Assessing Gammapapillomavirus infections of mucosal epithelia with two broad-spectrum PCR protocols.

BACKGROUND: Human papillomaviruses (HPVs) have been divided into mucosal and cutaneous types according to their primary epithelial tissue tropism. However, recent studies showed the presence of several cutaneous types in mucosal lesions and healthy mucosa from different anatomical sites. METHODS: Here, the HPV prevalence and type-specific distribution were assessed in a variety of mucosal samples from 435 individuals using a combination of two established broad-spectrum primer systems: Gamma-PV PCR and CUT PCR. RESULTS: Overall HPV prevalence in anal canal swabs, cervical cancer biopsies, genital warts and oral swabs was 85, 47, 62 and 4%, respectively. In anal canal swabs, Alpha-PVs were most frequently found (59%), followed by Gamma- (37%) and Beta-PVs (4%). The prevalence and persistence of HPV infection in the anal canal of 226 individuals were further explored. Overall HPV, Gamma-PVs and multiple HPV infections were significantly higher in men vs. women (p = 0.034, p = 0.027 and p = 0.003, respectively); multiple HPV infections were more common in individuals ≤40 years (p = 0.05), and significantly higher prevalence of Gamma-PVs and multiple HPV infections was observed in HIV-1-positive vs. HIV-1-negative individuals (p = 0.003 and p = 0.04, respectively). Out of 21 patients with follow-up anal swabs, only one persistent infection with the same type (HPV58) was detected. CONCLUSIONS: Our findings suggest that Gamma-PVs (except species Gamma-6) are ubiquitous viruses with dual muco-cutaneous tissue tropism. Anal canal Gamma-PV infections may be associated with sexual behavior and the host immune status. This study expands the knowledge on Gamma-PVs' tissue tropism, providing valuable data on the characteristics of HPV infection in the anal canal.

Tumor-specific and gender-specific pre-vaccination distribution of human papillomavirus types 6 and 11 in anogenital warts and laryngeal papillomas: a study on 574 tissue specimens.

Anogenital warts and laryngeal papillomas are two most important benign tumors etiologically linked with HPV. In the study, which included both the largest number of laryngeal papilloma tissue specimens (152 specimens from 152 patients) to date and the largest number of prospectively collected and histologically confirmed tissue specimens of anogenital warts obtained from both genders (422 specimens from 315 patients), HPV DNA was detected in 413/422 (97.9%) of anogenital warts and 139/152 (91.4%) of laryngeal papillomas. HPV-6 and/or HPV-11 were detected in 291/315 (92.4%) patients with anogenital warts and in 138/152 (90.8%) patients with laryngeal papillomas, indicating that the great majority of both tumors could be prevented with prophylactic quadrivalent vaccine. The HPV-6 gender-specific distribution in both anogenital warts and laryngeal papillomas was not statistically significant. In contrast, HPV-11 was found almost three times more often in males than in females with anogenital warts (16.5% vs. 6.3%; P = 0.008), with a gender neutral HPV-11 type distribution in laryngeal papillomas. The overall HPV DNA prevalence in anogenital warts was significantly different from that in laryngeal papillomas (97.1% vs. 91.4%; P = 0.01). In the first comparison of the HPV-6/HPV-11 type-specific distribution between patients suffering from anogenital warts and laryngeal papillomas with the same geographic and ethnic background, a significant imbalance in tumor-specific distribution of HPV-6 and HPV-11 was identified: HPV-6 was statistically more often present in anogenital warts than in laryngeal papillomas (79.0% vs. 59.2%; P = 0.000013), whereas HPV-11 was statistically more frequent in laryngeal papillomas than in anogenital warts (28.9% vs. 12.4%; P = 0.00003).

Usefulness of high-risk human papillomavirus mRNA silver in situ hybridization diagnostic assay in oropharyngeal squamous cell carcinomas.

AIMS: The transcriptional activity of high-risk human papillomaviruses (HR-HPV) within oropharyngeal squamous cell carcinomas (OPSCC) has been linked to improved survival of patients. HR-HPV mRNA silver in situ hybridization (SISH) was evaluated on a cohort of OPSCC and compared with viral HPV DNA tests and p16 expression. Clinical outcomes of HPV-driven OPSCC and non-HPV related OPSCC were also studied. METHODS: We evaluated 67 OPSCC and 3 papillomas, obtained from 62 patients, for detection of HR-HPV DNA by PCR tests. The positive samples were additionally studied by the SISH method using three probes of HPV16, HPV18, and HP33, and for p16 expression detected by immunohistochemistry. SISH assays were evaluated for the presence/number and intensity of signals in cancer cells. Prognostic significance of HPV status in our cohort was evaluated with univariate and multivariate statistics. RESULTS: According to the HR-HPV PCR tests, 46 (69%) OPSCC cases were HPV positive, while three papillomas were negative. Of total 46 HPV-positive OPSCCs, 43 cases were also SISH-positive, while p16 overexpression was found in 45 of 46 HPV positive OPSCC cases. In OPSCC specimens, the sensitivity and specificity of the combined SISH probes (HPV16 and 33) were both 100.00%, when compared to HPV PCR. HPV positivity of the tumors appeared significant for predicting progression-free survival, cause specific survival and overall survival in a multivariate setting. CONCLUSIONS: The recently developed mRNA SISH methodology can detect HPV-driven OPSCCs without any additional test in 79% of cases. Positive SISH signals enable the visualization of viral transcripts required to recognize clinically relevant HPV infection. However, rare and tiny signals require an experienced pathologist to establish a consensus interpretation of results. The currently applied HR-HPV mRNA SISH analysis may serve as a groundwork for additional studies.

GeneXpert enterovirus assay: one-year experience in a routine laboratory setting and evaluation on three proficiency panels.

A prospective unblinded comparative evaluation of three assays for the detection of enteroviral RNA performed on 83 positive and 79 negative cerebrospinal fluid samples showed initial and resolved sensitivities of 90.4% and 98.8%, respectively, for the Cepheid GeneXpert enterovirus assay; 94.0% and 97.6%, respectively, for the Argene enterovirus consensus kit; and 100% and 100%, respectively, for an in-house real-time PCR. The initial and resolved specificities were 100% for all three assays.

Distribution of HPV genotypes in Slovenian patients with anal carcinoma: preliminary results.

The aim of the present study was to obtain first data on the distribution of human papillomavirus (HPV) genotypes in patients with anal cancer (AC) in Slovenia. A total of 21 samples of AC (16 archival FFPE samples and 5 fresh-frozen tissue samples) collected from the same number of patients were analysed. All samples were tested for the presence of HPV DNA using a consensus GP5+/ GP6+ PCR and HPV genotypes determined by the INNO LiPA HPV Genotyping Extra test, capable of recognizing 28 different alfa-HPV genotypes. All 21 AC samples were HPV DNA positive. The most frequent HPV genotype, found in 19/21 AC samples, was HPV-16. Only low-risk HPV-6 was detected in one sample and infection with high-risk HPV-52 and low-risk HPV-61 was identified in one sample. Prophylactic HPV vaccination with currently available vaccines could potentially prevent the great majority of anal cancers in Slovenia.

Genomic distribution of beta papillomaviruses in single eyebrow hair samples and pools of eyebrow hair samples.

Human beta papillomaviruses (beta-HPVs) are frequently detected in hairs and the majority of people are infected with multiple beta-HPV genotypes. This study was conducted to investigate for the first time the distribution of beta-HPV genotypes in single hair specimens and to estimate the contribution of a single hair to the beta-HPV profile obtained from a specimen made of multiple hairs pooled together. A total of 85 eyebrow hair specimens, representing 64 single hairs and 21 pools of hairs, obtained from 21 immunocompetent individuals, were tested using a reverse-line blot-based beta-HPV genotyping assay that allows identification of 25 different beta-HPVs. Overall, beta-HPV DNA was detected in 82/84 (97.6%) samples. The great majority of hair pools (19/21; 90.5%) contained multiple beta-HPVs, the mean number of identified beta-HPV genotypes per hair pool was 5.2 (ranging from 1 to 12). In individual hairs, the great majority of individual hairs (43/63; 68.3%) contained multiple beta-HPVs, the mean number of identified beta-HPV genotypes was 4 (ranging from 1 to 12). Overall, HPV-23 was the most prevalent genotype, followed by HPV-24 and HPV-38. A comparison of beta-HPV genotype distribution in pooled hair specimens and in at least one individual hair within a single patient revealed that 5/20 patients had a complete match between the number and profile of identified genotypes, 2/20 patients had the same/similar number of HPV genotypes but different genotype profile, 9/20 patients had more HPV genotypes identified in pools than in the majority of individual hairs and 4/20 patients had at least one individual hair with more HPV genotypes identified than in the corresponding pool. Our results suggest that beta-HPVs are unevenly distributed over the eyebrows and even pools made of several hairs do not necessarily provide information on the whole spectrum of HPV genotypes present in eyebrows.