Characterization of type-specific HPV prevalence in a population of persistent cutaneous warts in Flanders, Belgium.

Cutaneous warts are benign skin lesions caused by the human papillomavirus (HPV). Even though they are considered benign, they can have a considerable impact on the quality of life and cause serious illness in certain immunocompromised populations. Studies have shown that the efficacy of wart treatment is dependent on the causative HPV type. Therefore, in this article, we aim to determine the HPV genotype-specific prevalence in cutaneous warts of a Flemish population as part of the Omnivirol-Salycilic acid randomized controlled trial. Swab samples of cutaneous warts (n = 269) were collected during enrollment. The DNA extraction was performed on the automated NucliSENS® easyMAG® system (bioMérieux). The samples were analyzed with two separate in-house PCR assays capable of detecting the most prevalent cutaneous HPV types (i.e. wart-associated HPV qPCR) as well as the most relevant mucosal types (i.e. RIATOL qPCR assay). In total, the type-specific prevalence of 30 distinct HPV genotypes was determined. The beta-globin gene was used as a cellularity control and for viral load quantification. Data concerning wart persistence, previous treatment, wart type, and other relevant wart and patient characteristics was collected through a baseline questionnaire. The study population consisted mostly of persistent warts considering that 98% (n = 263) of the sampled skin lesions were older than six months and 92% (n = 247) had undergone previous treatment. The most prominent wart type was the mosaic verruca plantaris (42%, n = 113). The most prevalent HPV types were cutaneous HPV types 27 (73%, n = 195), 57 (63%, n = 169), and 2 (42%, n = 113). Only 2% (n = 6) of the lesions was HPV negative. The highest median viral loads were observed with HPV27 and 57 (i.e. 6.29E+04 and 7.47E+01 viral copies per cell respectively). The multivariate analysis found significant associations between wart persistence and certain wart types, the number of warts, and HPV genotypes. Based on these findings, persistent warts are more likely to: (1) be verruca vulgaris, verruca plantaris simple or mosaic, (2) to manifest as multiple warts, (3) and to be negative for HPV type 2 or 4. These characteristics can be useful in the clinical setting for future risk stratification when considering treatment triage and management. Trial registration: NCT05862441, 17/05/2023 (retrospectively registered).

Intra- and interlaboratory reproducibility of the RIATOL qPCR HPV genotyping assay.

The implementation of cervical screening based on human papillomavirus (HPV) continues to progress rapidly across countries. Evidence has shown that assays detecting high-risk human papillomavirus (hrHPV) deoxyribonucleic acid (DNA) are more effective than cytology-based screening. Validation of new hrHPV DNA assays requires both noninferior clinical accuracy compared to a standard comparator for cervical precancer and good reproducibility. This study builds upon previous diagnostic accuracy assessments of the RIATOL HPV genotyping qPCR assay and aims to evaluate the international validation criteria for reproducibility. The intra- and interreproducibility of the RIATOL-qPCR assay were assessed using 550 remnant cervical cell material from the cytology archive of the National Reference Center for HPV in Belgium. Specimens were collected in the context of cervical cancer screening and tested in two different laboratories. The international reproducibility criteria include the lower bound of 95% confidence interval of the intra- and interlaboratory agreement regarding the detection of hrHPV DNA exceeding 87% with kappa ≥0.50. The RIATOL-qPCR assay demonstrated excellent intralaboratory reproducibility, achieving an overall agreement of 98.2 (95% CI 96.6-99.1%) and a kappa of 0.96. Interlaboratory testing showed an overall agreement of 98.5 (95% CI 97.1-99.4%) with a kappa of 0.97. The RIATOL-qPCR assay fulfills the third criterion for HPV test reproducibility requirement for use in cervical cancer screening.

Optimizing the pre-analytical phase for accurate HPV detection in skin disorders: insights from a cutaneous warts case study.

Background: In previous years, several cutaneous disorders have been associated with human papillomavirus (HPV); however, the exact role of HPV remains largely unknown. The lack of optimization and standardization of the pre-analytical phase forms a major obstacle. The aim of this study was to develop an accurate/patient-friendly sampling method for skin disorders, with cutaneous warts as a case study. Methods: Various sample processing techniques, pre-treatment protocols and DNA extraction methods were evaluated. Several sampling methods were examined, that is, skin scrapings, swabs and a tape-based method. Quantification of DNA yield was achieved by beta-globin real-time polymerase chain reaction (qPCR), and a wart-associated HPV genotyping qPCR was used to determine the HPV prevalence. Results: All samples tested positive for beta-globin. Skin scrapings had significantly higher yield than both swab and tape-based methods (p < 0.01), the latter two did not significantly differ from each other (p > 0.05). No significant difference in DNA yield was found between cotton and flocked swabs (p > 0.05). All swabs were HPV positive, and although there were some discrepancies in HPV prevalence between both swabs, an overall good strength of agreement was found [κ = 0.77, 95% CI (0.71-0.83)]. Conclusion: Although skin scrapings produced the highest DNA yield, patient discomfort was an important limitation of this method. Considering that in combination with our optimized DNA extraction procedure, all samples gave valid results with the less invasive swab methods preferred. Standardization of the pre-analytical phase is the first step in establishing a link between HPV and specific skin disorders and may have significant downstream diagnostic as well as therapeutic implications.

Development and validation of a wart-associated human papilloma virus genotyping assay for detection of HPV in cutaneous warts.

Cutaneous warts are infectious disorders caused by human papillomavirus (HPV). A recent study revealed that the HPV genotype influences the natural course and response to treatment for plantar warts, suggesting that HPV genotyping could potentially be used to optimize wart treatment schemes. For this purpose, a wart-associated HPV genotyping assay was developed. The assay was subjected to an intensive validation process including, i.a., empiric determination of the annealing temperature, primer-probe optimization, evaluation of the analytical specificity and sensitivity, viral load quantification, and qualitative as well as quantitative analysis of intra-run repeatability and inter-run reproducibility. The newly developed assay was employed in a small-scale HPV genotyping study of wart biopsies (n = 50). The assay exhibited an analytical type-specific sensitivity and specificity of 100% (95% confidence interval [CI]: 83.9%-100%). The limit of quantification of the tested sequences corresponded to less than 17 viral copies/µl, while the limit of detection was less than 5 copies/µl. Very good to excellent agreements were gained between intra- and inter-run measurements (κ = 0.85-1.00) and coefficients of variation of the quantitative agreements were less then 3%. 22.5% (95% CI: 11%-39%) of the analyzed biopsies were negative for the tested HPV types, while 35% (95% CI: 21%-52%) contained multiple infections. The wart-associated HPV quantitative polymerase chain reaction assay was proven to be highly sensitive and specific. Multiple HPV infections were detected in 35% of lesions, contradicting the current literature claiming that in immunocompetent patients only 4%-16% of warts exhibit multiple HPV infections. This assay is qualified to be implemented in development of future genotype specific wart treatment strategies.

Human Papillomavirus Prevalence in Oral and Oropharyngeal Rinse and Gargle Specimens of Dental Patients and of an HIV-Positive Cohort from Pretoria, South Africa.

INTRODUCTION: Studies on HPV prevalence in the head and neck region of South Africans are sparse. Of the available reports in the literature, there were no studies on the association between HPV-DNA presence in the mouth and oropharynx in relation to high-risk behaviours such as oral sex practice or tobacco and alcohol use. MATERIALS AND METHODS: Following ethical clearance and informed consent, patients attending a regional HIV-management clinic and patients attending a dental hospital were recruited to this study. The participants completed an interview-based questionnaire obtaining demographic information, data on HIV serostatus, and behavioural data including sexual practices and tobacco and alcohol use, and a rinse-and-gargle specimen was taken. Specimens were analysed for HPV DNA on 3 separate PCR/qPCR platforms. Statistical analyses were performed for associations between the study group and categorical variables, HPV status, and data from the questionnaires. RESULTS: Of 221 participants, 149 were from a general population and 72 from the HIV-management clinic. Smokers comprised 29.4% of the sample, and 45.2% of participants reported to have ever used alcohol. Open mouth kissing during teenage years was confirmed by 64.7% of participants, 40.3% have given oral sex with their mouth, and 44.8% confirmed to have received oral sex from their partner's mouth. Seven participants (3.2%) had detectable α-HPV DNA, and 1 (0.4%) had detectable ß-HPV DNA in their rinse-and-gargle specimens. Two participants were from the HIV-management clinic and 6 from the general dental population (overall 3.6%). CONCLUSION: Five high-risk HPV, 2 low-risk HPV, and one ß-HPV types were detected. The low prevalence of 3.6% compares well to similar studies in different cohorts studied in South Africa and falls within the global oral/oropharyngeal prevalence spectrum. Only 4 participants, all from the HIV-management clinic, had palatine tonsils. No significant relationships were found between HPV presence and demographic data or sexual, oral sexual, tobacco use, or alcohol use, and no associations were seen with numbers of sexual and oral-sex partners.

Efficacy of AV2-Salicylic acid combination therapy for cutaneous warts: Study protocol for a single-center randomized controlled trial.

Cutaneous warts comprise an extremely common condition caused by infection with the human papillomavirus (HPV). Although most verrucae will disappear spontaneously, many patients do seek treatment. Current wart treatments do not target the cause of the lesion directly, resulting in variable treatment efficacies and high wart recurrence rates. AV2 is a broad-spectrum antiviral drug, that is capable of deactivating HPV. It is however not able to destruct the already infected cells, which raises the need for an additional ablative treatment i.e. salicylic acid (SA). Implementation of AV2-Salicylic acid (AV2-SA) combination therapy would ensure permanent lesion clearance by on the one hand inactivation of HPV by AV2, and on the other hand elimination of the lesion by SA treatment. The primary aim of this study is to assess the efficacy of AV2-SA treatment versus standard SA treatment, by comparing cure and recurrence rates of cutaneous warts between the two treatment groups (at 12 weeks and six months after randomization). The second aim is to assess the safety and tolerability of AV2-SA therapy. The third aim is to identify subgroups of cutaneous warts that have favorable response to treatment, by comparing cure rates in an HPV genotype-specific manner. This randomized controlled trial will enroll 260 participants with cutaneous warts who will either receive the AV2-SA combination therapy or SA control treatment. Real time monitoring will be possible by daily photographs sent via WhatsApp™ (a messaging application) as well as online follow-up questionnaires administered on several occasions. HPV genotyping will be performed on swab self-samples.