Human Papillomavirus Detection Methods Research Articles

Humane Papillomaviren bei Kopf-Hals-Karzinomen

Human papilloma viruses (HPV) may cause head and neck squamous cell carcinomas. HPV associated carcinomas mostly occur in the oropharynx and particularly in the palatine tonsil and base of tongue. Carcinomas at other sites are infrequently caused by HPV. The viral proteins E6 and E7 are crucial for malignant transformation. They functionally interfere with cellular proteins maintaining genomic integrity such as p53 and cell cycle control such as retinoblastoma protein. However, unlike carcinogens such as tobacco smoke, the virus does not disrupt the coding genes. Reliable HPV detection methods in histological specimens include in situ hybridisation of high risk DNA and immunohistochemical detection of p16. HPV positivity does not proof a causal relation between HPV and tumour. High risk HPV are also detectable in 10-20% of normal mucosal specimens. Irrespective of the various modalities available for treatment, HPV associated head and neck cancers have a better prognosis than their carcinogen caused counterparts. HPV positivity does not object to surgical treatment.

P215 Evaluation of extraction and genotyping methods for human papillomavirus detection and analysis from liquid-based cytology specimens

To analyse the genetic variability of EG95 sequences and provide guidance for EG95 vaccine application against Echinococcus granulosus (E. granulosus).We analysed EG95 polymorphism by collecting total 97 different E. granulosus isolates from 12 different host species that originated from 10 different countries. Multiple sequence alignments and the homology were performed by Lasergene 1 (DNASTAR Inc., Madison, WI), and the phylogenetic analysis was performed by using MEGA5.1 (CEMI, Tempe, AZ, USA). In addition, linear and conformational epitopes were analysed, including secondary structure, NXT/S glycosylation, fibronectin type III (FnIII) domain and glycosylphosphatidylinositol anchor signal (GPI-anchor). The secondary structure was predicted by PSIPRED method.Our results indicated that most isolates overall shared 72.6–100% identity in EG95 gene sequence with the published standard EG95 sequence, X90928. However, EG95 gene indeed has polymorphism in different isolates. Phylogenetic analysis showed that different isolates could be divided into three subgroups. Subgroup 1 contained 87 isolates while Subgroup 2 and Subgroup 3 consisted of 3 and 7 isolates, respectively. Four sequences cloned from oncosphere shared a high identity with the parental sequence of the current vaccine, X90928, and they belonged to Subgroup 1. However, in comparison to X90928, several amino acid mutations occurred in most isolates besides oncosphere, which potentially altered the immunodominant linear epitopes, glycosylation sites and secondary structures in EG95 genes. All these variations might change their previous antigenicity and thereby affecting the efficacy of current EG95 vaccine.This study reveals the genetic variability of EG95 sequences in different E. granulosus isolates, and proposed that more vaccination trials would be needed to test the effectiveness of current EG95 vaccine against distinct isolates in different countries.

Detection of human papilloma viruses type 16 and 18 by PCR and RFLP in paraffin-embedded cervical cancer tissue specimens

Human papillomaviruses (HPVs) infect mucosal and cutaneous epithelia and cause malignant and benign neoplastic lesions. Human papillomaviruses cause 250000 deaths per year from cervical cancer most often in developing countries. This major public health problem make them important targets in the search for Human papillomaviruses detection methods that may play a role in cervical cancer screening. In the present study, we used a combination of polymerase chain reaction (PCR) amplification method and Restriction fragment length polymorphism (RFLP) analysis in order to identify HPV types in cervical cancer samples. Two of the five samples were HPV positive that one of them was HPV16 positive and the other was positive for HPV18. The method could facilitate the sensitive identification of a broad spectrum of genital Human papillomaviruses.

Presence of high-risk human papillomavirus sequences in breast cancer tissues and association with histopathological characteristics

Objective: To examine the presence of human papillomavirus (HPV) in breast cancer tissues. Design and methods: Four different PCR methods for detection and verification of genital HPVs were applied in frozen breast cancer specimens. Tumors were also evaluated for various histopathological parameters. Results: Seventeen samples out of 107 tested positive (15.9%). HPV RFLP typing identified a total of 21 high-risk viruses: fourteen HPV 16 (67% of all detected HPV types), three HPV 59, two HPV 58, one HPV 73 and one HPV 82 (one sample with double infection and two samples with triple infection). Breast cancer patients harboring high-risk HPV DNA sequences in their tumor were younger than the rest of the patients. Furthermore, they were less estrogen-receptor-positive and more proliferative as observed in the corresponding indices: Ki-67 staining, S-phase/proliferative fractions and percentage of cells with DNA content over 5C. Conclusion: The presence of high-risk HPV DNA sequences in the breast cancer tissues studied was verified, and a possible association with acceleration of malignancy was examined.

Resolving repeatedly borderline results of Hybrid Capture 2 HPV DNA Test using polymerase chain reaction and genotyping

The Hybrid Capture 2 HPV DNA Test (hc2) (Digene Corporation, Gaithersburg, MD) is at present the only FDA approved assay for routine detection of human papillomavirus (HPV) infections. A significant analytical inaccuracy of the hc2 near to cut-off was reported recently. To address this problem, 240 samples with repeatedly borderline/equivocal/indeterminate hc2 results (samples with repeated RLU/CO values between 0.4 and 4.0) were tested using the PGMY09/PGMY11 consensus PCR and genotyping in order to resolve their high-risk HPV status. All PGMY09/PGMY11 PCR negative samples were tested in addition using CPI/IIg consensus PCR. A false negative rate of 11.3% and false positive rate of 19.1% were recorded in the samples with repeatedly borderline hc2 results. The corresponding hc2 false reactivity rates in 95 samples selected at random which were clearly hc2 negative (samples with RLU/CO values less than 0.4) and 124 samples selected at random which were clearly hc2 positive (samples with RLU/CO values more than 4.0) were 4.2% and 5.6%, respectively. The proportion of hc2 false reactivity increased with proximity to the hc2 cut-off value. According to the results of the present study, the introduction of an hc2 grey-zone and retesting of samples with repeatedly borderline hc2 results by an alternate HPV detection method, such as the PGMY09/PGMY11 consensus PCR and genotyping, is recommended.

Comparison between the Hybrid Capture II Test and a PCR-Based Human Papillomavirus Detection Method for Diagnosis and Posttreatment Follow-Up of Cervical Intraepithelial Neoplasia

Human papillomavirus (HPV) infection is the major cause of cervical cancer and its precursor, cervical intraepithelial neoplasia (CIN), and HPV testing has therefore been proposed for improved triaging and follow-up of women treated for CIN. We compared two common HPV DNA detection tests (Hybrid Capture II [HCII] and PCR-enzyme immunosorbent assay (EIA) using the primers GP5+/GP6+ followed by HPV typing with reverse dot blot hybridization) for sensitivity and specificity for detection of CIN and of CIN recurrence after treatment. Two hundred and thirty-nine women referred to the Department of Obstetrics and Gynaecology in Västerås, Sweden, were enrolled because of atypical Pap smears; 177 of these were later treated for dysplasia by conization or loop diathermy. Samples for HPV DNA testing were taken before and 4 to 6 months after treatment. There was substantial agreement between the HCII and PCR-EIA (kappa, 0.70 before treatment and 0.72 after treatment). The sensitivity for histopathologically confirmed CIN III was 100.0% for PCR-EIA and 95.6% for HCII. For patients with CIN II or worse (CIN II+), the sensitivities were 92.9% (PCR-EIA) and 91.8% (HCII). The specificities for CIN II+ in the pretreatment setting were 30.4% for PCR-EIA and 24.1% for HCII. After treatment, the sensitivities for CIN III in cytology were 100.0% by both methods, and for CIN II+, sensitivities were 80.0% by both methods. The specificities for CIN II+ in the posttreatment setting were 83.5% for PCR and 85.4% for HCII. In conclusion, the sensitivities of both PCR-EIA and HCII are high and almost equal, suggesting that both methods are suitable as tools for detection and posttreatment follow-up of CIN II-III.

Comparison of Two Commercial Assays for Detection of Human Papillomavirus (HPV) in Cervical Scrape Specimens: Validation of the Roche AMPLICOR HPV Test as a Means To Screen for HPV Genotypes Associated with a Higher Risk of Cervical Disorders

Certain high-risk (HR) human papillomavirus (HPV) types are a necessary cause for the development of cervical disorders. Women with persistent HR HPV infections have an increased risk of developing high-grade cervical lesions, compared with those who have no or low-risk HPV infections. Therefore, implementation of HPV detection into cervical screening programs might identify women at risk of cervical cancer. Several HPV detection methods with different sensitivities and specificities are available. Recently, a new PCR-based technique, the Roche AMPLICOR HPV Test, was developed. This test recognizes a group of 13 HR HPV types simultaneously. This study was undertaken to validate and compare HPV detection in 573 cervical scrape specimens by the AMPLICOR HPV Test and the INNO-LiPA HPV detection/genotyping assay (SPF10-LiPA system version 1). Human beta-globin was not detected in nine specimens, which were therefore excluded from the comparison. Eleven scrape specimens containing HPV type 53 or 66 were also excluded from the comparison because these (probably) HR HPV types cannot be detected by the AMPLICOR HPV Test. The results of HPV detection by the Roche AMPLICOR HPV Test were confirmed by INNO-LiPA HPV detection/genotyping assay in 539/553 cases, showing an absolute agreement of 97.5% with a Cohen's kappa of 0.9327, indicating almost complete similarity of the two tests. Like the INNO-LiPA HPV detection/genotyping assay, the AMPLICOR HPV Test was sensitive, specific, feasible, and easy to handle. The value of the Roche AMPLICOR HPV Test with a broad-spectrum HR HPV detection has to be determined in prospective clinical studies.

Modifiable Risk Factors Associated with Clearance of Type-Specific Cervical Human Papillomavirus Infections in a Cohort of University Students

Previous findings regarding risk factors for human papillomavirus (HPV) persistence, other than viral determinants, identified from prospective cohort studies have been inconsistent in part because study designs have differed with respect to differing HPV detection methods and varying lengths of follow-up time. Therefore, the objectives of this study were to continue the search for epidemiologic risk factors of persistent cervical HPV infections and determine what behaviors differed between those women with transient HPV infections and those women who cannot clear their type-specific HPV infections. Female university students (n = 621) in Montreal were followed for 24 months at 6-month intervals. At each visit, a cervical cell specimen was collected. HPV DNA was detected using the MY09/MY11 PCR protocol and 27 HPV genotypes were identified by the line blot assay (Roche Molecular Systems, Inc., Alameda, CA). Proportional hazards regression was used to estimate the crude and adjusted hazard ratios of clearing a type-specific high-risk (n = 222) or low-risk (n = 105) HPV infection over time according to specific baseline and time-dependent covariates. Daily consumption of vegetables seemed to increase the rate of HPV clearance independent of type. The use of tampons was associated with a reduced rate of high-risk HPV clearance, whereas regular condom use was associated with an increased rate of low-risk HPV clearance only. Some proactive measures can be taken to increase the rate of HPV clearance, and there may be some differences between the sets of predictors of low-risk and high-risk HPV clearance.

DNA- versus RNA-based methods for human papillomavirus detection in cervical neoplasia

Objective. To compare DNA-based and mRNA-based methods for detection of high-grade cervical neoplasia in Norway. Methods. HPV prevalence was analyzed in 383 women with positive index cytology, selected from gynecology clinics. All patients were investigated by a new PAP smear, histology, and two commercially available HPV tests: Hybrid Capture II (Digene, Gaithersburg, MD) and the Pre Tect HPV-Proofer (NorChip AS). Cases with positive DNA test and negative mRNA test and cases with high-grade histology and negative HPV tests were retested with PCR and sequencing. We regarded the infection as latent or transient if sequencing revealed an HPV type included in both assays. Results. High-risk HPV was detected in 99.7% of the histological confirmed high-grade lesions (CIN2+) (290/291). The DNA test was positive in 95% (275/291), and the mRNA test was positive in 77% (225/291) of the histological confirmed high-grade lesions. All invasive carcinomas were mRNA positive. The DNA test was significantly more often positive in benign and low-grade lesions, some of which were found to be false positive due to cross-contamination with unrelated types. High-grade histology was detected in 83% of women with normal cytology and positive mRNA test. Latent or transient infections were detected in 11 low-grade and 12 high-grade preinvasive lesions. Sequencing revealed high-risk HPV types included only in the DNA test in 35 high-grade preinvasive lesions, HPV 52 and 58 were the most prevalent HPV types. Conclusions. These HPV tests have the potential to improve the detection rate of high-grade cervical neoplasia, with some limitations. The mRNA test seems to be more appropriate for risk-evaluation. Larger scale, population based studies are necessary to evaluate the predictive values of HPV testing in Norway.

Comparison of polymerase chain reaction and catalyzed signal amplification in situ hybridization methods for human papillomavirus detection in paraffin-embedded cervical preneoplastic and neoplastic lesions

Two molecular methods for HPV genotyping in formalin-fixed, paraffin-embedded tissue were evaluated: in house polymerase chain reaction assay (PCR) with consensus and type-specific primers and a novel procedure of in situ hybridization-a catalyzed signal amplification system (CSA-ISH, Genpoint, DAKO, Glostrup, Denmark). The number of HPV positive cases and detected viral types were compared in cervical biopsies and cone specimens according to histopathological diagnosis. Primer efficiency in detecting various types of HPV by PCR method was evaluated. DNA samples (101) were used as a template to amplify with three pairs of consensus (MY09/11, GP5+/6 +, CPI/IIG) and four type-specific HPV primers (HPV-6/11, 18, 16 and 33). The according histological tissue sections were analyzed with CSA-ISH method, using commercial HPV biotinylated probes HPV-6/11, 16/18 and 31/33/51. The degree of concordance for PCR and CSA-ISH was 64.4%. In 63 of 101 samples (62.4%), HPV was detected by PCR, while only 35 (34.7%) were positive using CSA-ISH. CSA-ISH found lower percentages for all HPV types, except HPV-6/11. A lower percentage of positive results in all high-grade lesions was detected by CSA-ISH. Multiple infections were detected by PCR in only one sample and in three samples by CSA-ISH. Detection with My09/11 primers followed by Gp5+/6+ primers, in nested reaction, gave the highest number of positive results: 58 of 63 (92%). None of the samples diagnosed as condylomata planum or CIN I was positive for HPV-6/11 (low risk type), which was detected exclusively in condylomata acuminatum group. A significantly higher number of positive samples was detected with PCR than with CSA-ISH method. CSA-ISH method should be improved, especially in detecting HPV in high-grade lesions. CSA-ISH may be more accurate in detection of multiple infections. GP5+/6+ in nested reaction after MY09/11 detected the highest number of positive results. Samples diagnosed as benign lesions positive on HPV-X must be monitored as possible candidates for progression. CIN I lesions, which were HPV negative, probably will not progress. This finding may be important in planning therapy and avoiding unnecessary treatment.

Detection and typing of human papillomavirus by e6 nested multiplex PCR.

A nested multiplex PCR (NMPCR) assay that combines degenerate E6/E7 consensus primers and type-specific primers was evaluated for the detection and typing of human papillomavirus (HPV) genotypes 6/11, 16, 18, 31, 33, 35, 39, 42, 43, 44, 45, 51, 52, 56, 58, 59, 66, and 68 using HPV DNA-containing plasmids and cervical scrapes (n = 1,525). The performance of the NMPCR assay relative to that of conventional PCR with MY09-MY11 and GP5+-GP6+ primers, and nested PCR with these two primer sets (MY/GP) was evaluated in 495 cervical scrapes with corresponding histologic and cytologic findings. HPV prevalence rates determined with the NMPCR assay were 34.7% (102 of 294) in the absence of cervical intraepithelial neoplasia (CIN 0), 94.2% (113 of 120) in the presence of mild or moderate dysplasia (CIN I/II), and 97.8% (44 of 45) in the presence of severe dysplasia (CIN III). The combination of all four HPV detection methods applied in the study was taken as "gold standard": in all three morphological subgroups the NMPCR assay had significantly (P < 0.0001) higher sensitivities than the MY09-MY11 and GP5+-GP6+ assays and sensitivities comparable or equal to those of the MY/GP assay. All 18 HPV genotypes investigated were detected among the clinical samples. The ratio of high- to low-risk HPV genotypes increased from 4:1 (80 of 103) in CIN 0 to 19:1 (149 of 157) in CIN I to III. Multiple infections were detected in 47.9% (124 of 259) of the patients. In conclusion, the novel NMPCR method is a sensitive and useful tool for HPV DNA detection, especially when exact HPV genotyping and the identification of multiple HPV infections are required.

Epidemiological and clinical aspects of human papillomavirus detection in the prevention of cervical cancer.

Cervical cancer is a major cause of death, and the second most frequent cancer in women worldwide. Many studies have indicated a causal relation between genital human papillomavirus (HPV) infections and cervical cancer. High-risk HPV genotypes have been detected in almost 100% of all cervical cancers, and the process of HPV mediated carcinogenesis has been partly clarified. The concept that HPV is a obligatory cause in cervical cancer development has lead to extensive research regarding the incorporation of HPV detection in (1) cervical cancer prevention programs, (2) triage of women with abnormal cervical scrapes in the screening program, and (3) follow-up of patients treated for cervical intraepithelial neoplasia (CIN). Epidemiological studies indicate that 50% of women becoming sexually active contract a genital HPV infection within 2 years. The lifetime risk of a genital HPV infection is estimated to be 80%, but very few of these women will develop cervical cancer. Several studies have shown that the persistence of genital HPV infections is especially related to the development of cervical cancer. As a result, HPV detection at a single moment was shown to be of limited clinical value in the triage of patients with abnormal cervical scrapes, or during follow-up after treatment for CIN. Furthermore, major variations in the sensitivity of different HPV detection methods, as well as differences in HPV detection related to the menstrual cycle, have shown that many facts regarding HPV transmission, replication, and detection need to be clarified, before HPV detection can be of clinical value.

A subset of head and neck squamous cell carcinomas exhibits integration of HPV 16/18 DNA and overexpression of p16INK4A and p53 in the absence of mutations in p53 exons 5-8.

Besides well-known risk factors such as tobacco use and alcohol consumption, oncogenic human papillomavirus (HPV) infection also has recently been suggested to promote head and neck tumorigenesis. HPV is known to cause cancer by inactivation of cell cycle regulators p53 and pRb via expression of viral oncoproteins E6 and E7. This indicates that p53 mutations are not a prerequisite in HPV-induced tumor development. However, discrepancy exists with respect to the frequency of head and neck squamous cell carcinomas (HNSCC) harboring DNA of oncogenic HPV and the fraction of these tumors showing p53 mutations. In our study, we examined the frequency of HNSCC demonstrating HPV 16/18 integration as identified by fluorescence in situ hybridization (FISH) and investigated their p53 (mutation) status by immunohistochemistry and single-strand conformation polymorphism (SSCP) analysis of exons 5-8. Paraffin-embedded, archival biopsy material from 27 premalignant mucosal lesions and 47 cases of HNSCC were analyzed. Ten of the 47 (21%) HNSCC unequivocally exhibited HPV 16 integration, including 8 of 12 (67%) tonsillar carcinomas. This is supported by the immunohistochemical detection of p16(INK4A) overexpression in all 10 HPV-positive tumors. Although FISH is considered to be less sensitive than PCR-based methods for HPV detection, our data clearly demonstrate clonal association of HPV with these tumors, as illustrated by the presence of integrated HPV 16 in both the primary tumor and their metastases in 2 patients. In contrast, HPV 16/18 DNA could not be detected in the premalignant lesions. In 30 of 47 (64%), HNSCC accumulation of p53 was observed, including 8 of the 10 HPV-positive carcinomas. However, in none of the latter cases could mutations in exons 5-8 be identified, except for a polymorphism in codon 213 of exon 6 in one patient. Evaluation of clinical data revealed a significant inverse relation between tobacco use with or without alcohol consumption, and HPV positivity of the tumors.

Screening for cervical cancer: new alternatives and research.

Evidence for the clinical utility of human papillomavirus (HPV) DNA testing has increased over the years and has now become very convincing. Some specific uses of HPV detection are a) triage of women with cytological determinations of atypical squamous cells of undetermined significance (ASC-US) and related management strategies, b) as a marker for test of cure post-treatment, and c) most importantly, as an adjunct to cytology in routine cervical disease screening programs. There are many studies that support each of these applications and include 8 studies on ASC-US triage, 10 on test of cure and 13 on adjunctive or stand-alone HPV screening. The most notable investigation of ASC-US triage was ALTS, a randomized controlled trial of 3,488 women. With respect to routine HPV screening the combined studies included 77,000 women, providing as a histological endpoint more than 1,000 cases of high-grade cervical intraepithelial neoplasia (CIN) or cancer. Testing methods were either the Hybrid Capture 2 (HC2) test or the polymerase chain reaction (PCR) test. HPV testing of women with ASC-US cytology had on average a higher sensitivity (90%) and specificity (70%) than repeating the cytological test (sensitivity 75%, specificity 60%) and was also more sensitive than colposcopy for follow-up. As an adjunct to the Papanicolaou (Pap) cytology test in routine screening, HPV DNA testing was a more sensitive indicator for prevalent high-grade CIN than either conventional or liquid cytology. A combination of HPV DNA and Papanicolaou testing had almost 100% sensitivity and negative predictive value. The specificity of the combined tests was slightly lower than the specificity of the Papanicolaou test. One "double-negative" HPV DNA and Papanicolaou test indicated a higher prognostic assurance against risk of future CIN 3 than three subsequent negative conventional Papanicolaou tests and may safely allow three-year or longer screening intervals for such low-risk women. It appears that HPV DNA testing is on the way to becoming a common testing strategy in cervical cancer prevention programs. Research continues into approaches for improving the performance and cost-effectiveness of HPV detection methods. Hybrid Capture 3 will offer improved HPV typing capabilities and the Rapid Capture machine allows for robot-assisted HPV DNA testing, permitting greater test throughput. PCR test improvements are expected to contribute to the growth of flexible accurate and cost-effective HPV DNA tests. It is likely that improved diagnostic technology along with HPV genotyping and quantitation may provide more value in future. A particularly promising approach is to combine HPV DNA testing with expression levels of other markers such as proliferative or cell cycle regulatory proteins to subdivide HPV-positive women into those who are at greater risk of cancer and those who can be safely followed by screening at longer intervals. This paper is available too at: http://www.insp.mx/salud/index.html.

Randomized clinical trial of PCR–determined human papillomavirus detection methods: Self-sampling versus clinician-directed–Biologic concordance and women's preferences

OBJECTIVE: The purpose of this study was to compare the high-risk human papillomavirus detection rates from self-sampled swabs and tampons with standard clinician-directed speculum sampling and to assess women's acceptance of self-sampling methods. STUDY DESIGN: One hundred three women who required a colposcopy underwent order randomization of the human papillomavirus sampling technique. Kappa and McNemar test statistical results were used to measure the agreement between clinician-directed and self-sampling techniques for high-risk types of human papillomavirus and the acceptance of self-sampling techniques. RESULTS: All self-directed samplings were equivalent to clinician sampling for all cervical intraepithelial neoplasia disease states. High-risk human papillomavirus was detected by self- and clinician-directed methods in 83% of the women with cervical intraepithelial neoplasia, grade 2/3. The 2 sequential swabs trend toward better detection of high-risk types of human papillomavirus than all other techniques for women with normal histologic factors (P =.0736, by McNemar's χ2 test). Ninety-four percent of women would accept self-sampling for their yearly cervical screen. CONCLUSION: Self-sampling is equivalent to clinician sampling for the detection of high-risk human papillomavirus and is acceptable to women as a yearly screen. (Am J Obstet Gynecol 2002;186:365-73.)

Human Papillomavirus Involvement in Esophageal Carcinogenesis in the High-Incidence Area of China: A Study of 700 Cases by Screening and Type-Specific in Situ Hybridization

Background: Human papillomavirus (HPV) DNA has been identified in esophageal precancerous lesions and carcinomas. However, there are marked variations in the prevalence of HPV infection reported in different studies. Most previous studies on HPV and esophageal carcinomas have been based on a limited number of biopsy samples studied by different HPV detection methods with highly variable sensitivity and specificy, making systematic studies of larger series clearly warranted. Methods: A series of 1876 surgical specimens (primary tumor, adjacent epithelium, regional lymph nodes, resection margins) from 700 patients surgically resected for an invasive squamous cell carcinoma of the esophagus in the high-incidence area of China was analyzed for the presence of HPV DNA with screening in situ hybridization (ISH) using biotinylated HPV DNA probes and followed by type-specific ISH for HPV 6, 11, 16, 18, 30, and 53. Results: Of the 700 esophageal carcinomas, 118 (16.9%) were shown to contain HPV DNA sequences by screening ISH. Positive signals were most frequent in the cancer cells (16.6%), more rare in the surrounding hyperplastic and dysplastic epithelia (5.6%), and infrequently present in the resection margins (0.2%). HPV signals were also detected in cancer cells in 6.9% of the lymph node metastases. HPV types 6, 11, 16, 18, and 30 account for 39.8% of the HPV-positive lesions, of which the high-risk types HPV 16 and 18 were present in 27.1% (32 of 118). Notably, 60.2% of the HPV-positive lesions contained DNA sequences other than HPV types 6, 11, 16, 18, 30, and 53. Conclusions: This study reports the largest series of esophageal cancers ever analyzed for the presence of HPV DNA. Our results confirm the presence of common mucosal HPV types in esophageal carcinomas but also suggest the involvement of other (novel?) HPV types that are unusually detected in genital cancers in a significant proportion of these lesions. The results further indicate that HVP has an etiologic role in esophageal carcinogenesis, at least in the high-incidence area of northern China.

High prevalence of human papillomavirus type 58 in Chinese women with cervical cancer and precancerous lesions

The prevalence of human papillomavirus (HPV) among 332 Hong Kong Chinese women with abnormal Papanicolaou smears were determined by polymerase chain reaction and restriction fragment length polymorphism analysis. The overall HPV positive rate was 44.3% with 18.6% (16/86) for normal/inflamed cervices, 36.4% (32/88) for condyloma, 64.7% (33/51) for cervical intraepithelial neoplasia grade 1 (CIN 1), 37.9% (11/29) for CIN 2, 68.3 (41/60) for CIN 3, and 77.8% (14/18) for carcinoma. Double HPV infection was detected in 17 of the 147 positive samples, with a significantly higher proportion in patients with normal or inflamed cervices than those with CIN or carcinoma (31.3% vs 10.5%, P = .029). The six most commonly identified genotypes were HPV 16 (33.3%), HPV 58 (23.8%), HPV 11, 18, 31 (8.8% each), and HPV 33 (6.8%). The worldwide uncommon genotype HPV 58 was found to be the second most common genotype detected in patients with cervical carcinoma (6 of 18 patients). HPV 58 infection showed a significant association with CIN/carcinoma (odds ratio [OR] = 3.98; 95% confidence interval [CI] = 1.22–14.35) and a significant trend of increase in prevalence with increasing severity of cervical lesion (χ2 = 5.84; P = .016). Among Hong Kong Chinese women with abnormal cervical cytology, the detection of HPV 58 carried a positive predictive value of 68.6% for a cervical lesion of CIN 1 or higher severity. The high prevalence of HPV 58 among Chinese women, particularly in patients with carcinoma, has an implication on the design of HPV detection methods and the development of vaccines. J. Med. Virol. 59:232–238, 1999. © 1999 Wiley-Liss, Inc.

High prevalence of human papillomavirus type 58 in Chinese women with cervical cancer and precancerous lesions

The prevalence of human papillomavirus (HPV) among 332 Hong Kong Chinese women with abnormal Papanicolaou smears were determined by polymerase chain reaction and restriction fragment length polymorphism analysis. The overall HPV positive rate was 44.3% with 18.6% (16/86) for normal/inflamed cervices, 36.4% (32/88) for condyloma, 64.7% (33/51) for cervical intraepithelial neoplasia grade 1 (CIN 1), 37.9% (11/29) for CIN 2, 68.3 (41/60) for CIN 3, and 77.8% (14/18) for carcinoma. Double HPV infection was detected in 17 of the 147 positive samples, with a significantly higher proportion in patients with normal or inflamed cervices than those with CIN or carcinoma (31.3% vs 10.5%, P =.029). The six most commonly identified genotypes were HPV 16 (33.3%), HPV 58 (23.8%), HPV 11, 18, 31 (8.8% each), and HPV 33 (6.8%). The worldwide uncommon genotype HPV 58 was found to be the second most common genotype detected in patients with cervical carcinoma (6 of 18 patients). HPV 58 infection showed a significant association with CIN/carcinoma (odds ratio [OR] = 3.98; 95% confidence interval [CI] = 1.22-14.35) and a significant trend of increase in prevalence with increasing severity of cervical lesion (chi(2) = 5.84; P =.016). Among Hong Kong Chinese women with abnormal cervical cytology, the detection of HPV 58 carried a positive predictive value of 68.6% for a cervical lesion of CIN 1 or higher severity. The high prevalence of HPV 58 among Chinese women, particularly in patients with carcinoma, has an implication on the design of HPV detection methods and the development of vaccines.

Prevalence of Human Papillomavirus in Squamous Cell Carcinoma of the Head and Neck Determined by Polymerase Chain Reaction and Southern Blot Hybridization: Proposal for Optimized Diagnostic Requirements

Human papillomaviruses (HPV) are discussed as cofactors in the carcinogenesis of squamous cell carcinoma of the head and neck (SCCHN). The prevalence of HPV infection in SCCHN is the subject of controversy since reported HPV prevalences range from less than 10% to almost 100%, depending mainly on the detection method employed. This study presents a realistic approximation to the real prevalence of HPV in SSCHN by applying polymerase chain reaction (PCR) and Southern blot hybridization (SBH), which are the most sensitive and specific HPV detection methods. Diagnostic procedures were optimized by applying a ''hot-start'' PCR protocol followed by a confirmatory SBH of the PCR products to reactions using both type-specific and consensus primers and probes. DNA of 75 tumour samples and 22 normal mucosa samples of the same patients were investigated. In 14 cases genomic SBH using complete HPV genomes as probes was performed additionally. HPV DNA was detected in 17/75 (22.7%) SCCHN specimens. HPV 16 was identified in four cases, HPV 45 in three cases, and HPV 6 and 18 in one case each. Hot-start PCR and SBH are the most reliable HPV detection methods, as they minimize both false-positive and false-negative results. With these methods, a HPV prevalence of 23% was achieved, which can be assumed to be representative for comparable study populations. The significant number of positives detected only by consensus primer PCR, along with the identification of HPV 45, indicate that further HPV types may play an important role in the genesis of SCCHN.

96/03638 Separation of Fischer-Tropsch catalyst/wax mixtures using dense-gas and liquid extraction

Prophylactic human papillomavirus (HPV) vaccines are recommended for prevention of HPV-associated cancers. Type-specific detection of HPV in formalin-fixed, paraffin-embedded (FFPE) tissues retrieved from diagnostic pathology laboratories is important in monitoring the impact of HPV vaccines. However, few typing assays have been validated for testing FFPE samples. Results of the Novaplex II HPV28 Detection (Novaplex) assay were compared with those from the reference assay (Linear Array with reflex Line Probe Assay) on 708 FFPE samples from cervical lesions. Novaplex showed high type-specific concordance with the reference method for HPV16/18, 9 types targeted by the Gardasil 9 vaccine, 14 high-risk types, and 21 types covered by comparison assays. The rate of inadequate samples was low in both approaches (reference, 3.4%; Novaplex, 1.7%). The proportion of discrepant types was less than 3.5% and positive concordance was greater than 75.0%. Furthermore, the type-specific positive agreement (92.0% to 98.0%), negative agreement (96.0% to 99.0%), and accuracy (97.0% to 99.0%) was high. Cohen's κ ranged from 0.86 to 0.89, indicating excellent agreement between Novaplex and reference assays. The results show that Novaplex is a suitable method for detection of HPV in FFPE tissues.