Diagnostic options for human polyomaviruses in clinical practice.

The members of the viral family Polyomavirae are widespread in the human population. According to serological studies, almost all adults are infected with at least one of this group of viruses. The primary infection usually occurs in childhood without any clinical signs, and after the primary infection, the viruses establish a persistent infection accompanied by occasional reactivation and shedding of the virus. These viruses often reactivate in immunosuppressed individuals, but only in a minority of these patients, the reactivation results in disease development. This biological property of human polyomaviruses makes laboratory diagnosis considerably difficult. The paper provides an overview of methods for diagnosing human polyomaviruses, which are commonly used for screening, and methods that are still validated by research but have the potential to improve detection and to identify patients at risk of developing diseases associated with polyomavirus infection.

Mucosal and skin HPV types in tumour-free tonsils and tonsillar tumours.

Human papillomavirus (HPV) infection is associated with tonsillar cancer (TC) whose incidence in humans is increasing. Tonsillar tumours are not ordinarily preceded by clinically apparent precancerous lesions, and no markers of the early stage disease are available. Therefore, we evaluated the presence of an active HPV infection also in tumour-free tonsillar tissue. Formalin-fixed paraffin-embedded (FFPE) tonsillar specimens from 114 patients with TC and 114 age and gender matched controls were screened for the presence of HPV DNA, expression of HR-HPV E6 mRNA, and p16 overexpression. HPV DNA was identified in 3.5% of tumour-free tissues, HR-HPV16 and 58 and LR-HPV111 and 17 were each detected in a single sample. No HR HPV E6 mRNA and p16 overexpression was found. The prevalence of HPV DNA in TC was 69.3%, with HPV16 being the most common (94.9%). Eighty-four percent of HPV16-positive tumours expressed HR HPV E6 mRNA, while no E6 mRNA was present in samples positive for HPV52 and 58. The overexpression of p16 correlated well with HPV DNA in TC, but in tumour-free tonsils no overexpression of p16 was detected.Our data provide further evidence of the etiological role of HPV16 in TC. In tumour-free tissue, the presence of HR-HPVs was rare and silent, as shown by direct and indirect markers.

High level of Tregs is a positive prognostic marker in patients with HPV-positive oral and oropharyngeal squamous cell carcinomas.

BACKGROUND: Human papillomaviruses (HPVs) have been proved as one of the etiological factors of oropharyngeal squamous cell carcinoma (OPSCC). Patients with tumors of viral etiology have a lower recurrence rate and better prognosis. OPSCC is linked to an alteration in the immune system. Only a limited number of studies have correlated both the immunological parameters and HPV status with patient prognosis. The aim of this study was to determine whether HPV infection and the immunological status influence patient prognosis individually or in concurrence. MATERIAL AND METHODS: Sixty patients with oral and oropharyngeal carcinomas were enrolled. They were divided into HPV-positive and HPV-negative groups based on the expression of HPV 16 E6 mRNA. Basic lymphocyte subpopulations were determined in the peripheral blood by means of flow cytometry. RESULTS: Significantly better disease-specific survival (DSS) was observed in patients with HPV-positive tumors. Nodal status, tumor grade, recurrence, and CD8+/Tregs ratio were identified as factors influencing DSS. A higher level of Tregs and a lower ratio of CD8/Tregs influenced overall survival (OS) independently of HPV status and age. Patients with HPV-positive tumors and high levels of Tregs survived significantly better than patients from the other groups. CONCLUSION: Better survival is associated with HPV positivity and elevated Tregs levels. Our data suggest that HPV infection and Tregs do not influence patient prognosis in concurrence.

Prevalence and genetic heterogeneity of SEN virus genotypes D and H in blood donors from Central and Western Europe and West Africa.

OBJECTIVES: To establish prevalence and phylogenetic relationship of SEN virus (SENV) D and H in blood donors from Scotland, Czech Republic and Ghana. AIM: To compare the data between three regions with differing prevalence of blood-borne viruses. BACKGROUND: Anelloviruses are a ubiquitous group of viruses without a clear disease association. Although there is little evidence that they are pathogenic per se, they may have the ability to modify ongoing disease processes. They have a high degree of heterogeneity both within populations and across geographic regions. MATERIALS AND METHODS: Three sets of donor samples were analysed by nested polymerase chain reaction (PCR) and hybridisation. A proportion of amplified samples were sequenced and phylogenetic analysis was carried out. RESULTS: The prevalence figures (including mixed D + H infection) were established for SENV D: 1·0, 8·4 and 25·2% and H: 12·5, 34·8 and 61·0% in Scottish, Czech and Ghanaian blood donors, respectively. The compilation of prevalence figures indicates the changing ratio of SENV D/H in west-east direction, most obvious between Western Europe (D/H < 1) and far East Asia (D/H > 1). Phylogenetic analysis grouped the samples mostly in accordance with geographic origin, despite the variability of short sequence analysed. The previously indicated link between SENV prevalence and age was statistically significant in this study, only for SENV H in Czech samples. CONCLUSION: SENV D and H appear to reflect the incidence of other blood-borne viruses in these locations. SENV H prevalence of 45·4% in Ghana represents the highest single-SENV-genotype prevalence described in blood donors to date.

Human papillomavirus infection and tumours of the anal canal: correlation of histology, PCR detection in paraffin sections and serology.

Human papillomavirus infection is an important etiological factor in squamous cell carcinoma of the anus (SCCA). Different histological variants of anal carcinomas displaying squamous differentiation, previously classified as separate tumours, were recently reclassified as SCCA by the WHO. In our recent study the presence of HPV was detected by PCR in biopsy specimens of 42 different anal tumours, including SCCA and its histological variants (n=22), adenocarcinomas (n=5), tubulovillous adenomas (n=5) and anal condylomas (n=10). HR HPV16 (high risk - HR) was detected in 18 of SCCA specimens (81.8%). All histological variants, i.e. tumours with basaloid, squamous and mixed histological patterns, were represented among the HPV-positive cancers. Four tumours (18.2%) were HPV negative. Low-risk (LR) HPV types were not detected within the SCCA group. HPV16 was identified in one adenocarcinoma, while four cases were HPV negative. Two adenomas showed presence of HPV16; one showed simultaneous positivity for HPV33. The remaining three tumours were HPV negative. Seven anal condylomas (70%) were LR HPV 6 and/or 11 positive, while three were HPV negative. The presence of HR HPV types was not observed in anal condylomas. Our results provide further evidence in support of the etiological role of HR HPV infection in the development of SCCA regardless of its histological appearance.

HPV and other risk factors of oral cavity/oropharyngeal cancer in the Czech Republic.

OBJECTIVE: An association between high-risk human papillomavirus (HR HPV) infection and a risk of development of a subgroup of head and neck cancers has been proposed recently. The main risk factors of oral and oropharyngal cancer observed in our population are smoking and alcohol consumption. The incidence of oral/oropharyngeal tumours in the Czech Republic is relatively high and there are no data available about the prevalence of HPV DNA presence in these tumours. MATERIALS AND METHODS: Eighty patients with a primary oropharyngeal cancer were enrolled. The presence of HPV DNA has been evaluated by polymerase chain reaction in 68 cases from which the tumour tissue and demographical and clinical data were available. The typing of HPV was performed by nucleotide DNA sequencing. RESULTS: The HPV DNA was detected in 51.5% of samples tested. Among the HPV DNA positive tumours, 80% contained HPV16. In the analysed group there were 54 men and 14 women. The prevalence of HPV DNA was lower in oral (25%) than in oropharyngeal (57%) tumours, and higher in never smokers (100%) and never drinkers (68.8%). HPV DNA presence was not related to gender, age, number of lifetime sexual partners or practice of oral-genital sex, size of tumour or presence of regional metastases. CONCLUSIONS: The difference in the prevalence of HPV DNA positive tumours between cases of oral cavity and oropharyngeal carcinoma exposed and not exposed to tobacco or alcohol support the theory that HPV DNA positive tumours form an aetiologically distinct subgroup of head and neck tumours.

Prospective study on cervical neoplasia: presence of HPV DNA in cytological smears precedes the development of cervical neoplastic lesions.

OBJECTIVES: The principal aim of the study was to verify whether HPV infection in healthy women, as determined by HPV DNA detection, was associated with an increased risk of development of cervical lesions. METHODS: Cervical smears collected at enrolment into the prospective study conducted in Prague during 1975-83 were tested for the presence of HPV DNA by means of a polymerase chain reaction (PCR) using the general GP5/6 primers and a mixture of primers specific for the E6 gene. 120 smears from patients in whom cervical neoplasia had been detected in the course of the prospective study and 208 smears from control women who had remained healthy throughout the observation period were analysed. Patients and controls were matched by age, number of sexual partners, age at first intercourse, and smoking habit. Patients were divided into three groups, A, B, and C, according to their cytological, colposcopic, and histological findings at enrolment. Group A consisted of 67 women found ill at enrolment, group B of 26 women with slightly suspicious findings, while group C comprised 27 women with normal findings at enrolment. In addition, sera taken at enrolment from these patients and controls were tested for the presence of antibodies reactive with virus-like particles (VLPs) of HPV 16, 18, and 33. RESULTS: For the whole cohort, there was a statistically highly significant difference in the presence of HPV DNA between patients and controls. Furthermore, the difference in the presence of HPV DNA between patients and controls was highly significant not only in those who had been found ill at enrolment (group A) but, most importantly, also in women who had developed the disease in the course of the follow up (groups B and C). Women positive for HPV DNA possessed HPV antibodies to VLP16, 18 and 33 significantly more often than those who were free of HPV DNA. CONCLUSION: This indicated that healthy women who were positive for HPV DNA at enrolment were at an increased risk of developing cervical neoplasia (OR = 18.5; CI 5.9 to 57.6).

Correlation between human papillomavirus-associated cervical cancer and p53 codon 72 arginine/proline polymorphism.

High-risk mucosal human papillomaviruses encode an E6 oncoprotein, which binds the cellular p53 tumor suppressor protein, thereby marking it for degradation through the ubiquitin-mediated pathway. A common p53 polymorphism at codon-72 of exon 4 results in translation to either arginine or proline. Recently reported data suggested an increased susceptibility to E6/ubiquitin-mediated degradation of the Arg72-p53 isoform and an over-representation of the homozygous Arg72-p53 genotype in cervical cancer patients. We have analyzed this polymorphism in a larger series of patients with cervical cancer and in controls in the Czech Republic. We found no statistically significant differences between the codon-72 p53 genotypes of cervical cancer patients and the control women. Based on these results, it is unlikely that Arg72-p53 is associated with an increased risk for human papillomavirus-associated cervical tumor development in Czech women.