Detection Of HPV Genotypes Research Articles

A single dose of quadrivalent HPV vaccine is highly effective against HPV genotypes 16 and 18 detection in young pregnant women eight years following vaccination: an retrospective cohort study in Fiji

In 2008/9, Fiji vaccinated >30,000 girls aged 9-12 years with the quadrivalent human papillomavirus (4vHPV) vaccine coverage for at least one dose was >60% (one dose only was 14%, two dose only was 13%, three doses was 35%). We calculated vaccine effectiveness (VE) of one, two and three doses of 4vHPV against oncogenic HPV genotypes 16/18, eight years following vaccination. A retrospective cohort study was undertaken (2015-2019) in pregnant women ≤23years old, eligible to receive 4vHPV in 2008/9, with confirmed vaccination status. The study was restricted to pregnant women due to the cultural sensitivity of asking about sexual behavior in Fiji. For each participant a clinician collected a questionnaire, vaginal swab and genital warts examination, a median eight (range 6-11) years post vaccination. HPV DNA was detected by molecular methods. Adjusted VE (aVE) against the detection of vaccine HPV genotypes (16/18), the comparison group of non-vaccine genotypes (31/33/35/39/45/51/52/56/58/59/66/68), and genital warts were calculated. Covariates included in the adjusted model were: age, ethnicity and smoking, according to univariate association with any HPV detection. Among 822 participants the prevalence of HPV 16/18 in the unvaccinated, one, two and three-dose groups were 13.3% (50/376), 2.5% (4/158), 0% (0/99) and 1.6% (3/189), respectively; and for the non-vaccine high-risk genotypes, the detection rate was similar across dosage groups (33.2%-40.4%, p=0.321). The aVE against HPV 16/18 for one, two and three doses were 81% (95% CI; 48-93%), 100% (95% CI; 100-100%), and 89% (95% CI; 64-96%), respectively. Prevalence of HPV 16/18 was lower among women with longer time since vaccination. A single dose 4vHPV vaccine is highly effective against HPV genotypes 16 and 18 eight years following vaccination. Our results provide the longest duration of protection for reduced dose 4vHPV schedule in a low- or middle-income country in the Western Pacific region. This study was supported by the Bill & Melinda Gates Foundation and the Department of Foreign Affairs and Trade of the Australian Government and Fiji Health Sector Support Program (FHSSP). FHSSP is implemented by Abt JTA on behalf of the Australian Government.

Impact of nucleic acid extraction procedures on human papillomavirus (HPV) detection and genotyping.

Human papillomavirus (HPV) infections are often asymptomatic, but some of the >200 HPV genotypes confer a high risk for precancerous cervical lesions and cervical cancer. Current clinical management of HPV infections relies on reliable nucleic acid testing detection and genotyping. We prospectively compared nucleic acid extraction without and with prior centrifugation enrichment for detecting and genotyping HPV in cervical swabs with atypical squamous or glandular cells. Consecutive swabs were analyzed from 45 patients with atypical squamous or glandular cells. Nucleic acids were extracted in parallel using three procedures, Abbott-M2000, Roche-MagNA-Pure-96 Large-Volume Kit without (Roche-MP-large) and with prior centrifugation (Roche-MP-large/spin) and tested using Seegene-Anyplex-II HPV28. In total, 54 HPV-genotypes were detected in 45 samples, 51 by Roche-MP-large/spin, 48 by Abbott-M2000 and 42 by Roche-MP-large. The overall concordance was 80% for detecting any HPV and 74% for specific HPV-genotypes. Roche-MP-large/spin and Abbott-M2000 showed the highest concordance for HPV detection (88.9%; kappa 0.78), and genotyping (88.5%). Two and more HPV-genotypes were detected in 15 samples, often with one HPV being more abundant. Dilution series confirmed the specific detection of multiple HPV-genotypes and their relative abundance. In 285 consecutive follow-up samples extracted by Roche-MP-large/spin, the top three detected genotypes were the high-risk HPV16, HPV53, HPV56 and the low-risk HPV42, HPV54 and HPV61. Rate and breadth of HPV detection in cervical swabs depends on extraction protocols being highest after centrifugation/enrichment. As multivalent HPV-vaccine coverage increases, detecting the evolving HPV virome depends on improved extraction and broader genotype coverage.

HPV genotyping by L1 amplicon sequencing of archived invasive cervical cancer samples: a pilot study

BackgroundHuman papillomavirus (HPV) is the primary cause of invasive cervical cancer (ICC). The prevalence of various HPV genotypes, ranging from oncogenically low- to high-risk, may be influenced by geographic and demographic factors, which could have critical implications for the screening and prevention of HPV infection and ICC incidence. However, many technical factors may influence the identification of high-risk genotypes associated with ICC in different populations.MethodsWe used high-throughput sequencing of a single amplicon within the HPV L1 gene to assess the influence of patient age, race/ethnicity, histological subtype, sample type, collection date, experimental factors, and computational parameters on the prevalence of HPV genotypes detected in archived DNA (n = 34), frozen tissue (n = 44), and formalin-fixed paraffin-embedded (FFPE) tissue (n = 57) samples collected in the Los Angeles metropolitan area.ResultsWe found that the percentage of off-target human reads and the concentration of DNA amplified from each sample varied by HPV genotype and by archive type. After accounting for the percentage of human reads and excluding samples with especially low levels of amplified DNA, the HPV prevalence was 95% across all ICC samples: HPV16 was the most common genotype (in 56% of all ICC samples), followed by HPV18 (in 21%). Depending upon the genotyping parameters, the prevalence of HPV58 varied up to twofold in our cohort. In archived DNA and frozen tissue samples, we detected previously established differences in HPV16 and HPV18 frequencies based on histological subtype, but we could not reproduce those findings using our FFPE samples.ConclusionsIn this pilot study, we demonstrate that sample collection, preparation, and analysis methods can influence the detection of certain HPV genotypes and must be carefully considered when drawing any biological conclusions based on HPV genotyping data from ICC samples.

Detekcija visokorizničnih HPV genotipova Real-time PCR metodom

Discovery of the causal relationship between the human papilloma virus and cervical cancer formation increased the significance of the real-time PCR in HPV diagnostics. Based on evidence showing that they caused cervical cancer, 14 HPV types have been classified as carcinogenic (HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68). This study analysed cervical smears taken from female patients, aged 19 to 25 years, using the Viasure diagnostic test for the detection of high-risk HPV genotypes and individual identification of HPV genotypes 16 and 18. A total of 110 cervical smears were analysed and 44 positive samples were detected (40%). DNA analysis of the positive samples found the following distribution of the HPV types: 27% HPV (31, 39, 56); 22% HPV (52, 59, 68); 18% HPV16; 13% HPV (33, 45, 51); 12% HPV (35, 58, 66); 8% HPV18. This study and the high positivity rate it found indicate that there is a lack of awareness among the youth on the measures of prevention, as well as a lack of understanding of HPV infection.

IL-6 597A/G (rs1800797) and 174G/C (rs1800795) Gene Polymorphisms in the Development of Cervical Cancer in Lithuanian Women.

Our study aimed to evaluate the distribution of genotypes and allele frequencies of IL-6 597A/G (rs1800797) and 174G/C (rs1800795) polymorphisms in HPV infected and uninfected healthy women and cervical cancer patients. A PCR based Multiplex HPV genotyping test kit was used for in vitro detection and differentiation of high risk HPV genotypes. Genotyping of two polymorphisms, IL-6 597A/G (rs1800797) and 174G/C (rs1800795), was performed using the KASP genotyping assay kit. Cervical cancer patients were more likely to be HPV positive than control patients. Allele C of IL-6 rs1800795 was associated with a higher risk of cervical cancer by 2.26-fold and genotype CC by 5.37-fold. Genotype CC of IL-6 rs1800795 was more frequent in the HPV positive group compared with the HPV negative group (p = 0.002). Allele G of IL-6 rs1800797 was more frequently found in women with HPV16/HPV18 compared to other HPV types (p = 0.045). Women with AA genotypes of IL-6 rs1800797 were less frequently infected with HPV16/HPV18 compared to other HPV types (p = 0.045). The major finding of the study is the significant association of C allele and CC genotype of IL-6 1800795 gene with cervical cancer in the Lithuanian population. Genotype CC of IL-6 rs1800795 has a significant association with HPV infection as well.

Flexible sensor based on conducting polymer and gold nanoparticles for electrochemical screening of HPV families in cervical specimens

Considering the low sensitivity of cytological exams and high costs of the molecular methods, the development of diagnostic tests for effective diagnosis of HPV infections is a priority. In this work, biosensor composed of polypyrrole (PPy) films and gold nanoparticles (AuNPs) was obtained for specific detection of HPV genotypes. The biosensor was developed by using flexible electrodes based on polyethylene terephthalate (PET) strips coated with indium tin oxide (ITO). Polymeric films and AuNPs were obtained by electrosynthesis. Oligonucleotides sequences modified with functional amino groups were designed to recognize HPV gene families strictly. The modified oligonucleotides were chemically immobilized on the nanostructured platform. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used for the analysis of the electrode modification and monitoring of molecular hybridization. Electrochemical changes were observed after exposure of the biosensors to plasmid samples and cervical specimens. The biosensor based on the BSH16 probe showed a linear concentration range for target HPV16 gene detection of 100 pg μL−1 to 1 fg μL−1. A limit of detection (LOD) of 0.89 pg μL−1 and limit of quantification (LOQ) of 2.70 pg μL−1 were obtained, with a regression coefficient of 0.98. Screening tests on cervical specimens were performed to evaluate the sensibility and specificity for HPV and its viral family. The expression of a biomarker for tumorigenesis (p53 gene) was also monitored. In this work, a flexible system has been successfully developed for label-free detection of HPV families and p53 gene monitoring with high specificity, selectivity, and sensitivity.

Microassay of High-Risk Human Papillomavirus Genotype Based on R6G-ddATP/SNaPshot-Gel Fluorescence Method

R6G-ddATP was used as a dideoxy fluorescence substrate to establish the single base end extension (SNaPShot)-gel fluorescence method for the rapid detection of the genotypes of three high-risk human papillomaviruses (HR-HPV) ( HPV18, HPV33 and HPV35) genotypes. HPV quality control products were used as as samples, and R6G-ddATP dideoxy fluorescence reagent was used as substrate. Firstly, HPV was amplified by using universal primers to obtain the first round of amplified products, which were purified and used as templates for subsequent SNaPShot reactions. Then, specific one-step extension primers were used to perform SNaPShot reaction to generate R6G-fluorescence-labeled DNA extension products. The product was subjected to agarose gel electrophoresis, the results of which were observed under a Gel Imager, and the HPV genotyping was done with different one-step extension primers. Each sample was tested three times and the results were compared with DNA sequencing results. The preferred annealing temperature for SNaPShot reaction is 55 ℃. Three HPV genotypes were examined by R6G-ddATP/SNaPShot gel fluorescence assay under optimal conditions, and the results were consistent with DNA sequencing results. The R6G-ddATP/SNaPShot-gel fluorescence method for the micro-detection methods of three HR-HPV genotypes was successfully established and can be used for rapid detection of HPV genotypes.

Targeted next generation sequencing panel for HPV genotyping in cervical cancer

Cervical cancer are generally caused by a persistent infection with the oncogenic virus, HPV. Patients with HPV integration are more prone to develop cervical cancer than patients without integration. In this proof-of-concept study, we aimed to develop a sensitive method based on targeted amplicon based NGS for early and precise detection of high-risk HPV-genotypes that are highly associated with the development of cervical cancer. Furthermore, we aimed to investigate if amplicon based NGS allowed for HPV genotyping in cervical lesions and whether it could detect HPV integration. The cohort included a group of CIN3+ biopsies (n = 64), CIN2 samples that progressed (n = 5), CIN2 samples that regressed (n = 3), healthy controls (n = 10), and plasma samples (n = 10) from cervical cancer patients. Sequencing was performed using a custom targeted NGS panel designed to detect all 25 high-risk and probably high-risk and two low-risk HPV genotypes. The method was validated by the SPF10 PCR-DEIA-LiPA25 assay. In the cohort, the following HPV genotypes were identified: HPV-16, 18, 31, 33, 35, 45, 51, 52, 56, 58, and 59. When comparing the results from the SPF10 PCR-DEIA-LiPA25 analyses with the NGS analyses, there was close to a perfect agreement (K = 0.92) among the genotyped HPV types, while in the two cases with complete disagreement, a third assay was applied, and here the results of the NGS analyses were confirmed. Whereas multiple HPV types were detected by the SPF10 PCR-DEIA-LiPA25 assay, the NGS analysis clearly suggest that there is one predomentant HPV type. The NGS assay was capable of detecting HPV-16 in a previous false-negative sample classified by the INNO-LiPA assay, emphasizing the importance of including multiple regions of the HPV genome when genotyping. For the 10 plasma samples, our NGS analyses showed full agreement with the digital droplet PCR (ddPCR) analyses of HPV positive as well as negative plasma samples. Lastly, the custom panel was capable of detecting the integration of HPV-16 in the SiHa cell line. The HPV panel provides a highly cost-effective method for HPV detection and genotyping, as exemplified by a list price of around 75 € per sample. In conclusion, the current study demonstrates that targeted NGS is capable of detecting and genotyping HPV in both FFPE biopsies and plasma samples. This method provides for early diagnosis and prognosis of cervical cancer disease progression, thereby optimizing the potential of recovery and survival for these patients.

The Frequency of Human Papilloma Virus in Laryngeal Squamous Cell Carcinoma.

The association of HPV in laryngeal squamous cell carcinoma (LSCC) is investigated in several studies but controversial results are established. This study aimed to investigate the HPV DNA positivity in LSCC patients diagnosed and treated in two otorhinolaryngology referral centres in northern region of Turkey. The study was planned as a retrospective investigation of LSCC patients. Fifty-two formalin-fixed, paraffin-embedded (FFPE) tissue blocks of laryngeal cancers-diagnosed and treated between 2010 and 2016, were included. Polymerase chain reaction (PCR) method was used for detection of HPV genotypes. PCR amplification was successful in 40 of 52 patients. Among the 40 LSCC samples, HPV DNA was detected in one patient (2.5%). The evaluated HPV positivity in LSCC as low; but larger studies are needed to confirm this.

Principles and analytical performance of Papilloplex® HR-HPV, a new commercial CE-IVD molecular diagnostic test for the detection of high-risk HPV genotypes

The accurate detection and genotyping of high-risk human papillomavirus (HR-HPV) are critical for cervical cancer screening and epidemiological investigations. GeneFirst Papilloplex® HR-HPV is a new CE-IVD–marked real-time PCR test based on patented multiplex probe amplification technology. Papilloplex® HR-HPV provides the simultaneous detection and differentiation of 14 HR-HPV genotypes (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68a/b) in a single closed-tube reaction ensuring rapid, cost-effective, and contamination-free results. In this study, the analytical performance characteristics in terms of the assay's sensitivity, specificity, range, reproducibility, and cross-reactivity were evaluated. Papilloplex® HR-HPV provided sensitive detection and differentiation of 14 HR-HPV types with highly reproducible results. The differential HR-HPV specificity and sensitivity were further confirmed through the participation in the WHO HPV Laboratory Network Proficiency Study (2014). Overall, GeneFirst Papilloplex® HR-HPV assay demonstrated a robust analytical performance with reproducible and reliable results in the detection of HR-HPV genotypes.

Detection of Hpv Genotypes and Their Association With p53 Codon 72 Polymorphism in Prostate Cancer

Aim: In this study, we focused on the p53 codon 72 polymorphism and HPV infections as a possible contributing factor in susceptibility to prostate cancer (PCa). Methods: Eighty histopathologically confirmed prostate biopsies comprising of 40 cancer cases and 40 benign prostate hyperplasia cases were used. Results: A significant difference was not observed between HPV presence and PCa progression. Our results showed that the p53 protein with Arg (p53-Arg72) variant contributes to the development of PCa. Conclusion: There is a near-significant association between Arg/Arg polymorphism and PCa.

Detection assay for HPV16 and HPV18 by loop‑mediated isothermal amplification with lateral flow dipstick tests.

Cervical cancer is the third highest cause of death in developing countries and most commonly results from high‑risk human papillomavirus (HR‑HPV) infection. Among HR‑HPV genotypes, HPV16 and HPV18 are the most prevalent in cervical cancers. Therefore, the present study aimed to develop a detection assay for HPV16 and HPV18 infection using loop‑mediated isothermal amplification (LAMP) with lateral flow dipstick (LFD) tests. This assay is a simplified, user‑friendly method for the visual detection of HPV genotypes. DNA was extracted from clinical tissue samples, and HPV genotyping was performed using nested polymerase chain reaction (PCR). The clinical samples were demonstrated to include 44 HPV16‑positive, 18 HPV18‑positive and 80 HPV‑negative samples. All DNA samples were also used as templates for a LAMP reaction (30min at 65˚C), and subsequently, a fluorescein isothiocyanate‑labelled probe was hybridized with the reaction product. Finally, the LFD test was performed. The sensitivity of the LAMP‑LFD test was higher than LAMP‑turbidity, exhibiting up to 100‑fold higher sensitivity for HPV16 and 10‑fold higher sensitivity for HPV18. All HPV16 and HPV18‑positive samples generated positive results in both assays; however, 22 samples detected as HPV‑negative by LAMP‑turbidity exhibited positive results by LAMP‑LFD test (22 of 80 samples). Therefore, these samples were further examined using quantitative (q)PCR. The results demonstrated that 20 out of the 22 samples designated positive by LAMP‑LFD, but negative by LAMP turbidity, gave a positive result with qPCR, while the remaining 2 samples were negative by qPCR. The present results suggested that LAMP‑LFD provided higher sensitivity than LAMP‑turbidity and nested PCR. Thus, the LAMP‑LFD test developed in the present study might be useful for the detection of HPV16 and HPV18 in local hospitals.

Efectividad del genotipado del virus del papiloma humano frente a la citología anal en la identificación de neoplasia intraepitelial de alto grado

IntroductionThe incidence of high-grade anal intraepithelial neoplasia (HGAIN) -with an aetiological based on high-risk types of human papillomavirus- is increasing in some high-risk groups. Screening for HGAIN includes routine anal cytology and, more recently, HPV genotyping. The main objective of this study was to determine the sensitivity and specificity of anal cytology and HPV genotyping for the detection of HGAIN. Materials and methodsThis is a study to determine the correlation of cytological and microbiological findings with anal biopsy findings in a cohort of patients at high risk of developing AIN referred to the department of sexually transmitted infections of the Hospital Costa del Sol, Spain, between January 2008 and December 2014. ResultsOf the 151 patients subjected to screening, a total of 92 patients, all of them with the result of three screening test (anal cytology, genotyping and biopsy) were included in the study. Just under two-thirds (62%) of them were HIV-positive. The sensitivity and specificity of anal cytology to detect HGAIN were 52.8 and 85.7%, respectively (k: 0.328), and 78 and 62.8% to detect two or more HPV oncogenic genotypes (k: 0.417). The detection of oncogenic HPV genotypes allowed the identification of 23 new cases of HGAIN that had been underdiagnosed with anal cytology, with 14 cases containing at least three high-risk genotypes. ConclusionAnal cytology did not show enough sensitivity in HGAIN screening. HPV genotyping has shown to be a useful tool to detect HGAIN cases, although it could lead to an over-diagnosis as a solitary screening procedure.

High-risk human papillomavirus genotypes in cervical lesions and vaccination challenges in China.

Cervical cancer, mostly progressing from cervical intraepithelial neoplasia, is a major cause of morbidity and mortality in Chinese women. This is largely due to high prevalence of high-risk human papillomaviruses (hr-HPVs) in the population. The prevalence of hr-HPV DNA in women and in cervical lesions women ranged from 9.9% to 17.% and from 50.5% to 70.9% in different regions of China, respectively. The most common genotypes somewhat differ between regions throughout the country and from those in many other countries. This may be a challenge to cervical cancer screening and prevention in China. Combined detection of particular HPV genotypes should be recommended in all geographical regions in China and greater attention must be paid to specific hr-HPV types during cervical cancer screening and follow-up of cervical lesions. Besides, vaccination for prevention of cervical cancer by particular HPV genotypes, has not been introduced to China so far. Updated knowledge on prevalent HPV genotypes should be provided to public health organizations to help with the development of more effective HPV vaccines, which can protect Chinese women against HPV types prevalent in local China and thus have a substantial impact on the cervical cancer burden.

Comparison of analytical and clinical performance of CLART HPV2 genotyping assay to Linear Array and Hybrid Capture 2: a split-sample study.

BackgroundHuman Papillomavirus (HPV) genotyping has an increasingly important role in cervical cancer screening and vaccination monitoring, however, without an internationally agreed standard reference assay. The test results from the most widely used genotyping assays are read manually and hence prone to inter-observer variability. The reading of test results on the CLART HPV2 genotyping assay is, on the other hand, automated. The aim of our study was to directly compare the detection of HPV genotypes and high-grade cervical intraepithelial neoplasia (CIN) by CLART, Linear Array (LA), and Hybrid Capture 2 (HC2) using samples stored in SurePath.MethodsResidual material from 401 routine samples from women with abnormal cytology was tested by CLART, LA, and HC2 (ClinicalTrial.gov: NCT01671462, Ethical Committee approval: H-2012-070). Histological outcomes were ascertained by linkage to the Danish nation-wide Pathology Data Bank. For comparison of CLART and LA in terms of genotype detection, we calculated κ-coefficients, and proportions of overall and positive agreement. For comparison of CIN detection between CLART, LA, and HC2, we calculated the relative sensitivity and specificity for high-grade CIN.ResultsThe κ-coefficient for agreement in detection of genotypes 16, 18, 31, 33, 35, and 51 was ≥0.90 (overall agreement: 98-99%, positive agreement: 84-95%). The values were slightly lower, but still in the “substantial” range for genotypes 39, 45, 52, 56, 58, 59, and several low-risk genotypes. The relative sensitivity of CLART for ≥ CIN2 and ≥ CIN3 was not significantly lower than that of LA and HC2, although CLART showed a higher specificity than HC2.ConclusionsIn Danish women with abnormal SurePath cytology, CLART and LA were highly comparable for detection of most high-risk and low-risk genotypes; and CLART’s sensitivity for high-grade CIN was comparable to that of both LA and HC2.

Significance of p16 positive expression in oropharyngeal cancers.

The involvement of HPV in oral and oropharyngeal carcinogenesis was first proposed in 2004, based on epithelial HPV tropism and detection of HPV genotypes in oral squamous cell carcinoma samples. While 60-70% of oropharynx tumors may be HPV-positive, only 10 to 19% of tumors of the oral cavity, larynx and hypopharynx appear to have HPV infection. The aim of the study was to evaluate HPV infection associated with oropharyngeal cancer. Seventy-eight cases were selected for p16 immunoexpression reactions, and demographic data were collected for comparisons. Most patients were over 60 years old, and 64.1% were smokers. Immunohistochemistry results showed that 86.3% of cases stained positive for p16 protein. The oropharyngeal cancer profile at Erasto Gaertner Hospital presented a high index of smokers over 60 years as well a high number of p16+ tumors, for what we can not determinate the main etiologic factor, but can be aware of the number of patients that presented HPV infection. Since prevention is still the best way to deal with cancer disease, it is important to analyze the interaction of these two etiologic factors and how to detect lesions at an early stage.

Correlation of EGFR, pEGFR and p16INK4 expressions and high risk HPV infection in HIV/AIDS-related squamous cell carcinoma of conjunctiva

BackgroundSquamous cell carcinoma of conjunctiva has increased tenfold in the era of HIV/AIDS. The disease pattern has also changed in Africa, affecting young persons, with peak age-specific incidence of 30-39 years, similar to that of Kaposi sarcoma, a well known HIV/AIDS defining neoplasm. In addition, the disease has assumed more aggressive clinical course. The contributing role of exposure to high risk HPV in the development of SCCC is still emerging.ObjectiveThe present study aimed to investigate if immunohistochemical expressions of EGFR, pEGFR and p16, could predict infection with high risk HPV in HIV-related SCCC.MethodsFFPE tissue blocks of fifty-eight cases diagnosed on hematoxylin and eosin with SCCC between 2005-2011, and subsequently confirmed from medical records to be HIV positive at the department of human pathology, UoN/KNH, were used for the study. Immunohistochemistry was performed to assess the expressions of p16INK4A, EGFR and pEGFR. This was followed with semi-nested PCR based detection and sequencing of HPV genotypes. The sequences were compared with the GenBank database, and data analyzed for significant statistical correlations using SPSS 16.0. Ethical approval to conduct the study was obtained from KNH-ERC.ResultsOut of the fifty-eight cases of SCCC analyzed, twenty-nine (50%) had well differentiated (grade 1), twenty one (36.2%) moderately differentiated (grade 2) while eight (13.8%) had poorly differentiated (grade 3) tumours. Immunohistochemistry assay was done in all the fifty eight studied cases, of which thirty nine cases (67.2%) were positive for p16INK4A staining, forty eight cases (82.8%) for EGFR and fifty one cases (87.9%) showed positivity for p-EGFR. HPV DNA was detected in 4 out of 40 SCCC cases (10%) in which PCR was performed, with HPV16 being the only HPV sub-type detected. Significant statistical association was found between HPV detection and p16INK4 (p=0.000, at 99% C.I) and EGFR (p=0.028, at 95% C.I) expressions, but not pEGFR. In addition, the expressions of these biomarkers did not show any significant association with tumor grades.ConclusionThis study points to an association of high risk HPV with over expressions of p16INK4A and EGFR proteins in AIDS-associated SCCC.

Evaluation of an automated SPF10-LiPA25 assay for detection and typing of human papillomavirus in archival samples

An automated blotter was evaluated for detection of the HPV SPF10-LiPA25 RHA for HPV genotyping with 63 FFPE tissue and 45 WHO proficiency panel samples. The results showed that 10–20% more types were detected and bands were of significantly stronger intensity (p<0.0001) compared to manual processing. Therefore it was concluded that automated detection of HPV genotypes using the HPV SPF10-LiPA25 is more sensitive for low copy number and mixed-type HPV samples.

P012 Systematic review of HPV test [Invader Technology/Real-Time PCR

ObjectiveHPV Test [Invader Technology/Real-Time PCR] is a molecular genetic test performed for confirming the presence of infection by detecting 14 types of HPV and additionally, for classifying types 16 and...

5. Detection of HIV DNA and HPV genotypes in the CD14+ fraction of anal cytology samples from HIV-positive human subjects

Background HIV-seropositive patients have a higher prevalence of other viral infections, including HPV, even while on anti-retroviral therapy. Anal infection with high-risk HPV genotypes can lead to anal dysplasia, which can progress to anal cancer. HIV and HPV can be contracted via sexual transmission; however, the interaction between these two pathogens and effect on the immune system are poorly understood. Since CD14 cells have been implicated in disease progression in other HIV-associated diseases, the objective of this study was to characterise CD14 cells from anal cytology specimens. Methods: Anal cytology samples were obtained from 14 HIV-positive patients enrolled in an anal dysplasia study. Samples were sorted for CD14 subsets. Following DNA extraction, HIV DNA copy was assessed by real-time PCR with primers/probes for β-globin and HIV gag genes. HPV DNA was amplified and genotypes identified using Roche’s LINEAR ARRAY HPV genotyping kit. Results: HIV DNA was detected in samples from 8 patients. Of those, 5 patients also tested positive for HPV genotypes. All 6 patients who tested HIV DNA-negative also tested HPV-positive. Due to low numbers, difference in HPV status relative to HIV DNA status was not statistically significant. HPV genotypes closely corresponded between CD14+ and CD14– fractions. Conclusions: Both HIV DNA and HPV genotypes can be detected from a single anal cytology specimen. Further studies are necessary to elucidate the significance of HPV genotypes detected in CD14+ fractions as well as the consequence of HIV and HPV coinfection in anal epithelia.