Comparison of the Digene Hybrid Capture 2 assay and Roche AMPLICOR and LINEAR ARRAY human papillomavirus (HPV) tests in detecting high-risk HPV genotypes in specimens from women with previous abnormal Pap smear results

The development of cervical cancer is strongly associated with the presence of persistent high-risk (HR) human papillomavirus (HPV) infection. Recently, the commercially manufactured PCR-based Roche AMPLICOR (AMP) and LINEAR ARRAY (LA) HPV tests have become available for HPV detection. However, knowledge of their clinical performance compared to the U.S. Food and Drug Administration-approved Hybrid Capture 2 (HC2) assay is limited. This study evaluated the concordance between the HC2, AMP, and LA tests in detecting HR-HPV among a cohort of 1,679 women with previous abnormal Pap smear results. Overall, 1,393 specimens (81.3%) generated concordant results for HR-HPV presence or absence by the three assays. The concordance levels were substantial between the HC2 and AMP tests (84.4%, kappa = 0.6419) and between the HC2 and LA tests (84.0%, kappa = 0.6341) and nearly perfect between the AMP and LA tests (97.8%, kappa = 0.9441). HR-HPV prevalence, as detected by the AMP or LA tests, was significantly higher among women with cytological or histological high-grade disease (CIN2 or greater) than that detected by HC2 (P < 0.0001). The AMP and LA tests exhibited greater sensitivity, but lower specificity, than HC2 for detecting HR-HPV among this cohort of women with underlying cervical abnormalities, particularly among subjects with histologically proven high-grade disease. Both PCR-based HPV tests may be valuable in the management of care for women with underlying cervical abnormalities, in predicting treatment success, and in studying the clearance or acquisition of new infections.     

Comparison of the AMPLICOR human papillomavirus test and the hybrid capture 2 assay for detection of high-risk human papillomavirus in women with abnormal PAP smear

Infection with human papillomavirus (HPV) is a necessary step in the progression to cervical cancer. Many methods for HPV testing are currently available, mostly developed to detect pools of HPV types. Hybrid Capture 2 (HC2) is one of the most widely used. A new PCR-based assay, the Roche AMPLICOR HPV test, has been recently developed. Both assays recognize a group of 13 HR HPV types contemporaneously. This study evaluated the performance of both methods for detecting high-grade cervical lesions as a part of management for abnormal PAP smears. The study population was composed of 213 women, all referred to colposcopy and histologic diagnosis following an abnormal PAP test. Biopsy-confirmed high-grade cervical intraepithelial neoplasia was used as a gold standard. Overall agreement was 84.9% with a kappa value of 0.6. When comparing the ability to detect moderate cervical intraepithelial neoplasia (CIN2+) and high-grade cervical intraepithelial neoplasia (CIN3+/cancer), AMPLICOR proved slightly more sensitive than HC2, a finding that is important when HPV testing is used in a triage of borderline smear results. Genotyping of discordant results showed a prevalence of LR-HPV types in HC2 positive/AMPLICOR negative samples, and a similar prevalence of HR- and LR-HPV types in AMPLICOR positive/HC2 negative samples. In conclusion, the study shows that the AMPLICOR assay is more sensitive than HC2, which makes it a valid alternative for routine clinical use.     

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.     

Comparison of the Third Wave Invader human papillomavirus (HPV) assay and the digene HPV hybrid capture 2 assay for detection of high-risk HPV DNA

This study compared the clinical performance of the Digene Hybrid Capture 2 (HC2) assay to that of a prototype Third Wave Invader human papillomavirus (HPV) (IHPV) analyte-specific reagent-based assay for the detection of oncogenic or "high-risk" (HR) HPV DNA using liquid-based cytology specimens. In total, 821 ThinPrep vials were tested using both assays. In accordance with the type-specific probes contained within each test, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) for the IHPV assay were 95.9%, 97.6%, 97.5%, and 96.1%, respectively, and those for the HC2 assay were 98.1%, 86.2%, 87.1%, and 97.9%. Overall, the sensitivity and NPV were comparable between the assays, but the IHPV assay demonstrated a better specificity and PPV, since the IHPV assay had fewer false-positive HR HPV results. The incorporation of an internal control to evaluate the cellularity of the test material is an important feature of the IHPV assay and should reduce the risk of false-negative results due to insufficient sample collection rather than the lack of HR HPV DNA. An additional benefit of the IHPV assay was the smaller sample volume required (1 ml versus 4 ml for the HC2 assay).     

Prospective evaluation of the Hybrid Capture 2 and AMPLICOR human papillomavirus (HPV) tests for detection of 13 high-risk HPV genotypes in atypical squamous cells of uncertain significance

The use of high-risk human papillomavirus (hrHPV) testing as an adjunct to cervical cytology in population-based screening programs is currently based on DNA hybridization and PCR assays. The aim of this study was to prospectively assess the diagnostic performance of the Hybrid Capture 2 test (HC2; Digene Corporation) in comparison with that of the recently developed PCR-based AMPLICOR HPV test (Roche Molecular Systems) for the detection of 13 hrHPV types. A reverse line blot hybridization assay (Innogenetics) was used as an internal reference standard in discordant cases. Two hundred seventy-one patients with atypical squamous cells of uncertain significance (ASCUS) in cervical samples underwent hrHPV testing. The chi-square test was performed to compare respective proportions. Totals of 160/271 (59%) and 156/271 (58%) were found to be positive for hrHPV with HC2 and AMPLICOR, respectively. Concordant results were obtained for 235 (86.7%) of the 271 samples (kappa statistic, 0.73 +/- 0.04). Considering types 26, 53, and 66 as oncogenic types, negative predictive values (NPVs) of HC2 and AMPLICOR were 92.8% and 87.8%, respectively (difference was not significant), and their respective accuracies were 94.8% and 91.9% (difference was not significant). Considering types 26, 53, and 66 as not oncogenic, the respective HC2 and AMPLICOR NPVs were 92.8% and 97.4% (difference was not significant), and accuracy was significantly higher for the AMPLICOR assay (95.9% versus 90.8% for HC2) (P<0.05). For ASCUS samples, the NPV was 92.8% for HC2 testing and might be compromised if the copy number of HPV DNA was low. The NPV was 97.4% for the AMPLICOR assay and might be compromised if HPV types 26, 53, and 66 were considered oncogenic. The accuracy of these two assays is good and is compatible with routine clinical use in the triage of ASCUS cases.     

Assessment of MagNA pure LC extraction system for detection of human papillomavirus (HPV) DNA in PreservCyt samples by the Roche AMPLICOR and LINEAR ARRAY HPV tests

Roche Molecular Systems recently released two PCR-based assays, AMPLICOR and LINEAR ARRAY (LA), for the detection and genotyping, respectively, of human papillomaviruses (HPVs). The manual specimen processing method recommended for use with both assays, AmpliLute, can be time-consuming and labor-intensive and is open to potential specimen cross-contamination. We evaluated the Roche MagNA Pure LC (MP) as an alternative for specimen processing prior to use with either assay. DNA was extracted from cervical brushings, collected in PreservCyt media, by AmpliLute and MP using DNA-I and Total Nucleic Acid (TNA) kits, from 150 patients with histologically confirmed cervical abnormalities. DNA was amplified and detected by AMPLICOR and the LA HPV test. Concordances of 96.5% (139 of 144) (kappa=0.93) and 95.1% (135 of 142) (kappa=0.90) were generated by AMPLICOR when we compared DNA extracts from AmpliLute to MP DNA-I and TNA, respectively. The HPV genotype profiles were identical in 78.7 and 74.7% of samples between AmpliLute and DNA-I or TNA, respectively. To improve LA concordance, all 150 specimens were extracted by MP DNA-I protocol after the centrifugation of 1-ml PreservCyt samples. This modified approach improved HPV genotype concordance levels between AmpliLute and MP DNA-I to 88.0% (P=0.043) without affecting AMPLICOR sensitivity. Laboratories that have an automated MP extraction system would find this procedure more feasible and easier to handle than the recommended manual extraction method and could substitute such extractions for AMPLICOR and LA HPV tests once internally validated.     

Comparison of PapType to Digene Hybrid Capture 2, Roche linear array, and Amplicor for detection of high-risk human papillomavirus genotypes in women with previous abnormal pap smears

PapType human papillomavirus (HPV) assay was compared to Hybrid Capture 2 (HC2), Amplicor (Amp), and Linear Array (LA) HPV tests in 894 women undergoing management for a high-grade Pap smear abnormality. The sensitivity in detection of underlying high-grade histological diagnosis by PapType was 90.3% and by HC2 was 79.8%, while by Amp and LA it was 92.4% and 91.6%, respectively. The specificities were 52.5%, 55.3%, 49.4%, and 51.7% for PapType, HC2, Amp, and LA, respectively.     

Digene HPV Genotyping RH Test RUO: Comparative evaluation with INNO-LiPA HPV Genotyping Extra Test for detection of 18 high-risk and probable high-risk human papillomavirus genotypes

BackgroundStandardized and validated methods for the specific detection and identification of a spectrum of high-risk (hr) HPV genotypes will be necessary if HPV genotyping gains an important role in the clinical management of HPV-related precancerous lesions and cancers.ObjectivesThe first comparative evaluation of novel HPV genotyping Digene HPV Genotyping RH Test RUO (Qiagen, Hilden, Germany) with standard INNO-LiPA HPV Genotyping Extra CE assay (Innogenetics, Gent, Belgium).Study designSeventy hr-HPV positive samples were tested in parallel with both genotyping assays. The results were interpreted taking into account 15 hr-HPV and 3 probable hr-HPV genotypes that can be identified by both assays (assay-common genotypes).ResultsConcordant results (a complete match of assay-common genotypes or negative using both assays) and compatible results (at least one genotype in common) were obtained in 42 (60.0%) and 28 (40.0%) samples, respectively. No discordant results for assay-common genotypes were obtained. Of 42 samples with compatible results, the presence of at least one assay-common genotype was detected in 37 samples, while no HPV was detected in two samples by both assays and only a single low-risk HPV was detected by INNO-LiPA in three samples.ConclusionsA novel Digene test is suitable for the detection of hr-HPV genotypes in clinical samples and it provides comparable results to the well established INNO-LiPA assay. Although INNO-LiPA identified significantly more samples with multiple HPV genotypes than the Digene test, the clinical benefit of such a difference is at present unclear.     

Comparative evaluation of AMPLICOR HPV PCR and Linear Array assays on SurePath liquid-based Pap samples for the detection of high-risk HPV genotypes

BackgroundPersistent cervical infection with high-risk [HR] HPV is a causative factor for cancer. Liquid-based [L-Pap] Pap samples are convenient for HPV testing and SurePath samples have been least studied. Most HPV tests have multiple step protocols and testing laboratories experience large volumes of samples.ObjectivesUsing SurePath L-Pap residual samples the objectives were as follows: [1] to test the performance of AMP-HPV and LA-HPV. [2] To perform an agreement study between two laboratories for the AMP-HPV test and [3] to compare agreement of results between AMP-HPV and LA-HPV and HC2.Study designSamples from 657 women were tested for Pap cytology then assayed for HR-HPV using AMP-HPV and LA-HPV tests. AMP-HPV performance was compared between 2 laboratories and agreement studies were conducted between AMP-HPV, LA-HPV and HC2.ResultsHR-HPV genotypes were associated with L-Pap readings as follows: HSIL 92% [23/25], LSIL 73.6% [162/220], ASCUS 70.4% [131/186], normal 31.9% [72/226]. More women less than 30 were infected with HR-HPV and multiple genotypes regardless of the L-Pap reading. AMP-HPV and LA-HPV testing had an overall raw agreement with each other of 84.2% [Kappa 0.66] and each had agreement of 94% with HC2 testing of 133 samples [Kappa 0.86/0.87]. AMP-HPV agreement between two laboratories was better at 93% [Kappa 0.84] compared to 76.1% [Kappa 0.40] when extraction was standardized.ConclusionIt is feasible to perform AMP-HPV and LA-HPV on SurePath samples to detect HR-HPV genotypes. HC2, AMP-HPV and LA-HPV showed strong agreement. The extraction component of the AMP-HPV assay needs careful attention to yield consistent results.     

Comparison of the Roche AMPLICOR MYCOBACTERIUM assay and Digene SHARP Signal System with in-house PCR and culture for detection of Mycobacterium tuberculosis in respiratory specimens.

Two commercial primer kits and detection systems, the Roche AMPLICOR MYCOBACTERIUM test and the Digene primer-probe kit with the SHARP Signal System, were compared to in-house PCR as well as standard culture techniques. For the 27 culture-positive specimens, the Roche AMPLICOR MYCOBACTERIUM test detected 20 specimens, the Digene system detected 19, and in-house PCR detected 21. Of the 86 culture-negative specimens, 13 were positive by the Roche AMPLICOR MYCOBACTERIUM test, 16 were positive by the Digene system, and 21 were positive by in-house PCR. When clinical situations were evaluated, 11 of 13 culture-negative Roche AMPLICOR MYCOBACTERIUM test-positive specimens, 10 of 16 culture-negative Digene system-positive specimens, and 13 of 21 culture-negative-in-house PCR-positive specimens were diagnosed as true-positive specimens. The sensitivities of Roche AMPLICOR MYCOBACTERIUM test, the Digene system, and in-house PCR were 73.81, 69.05, and 80.95%, and the specificities were 97.18, 91.55 and 88.73%, respectively. The positive predictive values were 93.94, 82.86, and 80.95%, and the negative predictive values were 86.25, 83.33, and 88.73%, respectively. For the commercial kits, the Roche AMPLICOR MYCOBACTERIUM test seems to be more sensitive and specific than the Digene system. However, the Roche AMPLICOR MYCOBACTERIUM test cannot be used on nonrespiratory specimens.     

Comparison of the GenoFlow human papillomavirus (HPV) test and the Linear Array assay for HPV screening in an Asian population

High-risk human papillomavirus (HR-HPV) DNA detection in cervical cytology samples is useful for primary screening of cervical cancer and for triage of patients with equivocal cytological findings. The GenoFlow HPV array test (GF assay; Diagcor Bioscience Inc., Hong Kong) was recently developed to detect 33 HPV genotypes by a "flowthrough" hybridization technology. In this study, we assessed the analytical sensitivity and reproducibility of the GF assay and compared its genotyping results with those of the Linear Array (LA) assay (Roche Molecular Diagnostics, Indianapolis, IN), using 400 archived liquid-based cytology samples representing the full range of cytology findings. Genotyping findings of the GF and LA assays were concordant or compatible for 93.44% of tested samples, with a good (κ = 0.797) to very good (κ = 0.812) strength of agreement for assay-common and oncogenic HPV types, respectively. The two assays showed good (κ = 0.635) agreement in detecting infections with multiple HPV genotypes. The lowest detection limits of the GF assay for HPV16 and HPV18 were 25 copies and 20 copies, respectively. Repeat testing of 60 samples by use of two different lots of the GF assay revealed no discordant results, suggesting good reproducibility of the assay. Both assays achieved approximately 80% and 100% sensitivity for identifying cases of atypical squamous cells of undetermined significance (ASC-US) and low-grade squamous intraepithelial lesions (LSIL) with subsequent detection of LSIL+ and high-grade squamous intraepithelial lesions or higher (HSIL+) in 2 years, respectively. Among ASC-US samples, the GF assay achieved the highest specificity (23.08%) for indicating subsequent identification of HSIL compared with the LA (19.23%) and Hybrid Capture 2 (HC2) (8.97%) assays. The GF and LA assays showed significant discrepancy in detecting HPV genotypes 11, 26, 39, 52, and 66. More sensitive detection of HPV52 by GF assay offers an advantage in regions where HPV52 is more prevalent. The sensitivity of the GF assay for detecting patients with HSIL+ was noninferior to that of the LA assay.     

Agreement between the AMPLICOR Human Papillomavirus Test and the Hybrid Capture 2 assay in detection of high-risk human papillomavirus and diagnosis of biopsy-confirmed high-grade cervical disease

The AMPLICOR HPV test (AMP) and the Hybrid Capture 2 assay (HC2) detect 13 high-risk human papillomavirus (HR-HPV) types. Evaluation of comparative performance with clinical samples is needed to allow informed implementation of AMP into clinical practice. AMP was used (i) to assess the prevalence of HR-HPV in 1,032 samples of known cytology, HC2 status, and/or confirmed histology; (ii) to determine agreement between AMP and HC2; (iii) to evaluate the clinical sensitivity and specificity for detecting HR-HPV; and (iv) to detect the presence of biopsy-confirmed high-grade cervical intraepithelial neoplasia. The prevalence of HR-HPV was 39.3% and 45.6% by AMP and HC2, respectively. Overall agreement was 89.2% (kappa value, 0.78). Of 509 HR-HPV-negative specimens by HC2, 488 (95.9%) were AMP negative. Of 427 HR-HPV-positive specimens by HC2, 347 (81.2%) were AMP positive. In comparing the ability to detect high-grade squamous intraepithelial lesions (HSIL), the two tests were positive for all HSIL samples. Both tests performed similarly on CIN2+ samples (clinical sensitivities were 96.7% and 97.8%, respectively, for AMP and HC2). The clinical specificities of AMP and HC2 were comparable (54.9% versus 51.6%; P=0.18). Genotyping of 20 HC2-negative/AMP-positive cases using alternative technologies revealed target HR genotypes in 63.1% of cases and low-risk types in 15.7% of cases, while 21% of cases were negative. In conclusion, AMP provides a viable alternative to HC2, with good agreement for samples with high-grade cytology and similar sensitivity in detecting CIN2+ lesions.     

Comparison of the Roche Cobas® 4800 HPV assay to Digene Hybrid Capture 2, Roche Linear Array and Roche Amplicor for Detection of High-Risk Human Papillomavirus Genotypes in Women undergoing treatment for cervical dysplasia

BackgroundThe recently FDA (U.S. food and drug administration) approved Roche Cobas^® 4800 (Cobas) human papillomavirus (HPV) has limited performance data compared to current HPV detection methods for test of cure in women undergoing treatment for high grade lesions.ObjectiveEvaluation of Cobas HPV assay using historical samples from women undergoing treatment for cervical dysplasia.Study designA selection of 407 samples was tested on the Cobas assay and compared to previous results from Hybrid Capture 2, HPV Amplicor and Roche Linear Array.ResultsOverall, a correlation between high-risk HPV positivity and high grade histological diagnosis was 90.6% by the Cobas, 86.1% by Hybrid Capture 2, 92.9% by HPV Amplicor and 91.8% by Roche Linear Array.ConclusionThe Cobas HPV assay is comparative to both the HPV Amplicor and Roche Linear Array assays and better than Hybrid capture 2 assay in the detection of High-Risk HPV in women undergoing treatment for cervical dysplasia.     

Comparison of Digene Hybrid Capture 2, GeneMatrix PapilloScreen,and a PCR sequencing assay in detecting high-risk and probable high-risk oncogenic HPV genotypes in specimens from Korean women

Most cervical cancers are caused by 15 high-risk (HR) and three probable high-risk (pHR) oncogenic types of human papillomavirus (HPV). However, current commercial HR HPV screening test products do not include pHR HPV genotypes. Recently, PapilloScreen has been developed to detect the 15 HR and three pHR HPV types. In this study, we evaluated the concordance levels and clinical performance of Hybrid Capture 2 (HC2), PapilloScreen, and a PCR sequencing assay in detecting HR and pHR HPV. The PapilloScreen (96.8 %) and PCR sequencing assay (96.8 %) demonstrated higher sensitivity than HC2 (80.7 %) for detecting HR and pHR HPV. The three assays showed similar specificities and positive or negative predictive values. The concordance levels were 86.5 % (κ = 0.68) and 86.5 % (κ = 0.67) between HC2 and PapilloScreen and between HC2 and PCR sequencing, respectively. A near-perfect concordance was observed between PapilloScreen and PCR sequencing (97.8 %, κ = 0.95). Overall, the agreement between the three assays suggests that the results obtained by the HC2 assay are more often discordant (12.6 %) than the PCR-based tests. In conclusion, PapilloScreen is highly sensitive for detecting high-grade CIN or cervical cancer. The PapilloScreen assay should be considered an accurate and sensitive method for detecting HR and pHR HPV infections and an epidemiological tool for prevalence studies as well as early diagnosis and intervention in HR and pHR HPV infections.     

Comparison of HPV genotyping assays and Hybrid Capture 2 for detection of high-risk HPV in cervical specimens

High-risk types of human papillomavirus (HR-HPV) are among the primary causes of cervical cancers. Hybrid Capture 2 (HC-II) (Digene, Gaithersburg, MD), which detects 13 HR-HPVs as a group, is the only HPV assay approved to date by the United States Food and Drug Administration. In Korea, several HPV genotyping assays are commercially available, including HPV RFMP (GeneMatrix Co., Seoul), HPVDNACHIP (Biomedlab Co., Seoul), and MyHPV Chip (Mygene Co., Seoul). We compared the results of these assays with those of HC-II. Among 553 residual samples of liquid-based Pap tests, a total of 435 (78.7%) were available for HPV assays. They were classified into four cytologic categories: normal, atypical squamous cells of undetermined significance (ASCUS), low-grade cervical squamous intraepithelial lesions (LSIL), and high grade SIL or carcinoma (HSIL+). Among these samples, 23.0%, 40.6%, 82.5%, 93.8% were HR-HPV positive by HC-II, respectively; 6.6%, 18.1%, 44.4%, 84.4%, by HPV RFMP, respectively; 5.7%, 24.5%, 54.0%, 84.4%, by HPVDNACHIP, respectively; and 6.6%, 11.6%, 42.9%, 84.4%, by MyHPV, respectively. Compared with HC-II, the concordance rates and kappa values were 70.6% and 0.421 for HPV RFMP; 75.4% and 0.514 for HPVDNACHIP; and 67.8% and 0.367 for MyHPV. The concordance rates and kappa values between genotyping assays were 85.1% and 0.644 for HPV RFMP and HPVDNACHIP; 83.4% and 0.574 for HPV RFMP and MyHPV Chip; and 82.8% and 0.579 for HPVDNACHIP and MyHPV Chip. In conclusion, compared with HC-II test, the genotyping tests showed more than fair concordance but lower sensitivity in the detection of HR-HPVs, limiting their usefulness as HR-HPV screening tools.     

Enhanced detection and typing of human papillomavirus (HPV) DNA in anogenital samples with PGMY primers and the Linear array HPV genotyping test

The Roche PGMY primer-based research prototype line blot assay (PGMY-LB) is a convenient tool in epidemiological studies for the detection and typing of human papillomavirus (HPV) DNA. This assay has been optimized and is being commercialized as the Linear Array HPV genotyping test (LA-HPV). We assessed the agreement between LA-HPV and PGMY-LB for detection and typing of 37 HPV genotypes in 528 anogenital samples (236 anal, 146 physician-collected cervical, and 146 self-collected cervicovaginal swabs) obtained from human immunodeficiency virus-seropositive individuals (236 men and 146 women). HPV DNA was detected in 433 (82.0%) and 458 (86.7%) samples with PGMY-LB and LA-HPV (P = 0.047), respectively, for an excellent agreement of 93.8% (kappa = 0.76). Of the 17,094 HPV typing results, 16,562 (1,743 positive and 14,819 negative results) were concordant between tests (agreement = 96.9%; kappa = 0.76). The mean agreement between tests for each type was 96.4% +/- 2.4% (95% confidence interval [CI], 95.6% to 97.2%; range, 86% to 100%), for an excellent mean kappa value of 0.85 +/- 0.10 (95% CI, 0.82 to 0.87). However, detection rates for most HPV types were greater with LA-HPV. The mean number of types per sample detected by LA-HPV (4.2 +/- 3.4; 95% CI, 3.9 to 4.5; median, 3.0) was greater than that for PGMY-LB (3.4 +/- 3.0; 95% CI, 3.1 to 3.6; median, 2.0) (P < 0.001). The number of types detected in excess by LA-HPV in anal samples correlated with the number of types per sample (r = 0.49 +/- 0.06; P = 0.001) but not with patient age (r = 0.03 +/- 0.06; P = 0.57), CD4 cell counts (r = 0.06 +/- 0.06; P = 0.13), or the grade of anal disease (r = -0.11 +/- 0.06; P = 0.07). LA-HPV compared favorably with PGMY-LB but yielded higher detection rates for newer and well-known HPV types.     

Human papillomavirus (HPV) in atypical squamous cervical cytology: the Invader HPV test as a new screening assay

In surveillance for cervical neoplasia, a diagnosis of cytologically atypical squamous cells of undetermined significance (ASCUS) presents a significant clinical issue, often dependent on testing for high-risk (HR) human papillomavirus (HPV) for the triage of patients. HPV type 16 now appears to be a critical concern in the follow-up of patients with ASCUS. The Invader HPV (Inv2) test, by Third Wave Technologies, Inc., is a recently developed analyte-specific reagent assay that uses probe sets for the detection of 14 HR HPV subtypes. These probe sets are A5/A6 (HPV types 51, 56, and 66), A7 (HPV types 18, 39, 45, 59, and 68), and A9 (HPV types 16, 31, 33, 35, 52, and 58). This report describes the performance characteristics of the Inv2 test in the screening of ASCUS cervical cytology specimens and correlates the results of the Inv2 test with those of the Hybrid Capture II HPV (HC2) test by Digene. The linear array HPV genotyping test (Roche Molecular Systems) was used as a reference method for the testing of samples with discordant results. Ninety-four Pap smear samples with a cytological diagnosis of ASCUS and 39 samples with a negative diagnosis were tested. The results of the Inv2 test demonstrated a good (86.6%) concordance with those of the HC2 test, with an overall sensitivity and specificity of 96% for the Inv2 test. Additionally, the Inv2 assay, which offers high-throughput, semiautomated DNA extraction, allows the subgrouping of HPV types by differential probe sets, could provide a useful test for screening for HPV, and has the potential to provide an improved means of risk stratification and the selection of patients for further HPV subtyping.     

Detection of high-risk subtypes of human papillomavirus in cervical swabs: routine use of the Digene Hybrid Capture assay and polymerase chain reaction analysis

Human papillomaviruses (HPVs) are major causative agents in the pathogenesis of cervical cancer, and more than twenty types are associated with its development. With the introduction of liquid-based preparation systems, it is envisaged that large-scale HPV testing will be established in the near future. Preliminary studies demonstrate the accessibility of these samples for DNA testing using both the Digene Hybrid Capture assay (DHCA) and polymerase chain reaction (PCR) techniques. This study aims to assess the validity and sensitivity of the DHCA system to detect high-risk HPV DNA, using two sets of HPV consensus primers (Gp5+/Gp6+ and MY09/MY11) in tandem with routine assessment of cervical smear and biopsy samples. Results indicate that the combination of DHCA and PCR detects more high-grade lesions than does the DHCA alone. DHCA-negative cases were categorised by subsequent PCR amplification into low-grade HPV-negative (12/16) cervical lesions and high-grade HPV-positive (7/9) cervical lesions. Gp5+/Gp6+ primers were less sensitive in detecting HPV-positive samples than was the MY09/MY11 primer set. These results support the use of high-risk HPV testing by DHCA, with subsequent analysis of DHCA-negative samples by PCR using the MY09/MY11 primers.     

Pap cytopathology and the presence of high-risk human papillomavirus in SurePath liquid preservative and Digene cervical sampler specimens

The Digene Hybrid Capture 2 (HC2) assay for high-risk human papillomavirus (HPV) identified 92 (28.7%) infected women by testing 320 SurePath liquid-based Pap samples and a hybrid capture 2 (HC2) sampler collection. HPV positivity was predictive for high-grade lesions. A majority of women had normal cervical readings although 15.8% were HPV-positive. Half of the patients with a reading of atypical squamous cells of undetermined significance (ASCUS) (n=16) were HPV-positive.     

Comparison of real-time multiplex human papillomavirus (HPV) PCR assays with INNO-LiPA HPV genotyping extra assay

Real-time type-specific multiplex human papillomavirus (HPV) PCR assays were developed to detect HPV DNA in samples collected for the efficacy determination of the quadrivalent HPV (type 6, 11, 16, and 18) L1 virus-like particle (VLP) vaccine (Gardasil). Additional multiplex (L1, E6, and E7 open reading frame [ORF]) or duplex (E6 and E7 ORF) HPV PCR assays were developed to detect high-risk HPV types, including HPV type 31 (HPV31), HPV33, HPV35, HPV39, HPV45, HPV51, HPV52, HPV56, HPV58, and HPV59. Here, we evaluated clinical specimen concordance and compared the limits of detection (LODs) between multiplex HPV PCR assays and the INNO-LiPA HPV Genotyping Extra assay, which detects 28 types, for the 14 HPV types common to both of these methods. Overall HPV detection agreement rates were >90% for swabs and >95% for thin sections. Statistically significant differences in detection were observed for HPV6, HPV16, HPV18, HPV35, HPV39, HPV45, HPV56, HPV58, and HPV59 in swabs and for HPV45, HPV58, and HPV59 in thin sections. Where P was <0.05, discordance was due to detection of more HPV-positive samples by the multiplex HPV PCR assays. LODs were similar for eight HPV types, significantly lower in multiplex assays for five HPV types, and lower in INNO-LiPA for HPV6 only. LODs were under 50 copies for all HPV types, with the exception of HPV39, HPV58, and HPV59 in the INNO-LiPA assay. The overall percent agreement for detection of 14 HPV types between the type-specific multiplex HPV PCR and INNO-LiPA genotyping assays was good. The differences in positive sample detection favored multiplex HPV PCR, suggesting increased sensitivity of HPV DNA detection by type-specific multiplex HPV PCR assays.