Journal of Clinical Microbiology

Comparative performance of cobas 4800 HPV Test and Anyplex II HPV HR for high-risk human papillomavirus detection

ABSTRACT Numerous molecular tests are available to detect human papillomavirus (HPV). We compared the analytical performance of cobas and Anyplex for detection of high-risk (HR) carcinogenic HPV genotypes, assessed the composition of HPV types (other than 16 and 18) that influenced cobas performance, and considered the impact of viral load on test performance. We used data from the Early Detection of Cervical Cancer in Hard-to-Reach Populations of Women Through Portable and Point-of-Care HPV Testing project, which involved collection (2019–2022) of cervicovaginal samples from 1,042 women aged 21–74 years in Belgium ( n = 244), Portugal ( n = 309), Brazil ( n = 244), and Ecuador ( n = 245). Samples were tested by cobas (provides individual results for HPV16 and HPV18 and a pooled result for 12 other HR-HPV types) and Anyplex (provides separate results for 14 HR-HPVs). We calculated HPV positivity by each test and compared performance between tests by calculating Cohen’s kappa statistics. Based on 938 samples with complete data from both tests, positivity rates by cobas were 13.4%, 3.6%, 34.3%, and 45.3% for HPV16, HPV18, 12 pooled HR-HPVs, and any HR-HPV, respectively. Corresponding HPV positivity rates by Anyplex were 14.9%, 3.7%, 37.9%, and 50.0% for the same categories, respectively, with high concordance; kappa statistics were 0.90, 0.87, 0.82, and 0.85, respectively. Based on 355 samples that tested positive for at least 1 of the 12 pooled HR-HPVs, most types showed high agreement (80.9%–100.0%) between individual-Anyplex and pooled-cobas HPV results, except for HPV68 (61.3% agreement). Our findings suggest that the two commercial tests may have different performances, depending on the specific HPV types detected, emphasizing the need for continued research on conditions that may affect these tests, especially for less common or less studied HPV types. IMPORTANCE This study compared two commercial tests—cobas and Anyplex—for detecting high-risk HPV types in women undergoing routine cervical cancer screening or referred for colposcopy. Both tests provide separate results for HPV16 and HPV18, but Anyplex also identifies the remaining 12 high-risk HPV types individually, while cobas groups them together. Overall, we found a high level of agreement between the two tests, supporting their use in clinical practice. However, differences in detecting certain HPV types, particularly those that are less common or less studied, emphasize the importance of choosing the right test. As more countries switch to HPV-based cervical cancer screening, using tests that provide detailed results could help improve risk assessment and optimize patient care.

Self-collection for primary HPV screening using dry swabs: a review of clinical performance, laboratory considerations, and patient preferences

ABSTRACT For cervical cancer screening, testing for high-risk human papillomavirus (hrHPV) detects more high-grade precancerous lesions, has a higher negative predictive value, and requires fewer lifetime screenings compared to cytology. As a result, both the US Preventive Services Task Force and the World Health Organization have endorsed hrHPV testing as the preferred screening method. Despite the utility, there are significant implementation challenges to adopting hrHPV primary screening across patients, providers, and institutions that must be addressed to ensure its widespread effectiveness. Here, we take a laboratory-centric approach to reviewing hrHPV primary screening, including discussion of specimen types and collection methods. For user experience, clinical and analytical validations, we focused on self-collected vaginal swab specimens stored dry during transport. Our analysis indicates that clinical laboratories should do their part to engage with institutional and clinical leadership to validate and promote the use of vaginal self-sampling for cervical cancer screening options within and outside the clinic. This work highlights the multiple studies that have validated dry swab collection as a simplified and high-quality method for hrHPV detection.

Clinical Validation of the HPV-Risk Assay, a Novel Real-Time PCR Assay for Detection of High-Risk Human Papillomavirus DNA by Targeting the E7 Region

ABSTRACT The HPV-Risk assay is a novel real-time PCR assay targeting the E7 region of 15 high-risk human papillomavirus (HPV) types (i.e., HPV16, -18, -31, -33, -35, -39, -45, -51, -52, -56, -58, -59, -66, -67, and −68), and provides additional genotype information for HPV16 and HPV18. This study evaluated the clinical performance and reproducibility of the HPV-Risk assay with cervical scraping specimens and its utility with self-collected (cervico)vaginal specimens. The clinical performance of the HPV-Risk assay for cervical intraepithelial neoplasia of grade 2 or worse (CIN2+) with cervical scraping specimens was evaluated by a noninferiority analysis, relative to high-risk HPV GP5+/6+ PCR, following international guidelines for HPV test requirements for cervical cancer screening. The HPV-Risk assay showed clinical sensitivity for CIN2+ of 97.1% (95% confidence interval [CI], 89.1 to 99.3%; 67/69 samples) and a clinical specificity for CIN2+ of 94.3% (95% CI, 92.5 to 95.7%; 777/824 samples). The clinical sensitivity and specificity were noninferior to those of GP5+/6+ PCR (noninferiority score test, P = 0.006 and 0.0003, respectively). Intralaboratory reproducibility over time (99.5% [95% CI, 98.6 to 99.8%]; 544/547 samples, kappa = 0.99) and interlaboratory agreement (99.2% [95% CI, 98.6 to 99.8%]; 527/531 samples, kappa = 0.98) for the HPV-Risk assay with cervical scraping specimens were high. The agreement of the HPV-Risk assay results for self-collected (cervico)vaginal specimens and clinician-obtained cervical scraping specimens was also high, i.e., 95.9% (95% CI, 85.1 to 99.0%; 47/49 samples, kappa = 0.90) for self-collected lavage samples and 91.6% (95% CI, 84.6 to 95.6%; 98/107 samples, kappa = 0.82) for self-collected brush samples. In conclusion, the HPV-Risk assay meets the cross-sectional clinical and reproducibility criteria of the international guidelines for HPV test requirements and can be considered clinically validated for cervical screening purposes. The compatibility of the HPV-Risk assay with self-collected specimens supports its utility for HPV self-sampling.

Validation of a simplified HPV genotyping assay designed for cervical screening in low-resource settings

ABSTRACT Human papillomavirus (HPV) genotype predicts cervical cancer risk, and genotyping could help guide the management of HPV positives as part of cervical screening. An isothermal amplification HPV extended genotyping test (ScreenFire HPV RS assay) can assay up to 96 samples/controls in 1 hour plus preparation time. A novel format with pre-aliquoted reagents and an anti-contamination component (Zebra BioDome) could simplify the HPV testing process and reduce the chances of post-amplification contamination. We validated the Zebra BioDome formulation prior to its clinical use. Residual provider-collected cervical samples ( n = 450) from a population-based study in rural Nigeria were retested with ScreenFire, once using the standard assay version (liquid reagents combined onsite) and twice with Zebra BioDome. HPV results with adequate DNA ( N = 427) were analyzed channel-by-channel and using the cervical cancer risk-based hierarchy of HPV type channels (HPV16, else 18/45, else 31/33/35/52/58, else 39/51/56/59/68, else high-risk HPV negative) to evaluate Zebra BioDome repeatability and accuracy against the standard version. Zebra BioDome reduced the number of pipetting steps to run the ScreenFire HPV assay. Following amplification, the BioDome material formed a sealant layer above the reaction components. Zebra BioDome had excellent repeatability and agreement with the standard version, both at the channel-specific analysis (positive percent agreement between 88.4% [HPV39/51/56/59/68] and 100% [HPV16]; negative percent agreement between 97.8% [HPV31/33/35/52/58] and 100% [HPV39/51/56/59/68]) and hierarchical analysis (overall agreement 97.2%). The assay version utilizing Zebra BioDome performed similarly to the previously validated standard version of the ScreenFire HPV assay and is now undergoing field evaluation. This solution has the potential to reduce assay preparation time and risk of contamination, providing a simpler, low-cost, near-point-of-care HPV testing and extended genotyping solution for cervical screening in lower-resource settings. The potential application of Zebra BioDome technology to other PCR assays should be considered. IMPORTANCE This work validates a novel pre-packed formulation for the ScreenFire human papillomavirus (HPV) assay, which has the potential to simplify the HPV testing process and to reduce the chances of post-amplification contamination, providing a simpler, low-cost, near-point-of-care HPV testing, and extended genotyping solution for cervical screening in resource-limited settings as part of the ultimate public health goal to accelerate cervical cancer prevention. This technology can also have broad applications for other DNA amplification assays beyond HPV.

Evaluation and Optimization of the Clinical Accuracy of Hybribio's 14 High-Risk HPV with 16/18 Genotyping Assay within the VALGENT-3 Framework

Hybribio’s 14 High-Risk HPV with 16/18 genotyping real-time PCR (HBRT-H14) is a human papillomavirus (HPV) assay with approval from the China Food and Drug Administration that is widely used in China. VALGENT (VALidation of HPV GENotyping Tests) is an established framework for evaluating HPV tests’ clinical performance relative to validated comparators. The aim of this study was to assess the clinical accuracy of HBRT-H14 following international validation criteria. Within VALGENT-3, clinical performance of HBRT-H14 was compared with Hybrid Capture 2 (HC2), Linear Array HPV genotyping test (Linear Array), and Cobas 4800 HPV test (Cobas).

Testing for Human Papillomaviruses in Urine, Blood, and Oral Specimens: an Update for the Laboratory

Twelve high-risk alpha human papillomavirus (HPV) genotypes cause approximately 690,000 cancer cases annually, with cervical and oropharyngeal cancer being the two most prominent types. HPV testing is performed in laboratory settings for various applications of a clinical, epidemiological, and research nature using a range of clinical specimens collected by clinicians or by individuals (self-collected specimens).

HPV testing with 16/18 genotyping for risk stratification among women with normal cytology: a multicenter prospective cohort study from China

ABSTRACT To evaluate the clinical performance of Hybribio’s 14-type HPV real-time PCR with 16/18 genotyping (HBRT-H14) and its risk stratification utility among women with normal cytology (NILM). From 2017 to 2020, a multicenter cohort enrolled 8,401 women aged 30–64 years with NILM cytology. Baseline HPV testing used HBRT-H14. Women positive for HPV 16/18 were referred for colposcopy; follow-up was annual for 3 years or until the detection of cervical intraepithelial neoplasia grade 2 or worse (CIN2+). Analyses included 6,679 women who completed follow-up. Overall HPV positivity was 11.4%, including 2.3% HPV 16/18. Over 3 years, sensitivity and specificity of HPV positivity for CIN2+ were 92.3% (95% confidence interval [CI]: 84.2–96.4) and 89.6% (88.8–90.3). For HPV 16/18 positivity, sensitivity and specificity were 41.0% (30.8–52.1) and 98.2% (97.8–98.5). Three-year cumulative CIN2+ risk was 20.9% (15.2–28.1) for HPV 16/18-positive women, 6.6% (4.9–8.9) for other types, and 0.1% (0.04–0.2) for HPV-negative women. HBRT-H14 shows strong clinical performance for detecting CIN2+, and HPV 16/18 genotyping provides effective risk stratification among women with NILM cytology. Findings support integration of HBRT-H14 into HPV-based screening pathways with HPV 16/18 genotyping and cytology triage of other types. IMPORTANCE This multicenter prospective study evaluated the Hybribio 14 high-risk HPV real-time PCR assay (HBRT-H14) in 8,401 women with normal (NILM) cytology under guideline-based follow-up. The assay showed high clinical sensitivity and a very low risk among HPV-negative women, and HPV 16/18 genotyping provided clear risk stratification. These findings deliver large-scale, practice-oriented evidence supporting integration of HBRT-H14 into HPV-based screening pathways that use HPV 16/18 genotyping with cytology triage of other types.

Clinical and Analytical Performance of the BD Onclarity HPV Assay with SurePath Screening Samples from the Danish Cervical Screening Program Using the VALGENT Framework

The Val idation of HPV Gen otyping T ests (VALGENT) framework is an international cooperation designed to evaluate human papillomavirus (HPV) assays with genotyping capabilities.

Diagnostic accuracy of the Daye diagnostic tampon compared to clinician-collected and self-collected vaginal swabs for detecting HPV: a comparative study

ABSTRACT Cervical cancer screening is vital for achieving global elimination of this preventable disease. Vaginal self-sampling (VSS) for human papillomavirus (HPV) has the potential to increase screening uptake, particularly among individuals who may be underserved by clinician collection. Expanding self-sampling options with accurate, acceptable collection devices is essential. The Daye diagnostic tampon (DDT) offers an innovative approach, utilizing a tampon for HPV-based detection. This study assessed the diagnostic accuracy of DDT in detecting high-risk HPV infections, using vaginal clinician-collected swabs (CCS) as the reference standard. In this UK-based study, 260 participants provided CCS and VSS (with Copan FLOQSwabs) and DDT samples for HPV testing. Samples were analyzed using the Aptima HPV assay, which detects 14 high-risk HPV types. The sensitivity, specificity, positive predictive value, and negative predictive value of the DDT were evaluated against the CCS. Invalidity rates—HPV-negative results with negative internal controls—were compared across sampling methods. The DDT showed a sensitivity of 82.9% [95% confidence interval (CI): 72.4%–89.9%], specificity of 91.6% (CI: 86.4%–94.9%), and overall accuracy of 89.0% (CI: 84.4%–92.4%) relative to CCS. McNemar’s test showed no significant difference between CCS and DDT results ( P = 0.845). Valid result rates were highest for DDT (99.2%), followed by VSS (95.4%) and CCS (90.8%). The DDT demonstrates comparable accuracy to CCS for detecting high-risk HPV. This novel device shows promise as a self-sampling method. Furthermore, complementary research should focus on assessing DDT’s clinical performance in detecting HPV associated with cervical disease endpoints. IMPORTANCE Cervical cancer remains a leading preventable cause of cancer death globally, with persistent disparities in screening access. Self-sampling for HPV has emerged as a critical tool to improve screening uptake, particularly among underserved populations, yet device acceptability and diagnostic reliability remain barriers to equitable implementation. This study demonstrates that the Daye diagnostic tampon (DDT), a novel, tampon-based self-sampling method, achieves diagnostic accuracy comparable to clinician-collected swabs (sensitivity 82.9% and specificity 91.6%) while yielding fewer invalid results (0.8%) than conventional swabs. By aligning with a familiar menstrual product, the DDT addresses usability concerns that hinder confidence in existing self-sampling devices, as evidenced by 70.5% participant preference in focus groups. These findings advance progress toward World Health Organisation (WHO) cervical cancer elimination targets by validating a culturally resonant, high-performance alternative to clinic-based sampling. The DDT’s potential to expand screening access, especially in low-resource settings or among individuals avoiding pelvic exams, could transform preventive care landscapes, reducing disparities in a disease rooted in healthcare inequity.

Analytical and Clinical Sample Performance Characteristics of the Onclarity Assay for the Detection of Human Papillomavirus

The objective of this study was to determine the result reproducibility and performance of the BD Onclarity human papillomavirus (HPV) assay (Onclarity) on the BD Viper LT platform using both contrived and clinical specimens. Reproducibility was assessed in BD SurePath liquid-based cytology (LBC) medium (SurePath) using contrived panels (HPV genotype 16 [HPV16] positive, HPV18 positive, or HPV45 positive) or clinical specimens (HPV16, -18, -31, -33/58, -45, or -52 positive or HPV negative). In addition, specimens from 3,879 individuals from the Onclarity trial were aliquoted prior to or following cytology processing and tested for HPV.

Test Accuracy of Human Papillomavirus in Urine for Detection of Cervical Intraepithelial Neoplasia

The objective was to assess the diagnostic test accuracy of high-risk human papillomavirus (hrHPV) testing of self-collected urine and cervicovaginal samples for the detection of cervical intraepithelial neoplasia grade 2 or higher (CIN2+). We recruited a convenience sample of women 25 to 65 years of age who were undergoing clinically indicated colposcopy at two medical centers in North Carolina between November 2016 and January 2019.

Clinical and Analytical Evaluation of the Alinity m HR HPV Assay within the VALGENT-3 Framework

Only clinically validated human papillomavirus (HPV) tests should be used in cervical cancer screening. VALGENT provides a framework to validate new HPV tests.