most HPV infections are transient, some develop into high-grade squamous intraepithelial lesions or invasive cervix cancer. HPV16 is the most carcinogenic HPV type circulating worldwide and still represents over 50% of the cervix cancers identified . In contSince HPV vaccine licensure in 2006, indisputable evidence has accumulated establishing the effective prevention of HPV infection and squamous intraepithelial lesion development ,5,6. FurAntibodies to conformational epitopes on synthetically produced virus-like particles (VLPs) are morphologically indistinguishable from authentic HPV virions and have been shown to develop in response to infection ,24,25,26Detection of serum antibodies to HPV capsids is a valid marker for immune response prior to vaccination . Since a®, Merck & Co., Inc., Kenilworth, NJ, USA) targeting HPV types 6, 11, 16, and 18. The study includes follow-up visits every six months until 25 years of age, with a collection of information by a self-reported questionnaire on lifetime and recent sexual behaviors, including the number of vaginal, anal, and oral sex partners, and a gynecological examination. Serum samples were collected at the enrollment visit, within six months of receipt of a third vaccine dose if later, and every two years thereafter for all returning participants. Additional details on the study design and protocol are described elsewhere [Study participants were selected from a large ongoing prospectively enrolled Phase 4 HPV vaccine cohort study of adolescent female patients attending Mount Sinai Adolescent Health Center (MSAHC) in New York City as previously described ,34. Studlsewhere . The Insn = 26) or HPV31 (n = 17) after three doses of 4vHPV amongst baseline HPV16/31 DNA-negative individuals. Two control individuals (HPV16/31-negative) per case were selected. A random control (RC) was defined as being HPV16/31-negative throughout the study period, having received three doses of the vaccine and matched to a case on age at first dose of 4vHPV (±one year) and follow-up time. A high-risk control (HRC) was defined as a participant who was in the upper quartile of a sexual risk behavior score (score > 9), HPV16/31-negative throughout the study period, and matched to cases on age at first dose of 4vHPV (±one year) and follow-up time with complete vaccination (three doses). A linear sexual risk behavior score was derived based on the following variables, as shown in Table 1: lifetime vaginal sex partners (1–4), recent (past six months) number of partners (0–2), history of chlamydia trachomatis (0–1), any pregnancy (0–1), emergency contraception used (0–1), condom usage (0–1), ever anal sex (0–1), and lifetime number of anal sex partners (0–2). The linear score ranged from 0 and 13, and more weights were given for the number of sex partners.We used a nested case–control study design. The criteria for the case groups were incident detection of cervical HPV16  in cervical Pap specimens collected at baseline (enrollment) and each follow-up clinic visit. Details on specimen collection, HPV DNA detection, and typing have been described in detail elsewhere ,33,34,35Serum samples were tested for antibodies to HPV16 VLPs as previously described ,37. BrieThe ELISA protocol was performed as described . Serum sn = 43; average time since the last dose of 4vHPV 3.0 ± 2.9  years); (2) at the time of incident detection of cervical HPV16/31 ; and (3) the serum obtained after HPV16/31 incident detection . Serum samples from controls were matched by time-to-case pre-infection samples .Serum samples from cases were tested at three time points: (1) blood draw immediately prior to the visit in which the incident detection of cervical HPV16/31 occurred  and case–control status (incident HPV16/31) was investigated using conditional logistic regression. A logistic regression model was fitted to assess the log serum titers’ association with HPV infection under subgroup analysis.Descriptive statistics were used to summarize adolescent and youth characteristics. Continuous scale variables were summarized using mean and standard deviation; non-normal variables were log-transformed. The difference in continuous variables was assessed using analysis of variance and An analysis examining the influence of infection on log serum titers post-vaccination among cases was performed. For this analysis, the changes in log serum titer before and after infection were fit using a piecewise linear mixed-effects model with random intercept and slope. Initially, actual time was computed as the difference between the last vaccination (dose 3) and pre-, peri-, and post-infection dates. Further, (relative) time for the model was recoded as time since infection and could be positive or negative. The estimation was performed using a restricted maximum likelihood approach with Kenward–Roger degrees of freedom adjustment. This model allowed for the modeling of inherently unbalanced repeated measures data and missing information under “missing at random” assumptions. All analyses were performed using SAS software version 9.4 .n = 1398) through May 2021, of whom 43  developed infection after having completed the full set of immunizations . The selected individuals (43 cases and 86 controls) were 18 ± 1.3 (SD) years of age, and 95% reported being either Hispanic or African American. Over 62% (81/129) had more than three lifetime sexual partners. The average ages at coitarche and receiving the first vaccine dose were 14.7 ± 1.4 (SD) and 15.2 ± 2.3 (SD) years, respectively. Slightly more than one-third (38%) initiated sexual activity after they had at least one dose of vaccination. We selected two different control groups to compare women with incident HPV16 or HPV31 infections. Specifically, we included a random control group (RC) that was matched on age at first dose of 4vHPV, having completed three doses of HPV vaccine and being HPV-negative for HPV16 and 31. We selected a second control group (HRC), reasoning that women with high risk for a cervical HPV infection should have higher antibody levels, affording greater protection consistent with no detectable HPV16 or HPV31 infections after immunization.The characteristics of the participants in this nested case–control study are presented in Titration curves of anti-HPV16 VLP antibody titers were determined for all cases and controls in duplicate. As previously described, a serological titer equal to 0.2 OD units was defined as the seropositivity cutoff value . Mean sep = 0.07 level among the groups using the analysis of variance procedure. Compared to each control separately, the difference between case titers was significant with high-risk controls (p = 0.01) but not with random controls (p = 0.19)  .p = 0.04), adjusting for the receipt of the 4vHPV before sexual coitarche vs. not, as shown in p = 0.04) of having a one-unit increase in log antibody titers. However, the relative odds  were attenuated and were not significant (p = 0.21) compared to the random control group. Thus, high-risk controls had higher antibody titers compared to random controls and cases. A subgroup analysis of HPV16 or HPV31 types suggested similar scenarios with higher odds among the controls than cases for one log unit increase in antibody titers. This observation was more pronounced among HPV31 incident cases, where we observed the odds ratio was 67%  higher among the controls when compared to cases  had a 41% higher likelihood (relative odds) of having a one log unit increase in antibody titers compared to the cases , while the average log serum titers decreased by 0.11 units (SE: 0.13) from peri- to post-infection (p = 0.41). This case analysis suggests that potential “false negative” undetected HPV infections were not driving the higher antibody titers detected in the high-risk control group.Based on the piecewise linear mixed model, the average log serum titers increased from pre- to peri-infection by 0.16 units , although this was not statistically significant  prevented the acquisition of new HPV16/31 infections, or if continuous or frequent exposure  induced the higher titers detected in the high-risk control group.p = 0.037). The results included a phylogenetic group approach of different HPV types, in which the presence of prior anti-HPV16 antibodies were markers for immunity against HPV types other than HPV16. Based on the cross-protection of HPV31 by HPV16 vaccination [Though neutralization assays are often considered the “gold standard” in assessing the protective immunogenicity of prophylactic vaccines, they are highly correlated to ELISA titers. Thus, the anti-HPV16 VLP ELISA represents a surrogate assay for neutralizing anti-HPV IgG antibodies ,37. Addicination ,41,42, wp = 0.08). Studies conducted in females vaccinated < 15 years of age have shown high effectiveness in reducing HPV vaccine type prevalence [Real-life evidence of HPV vaccine effectiveness has become increasingly evident, especially in females vaccinated before sexual exposure to HPV . In our evalence ,45,46,47evalence .Vaccine clinical trial studies on bivalent and quadrivalent HPV vaccines have shown an efficacy of over 80% and 36% in preventing HPV16 or HPV31 persistent infections, respectively ,49. In oIn this study, we also evaluated the serum titers in cases at three time points . We observed a stable level of antibody titer over the window of examination. This implies that infection with a vaccine-type HPV did not boost the level of antibody titers acquired after immunization with three doses. Therefore, the higher antibody titers detected in the high-risk control group were unlikely to be related to prior HPV infections. We would argue that we did not detect incident HPV16/31 infections in this group because of their robust antibody titers from vaccination.There are some limitations to this study. One limitation is the sample size, directly correlated to the low prevalence of HPV16/31 infections in our MSAHC vaccinated cohort. Another limitation is that we did not directly measure neutralizing antibodies, although we believe titers against HPV16 VLPs are a reasonable surrogate. Lastly, we could not absolutely control for exposure to HPV16/31 infections in both groups.In summary, it is accepted that HPV vaccines substantially modify the incidence and duration of HPV16-induced cervicovaginal neoplasia by restricting virus infections through neutralizing antibodies. We show a prospective association between anti-HPV16 VLP antibody titers and the acquisition of an HPV16/31 infection subsequent to the receipt of three doses of the 4vHPV vaccine. Continued follow-up of vaccinated individuals will be required to determine if waning antibody titers might correlate with an increased risk of HPV infection. However, the elimination of HPV16 and other types by breaking the chain of transmission could effectively wipe out the burden of future infections and related neoplasia and cancers.


We investigated the relationship between attentional resources and pedaling cadence using electroencephalography (EEG) to measure P300 amplitudes and latencies. Twenty-five healthy volunteers performed the oddball task while pedaling on a stationary bike or relaxing . We set them four conditions, namely, (1) performing only the oddball task , (2) performing the oddball task while pedaling at optimal cadence , (3) performing the oddball task while pedaling faster than optimal cadence , and (4) performing the oddball task while pedaling slower than optimal cadence . P300 amplitudes at Cz and Pz electrodes under optimal, fast, and slow conditions were significantly lower than those under control conditions. P300 amplitudes at Pz under fast and slow conditions were significantly lower than those under the optimal condition. No significant changes in P300 latency at any electrode were observed under any condition. Our findings revealed that pedaling at non-optimal cadence results in less attention being paid to external stimuli compared with pedaling at optimal cadence. Humans experience situations that require the execution of multiple tasks simultaneously. Dual tasks, where individuals perform two tasks simultaneously, degrade the performance . EspeciaIn previous studies, participants were imposed a dual task, namely, a cognitive task and an alternate leg movement. Performing cognitive tasks during walking reduced not only gait parameters but also the cognitive ability . NotablyIn our study, to clarify attentional resources during alternating movement of lower limbs as a function of gait speed, participants were asked to perform oddball tasks while pedaling on a stationary bike. Pedaling has three advantages. First, the trunk and head of the participants are stabilized during pedaling compared to walking. Second, the pedaling device is light and portable. Third, studies investigating characteristics of brain activity reported that EEG can be performed during pedaling . TherefoP300, an event-related potential (ERP) component, is suited well to testing attention. Studies have reported that the P300 peak amplitude is proportional to attentional resources devoted to a given task, and latency reflects stimulus evaluation time . P300 caSensory inputs, including visual, auditory, and somatosensory ones, allow detecting the changes in the environment. To effectively function in our environment, we employed a set of neural mechanisms that extract the sensory inputs, which are most relevant to our current goals . TherefoWe determined the minimum sample size of the present study using the G*power software from partial η-squared. This η-squared value was determined from the previous study . The effElectrical stimuli were randomly presented to the right index and fifth finger for 0.2 ms through ring electrodes attached to the first  and second  interphalangeal joints. The index finger was stimulated for the target stimulus and the fifth finger for the standard stimulus. The stimulus intensity was adjusted to three times the sensory threshold of the participants . StimuliAll participants performed the oddball task during either pedaling on a stationary bike or relaxing , and EEG was recorded. Participants fixated on the crosshairs in the middle of the screen during EEG recording. They were asked to perform two tasks, namely, motor and count tasks. In the motor task, participants were instructed to pedal on a stationary bike at optimal cadence, 30% faster than optimal cadence, and 30% slower than optimal cadence. We fixed the optimal cadence that makes the participants pedal comfortably. Participants were trained to maintain a smooth consistent pedaling cadence in synchrony with a metronome before recording EEG. In the count task, participants silently counted the number of target stimuli presented to their index finger on the stationary bike. We set them four conditions, namely, (1) performing only the count task , (2) performing the count task during pedaling at optimal cadence , (3) performing the count task while pedaling faster than optimal cadence , and (4) performing the count task during pedaling slower than optimal cadence . One run varied randomly between 80 and 120 epochs (mean: 100 epochs), so the participants could not estimate the number of target stimuli. All participants performed three runs at each condition . The order of these tasks was randomized for each run. Participants had time to recover between runs. We used the modified Borg scale to measure the perceived intensity of physical activity of the participants. We recorded the rating of perceived exertion before and after each run. Before the recording session was started, all participants practiced motor tasks until they felt comfortable.Using the Neuropack system , PowerLab, and LabChart software , EEG and EMG signals were digitized and recorded. A study reported that P300 amplitudes are observed better on the midline, especially Pz . EEG wasPedaling could lead to ocular artifacts that synchronize with movement . TherefoElectromyogram was measured using a pair of Ag/AgCl electrodes  mounted over the right vastus medialis (VM) and the short head of biceps femoris (BF). EMG signals  were sampled at 1,000 Hz  and bandpass filtered at 1–300 Hz. Then, the full-wave rectification and smoothing were performed. The values of moving window were 501 ms. EMG signals were normalized to maximum voluntary contractions. The averages of normalized EMG values over three pedal cycles were calculated. t-test was performed on P300 under two stimuli between the target and standard. Then, repeated measures ANOVAs were performed to determine the effect of each condition  on the amplitude and latency of P300 elicited by target and standard stimuli, the correlation coefficients between EEG and EOG, EMG, error rate, and physical activity intensity. The post-hoc tests were performed for significant differences in ANOVAs, with the Bonferroni correction for multiple comparisons. In addition, we analyzed bivariate correlations between P300 amplitudes and EMG activities. The significance was set at p < 0.05.Statistical analyses were performed using the IBM SPSS 25 software , and all data were expressed as the mean ± SD. The Shapiro–Wilk test was used to assess the normality. To analyze the assumption of sphericity prior to repeated measures ANOVA, Mauchly’s test of sphericity was used; if the result of the test was significant, indicating that the sphericity assumption was violated, the Greenhouse–Geisser adjustment was used to correct this violation. The paired Optimal, fast, and slow cadence were 51.56 ± 9.67, 65.76 ± 13.99, and 35.88 ± 7.11 rpm, respectively. The Shapiro–Wilk test confirmed that all data, except physical activity intensity and %EMG, were normally distributed.F = 4.047, p = 0.011; Cz, F = 21.057, p < 0.001; Pz, F = 16.551, p < 0.001] but not P300 latency . Post-hoc tests revealed that P300 amplitude at Fz decreased under fast and slow conditions compared with that under the control condition . P300 amplitude at Cz decreased under optimal, fast, and slow conditions compared with that under the control condition  and under the slow condition compared with that under the optimal condition (p = 0.037). P300 amplitude at Pz decreased under optimal, fast, and slow conditions compared with that under the control condition  and under fast and slow conditions compared with that under the optimal condition .p = 0.098; optimal, p = 0.029; fast, p = 0.041; slow, p = 0.128; Cz: control, p < 0.001; optimal, p < 0.001; fast, p = 0.007; slow, p = 0.006; Pz: control, p < 0.001; optimal, p < 0.001; fast, p < 0.001; slow, p < 0.001). No significant differences were found in the latencies of P300. In contrast, the different conditions exerted significant effects on P300 amplitude . When evoked by standard stimuli, P300 amplitude decreased under optimal, fast, and slow conditions compared with that under the control condition , with no significant differences among the optimal, fast, and slow conditions. No significant differences were found regarding the latencies of P300.The P300 amplitude of all conditions, except the control and slow conditions at Fz, decreased under standard stimuli in contrast to the target stimuli . The different conditions had no significant effects on the correlation coefficients. These findings suggest that pedaling did not lead to ocular artifacts that synchronize with movement nor did it have any effect on P300.p < 0.001; BF, p = 0.005). The post-hoc analysis showed that the EMG activity in VM increased under the fast condition compared with that under slow and optimal conditions  and under the optimal condition compared with that under the slow condition . When the relationship between P300 amplitude and %EMG was analyzed by Spearman’s rank correlation test, no significant correlation under any condition was observed  = 5.020, p = 0.003]. The post-hoc analysis showed that the error rate increased under the slow condition compared with that under the control condition (p = 0.005). Intensities of physical activity were not significantly different under any condition.The results showed that (1) P300 amplitudes elicited by target stimuli decreased under optimal, fast, and slow conditions relative to the control condition at Cz and Pz, (2) P300 amplitudes elicited by target stimuli decreased under fast and slow conditions compared with that under the optimal condition at Pz, and (3) P300 amplitudes elicited by target stimuli were not significantly different between fast and slow conditions at Fz, Cz, and Pz. Our results showed that the optimal cadence is slow compared with that in previous studies . It coulStudies have reported that P300 amplitude is proportional to the amount of attentional resources devoted to a given task  and decrIn contrast, the P300 amplitude elicited by target stimuli at Pz under fast and slow conditions decreased compared with that under the optimal condition. A study suggested that pedaling at optimal cadence may be a good reflection of movement frequency output generated by the central pattern generator CPG; . CPG, whIn this study, P300 amplitudes elicited by target at all electrodes were not significantly different between fast and slow conditions. We hypothesized that the P300 amplitude increased under the fast condition and decreased under the slow condition . In thatIn this study, the P300 amplitude decreased under standard stimuli compared with that under target stimuli. These results indicate that the oddball paradigm was successfully implemented. However, there were no significant differences between the control and slow conditions on Fz. The frontal component could reflect novel events or passive attention. The posterior component could reflect active attention or activation by task-relevant events, such as targets . In our Our results revealed that EMG activities in the VM and BF increased under the fast condition compared with slow and optimal conditions. A study reported that the value of EMG is decided by the activation of the primary motor cortex , and P30The error rate of the slow condition was the highest; thus, slow conditions may be the most difficult tasks. A study revealed that executing smooth rhythmic motions very slowly is challenging for humans . SimilarIn this study, no significant changes in the P300 latency were observed under any condition. Studies have reported that the P300 latency is related to stimulus evaluation . Kida etThis study has several limitations. First, this study has not considered the effect of arousal. The P300 amplitude is associated with arousal . While, This study assessed the relationship between cadence of alternating movement of lower limbs and attentional resources. Our findings indicated that pedaling at non-optimal cadence results in less attention being paid to external stimuli compared with pedaling at optimal cadence.The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.The studies involving human participants were reviewed and approved by the Sapporo Medical University (No. 1-2-62). The patients/participants provided their written informed consent to participate in this study.MA, KS, and HS performed the material preparation and data collection. MA, KI, and TS performed the formal analysis. MA wrote the first draft of the manuscript. KS wrote the review and editing of the manuscript. All authors commented on previous versions of the manuscript, contributed to the study conception and design, and 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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.