Testing and Genotyping of High-Risk Human Papillomavirus by the cobas HPV Test and the Hybrid Capture 2 High-Risk HPV DNA Test Using Cervical and Vaginal Samples

Abstract

cThe accurate detection and typing of high-risk human papillomavirus (HPV) are critical for cervical cancer screening. The Hybrid Capture 2 (hc2) and cobas HPV tests showed high agreement for cervical samples (94.4%, ‫؍‬ 0.72, n ‫؍‬ 693) and moderate agreement for vaginal samples ( ‫؍‬ 0.62, n ‫؍‬ 108). The HPV16 and HPV18 results were highly consistent between the cobas and Linear Array tests ( > 0.96, n ‫؍‬ 197). Three hc2-negative vaginal samples were repeatedly invalid by the cobas test due to ␤-globin control failures, highlighting amplification control benefits. No cross-contamination was detected in a challenge experiment. Cervical cytology screening has significantly decreased the incidence and mortality of cervical cancer. In 2013, an estimated 12,340 new cases of cervical cancer will be diagnosed and 4,030 deaths will occur in the United States (1). Persistent infection with high-risk (HR) human papillomavirus (HPV), particularly genotypes HPV16 and HPV18, is the major cause of cervical cancer. The American Cancer Society, the American Society for Colposcopy and Cervical Pathology, and the American Society for Clinical Pathology recommend cotesting (concurrent cytology and HPV screening) every 5 years for cervical cancer screening in women between 30 and 65 years of age (2-4). In women who have undergone a total hysterectomy for cervical precancer or cancer, HPV testing may be used (but has not been cleared by the U.S. FDA) with vaginal cytology specimens to detect vaginal neoplasias.There are currently four U.S. FDA-approved tests for HPV testing using cervical cytology samples in PreservCyt solution (ThinPrep; Hologic, Bedford, MA), including the Digene Hybrid Capture 2 High-Risk HPV DNA Test (hc2) (Qiagen, Gaithersburg, MD) and the cobas HPV test (cobas) (Roche Diagnostics, Indianapolis, IN). The hc2 test uses nucleic acid hybridization and chemiluminescent signal amplification to detect a pool of 13 HR-HPV types (5). The recently FDA-cleared cobas HPV test is an automated multiplex real-time PCR assay to detect separately HPV16, HPV18, a pool of 12 other-high-risk HPV (HR-HPV) types, and ␤-globin as a process control (6).The clinical performance of newer HPV tests is less well understood but almost impossible for most clinical laboratories to assess, since this requires very large prospective clinical trials. However, new tests offer advantages, including processing controls, simultaneous HPV16/18 genotyping, and automated workflow, which are important factors for clinical laboratories faced with choosing an HPV test. In addition, accurate HPV16/18 detection has increasingly important implications for triaging HR-HPVpositive women. We performed a comprehensive analytical performance evaluation with cervical specimens and assessed the genotyping accuracy, testing of vaginal specimens, precision, and carryover risk for the cobas test.Consecutive residual cervical cytology samples in PreservCyt medium (n ϭ 693) from women Ͼ30 years of age (range, 30 to 79 years; median, 45 years) with Ն8 ml residual v...

Comparison of CerviHPV and Hybrid Capture 2 HPV tests for detection of high‐risk HPV infection in cervical swab specimens

“…The concordance rate between the two assays was 84.5% with a kappa value of 0.579. Whereas the result is in line with various published studies comparing HC2 and other HR‐HPV screening tests, the agreement between the two tests may have been underestimated because the alkali condition of denatured HC2 samples could lead to DNA degradation through time, thus reducing the level of remaining HR‐HPV DNA for CerviHPV testing. This was also one reason that we focused our subsequent analyses mostly on the 15 HC2 positive discordant cases with RLU/CO values greater than 3.0.…”

“…cobasHPV also employs multiplex qPCR to detect the same set of 14 HR‐HPV with the ability to genotype HPV16 and 18, and also include a cellular control to avoid false negative results. Multiple studies have demonstrated comparable performance of cobasHPV and HC2 tests to detect HR‐HPV with overall agreement of 82.8% to 97.3%, similar to the agreement between CerviHPV and HC2 tests observed in this study. Thus, it would be interesting to know how CerviHPV test compares to cobasHPV test.…”

E6/E7 proteins are potential markers for the screening and diagnosis of cervical pre‑cancerous lesions and cervical cancer in a Chinese population

“…Denaturant (150 µl; digeneHC2HPVDNA detection kit; Qiagen Sciences, Inc.) was then added to the mixture. Following TCT, the residual samples were denatured by asymmetric PCR (Piko ® Thermal Cycler 96-well system; Thermo Fisher Scientific, Inc., Waltham, MA, USA) to obtain single-stranded DNA (13), which was then mixed and reacted with an RNA probe cocktail (BD Pharmingen; BD Biosciences, San Jose, CA, USA) for 13 HR-HPV oncogenic types (16,18,31,33,35,39,45, 51, 52, 56, 58, 59 and 68) to capture oncogenic HPV subtypes using DML-2000 gene hybridization amplifiers (Digene Corporation). The type of positive point HPV was determined according to the distribution pattern of HPV.…”

“…HC2 test (Qiagen Sciences, Inc., Gaithersburg, MD, USA), a signal amplification test based on the hybridization of a RNA probe cocktail for 13 high-risk oncogenic types with the target DNA, and capture and detection of the DNA-RNA hybrid by chemiluminescence, was applied to quantitate the level of HPV DNA for its standardization approved by the USA Food and Drug Administration (13). This test was performed with 4 ml of PreservCyt samples, and then mixed with 400 µl transfer buffer (digeneHC2HPVDNA detection kit; Qiagen Sciences, Inc.).…”

Comparative Evaluation of Three Commercial Systems for Detection of High-Risk Human Papillomavirus in Cervical and Vaginal ThinPrep PreservCyt Samples and Correlation with Biopsy Results

“…The pathogenesis of HPV is dependent on a number of factors, but it is now understood that certain genotypes of the virus are more closely associated with the development of severe disease, including cancer. HPV genotypes are commonly separated into two broad categories, (i) "low risk" (types 6,11,40,42,43,44,53,54,61,72,73, and 81) and (ii) "high risk" (types 16,18,31,33,35,39,45, 51, 52, 56, 58, 59, 66, and 68), based on the degree of correlation between infection and the development of cervical cancer. Among the 14 high-risk (HR) genotypes, HPV-16 and HPV-18 have been estimated to account for approximately 70% of cervical cancers (2).…”