Development of a fluorescence–based multiplex genotyping method for simultaneous determination of human papillomavirus infections and viral loads

Abstract

BackgroundPersistent high-risk human papillomavirus (HPV) infection is correlated with an increased risk of developing intraepithelial lesion or malignancy (NILM). The aims of the current study is to establish a method named BioPerfectus Multiplex Real Time (BMRT) HPV assay for simultaneous typing and quantifying HPVs, and to evaluate it by comparison with HPV GenoArray test and PCR-sequencing method, as well as histological status.MethodsA total of 817 cervical specimens were evaluated by BMRT method and HPV GenoArray test, using PCR-sequencing method as the reference standard; simultaneously, high-risk HPV-16 and -18 DNA loads were assessed in 443 specimens to investigate the correlation with infection outcomes.ResultsThe overall detection coincidence rate between BMRT assay and HPV GenoArray test is 96.6 % and the Kappa value is 0.760. In addition, the sensitivity and positive predictive value of BMRT is 98.4 % and 95.7 % compared with the results detected by PCR-sequencing method, respectively. HPV-16 viral load has a correlation with CINs or worse lesions. By comparing with infected women presenting NILM /cervicitis, the cutoff value for HPV-16 from patients with CINs was 0.827. With this cutoff value, 74.6 % sensitivity and 72.5 % specificity for prediction of HPV-16 infected patients with CINI and higher CIN were achieved. High significance was obtained when comparing the infected women presenting NILM/cervicitis with women either with CIN and cervical carcinomas (p < 0.001).ConclusionsThe BMRT assay seemed to be a good alternative approach for HR-HPV testing, due to its high level of automation and ability to quantify HPV-16, HPV-18 and other HR-HPVs.

Multiple HPV Infections and Viral Load Association in Persistent Cervical Lesions in Mexican Women

“…HPV viral load in serial measuring could predict cervical lesion progression [22]. The risk of CC and HSIL increases with the viral load, which could triage HR-HPV-positive women [23,24].…”

Type-specific high-risk human papillomavirus viral load as a viable triage indicator for high-grade squamous intraepithelial lesion: a nested case&ndash;control study

“…A previous study reported 24 the BioPerfectus Multiplex Real-Time PCR (BMRT) assay, which was a newly developed, highly automated approach for quantifying viral loads for 14 HR-HPV types. This method provides an absolute quantitative measure of individual types with normalization according to the amount of human DNA.…”

“…This method provides an absolute quantitative measure of individual types with normalization according to the amount of human DNA. 24 , 25 In order to confirm that type-specific HPV viral loads correlate with the grade of cervical lesions and determine whether the type-specific HPV viral load is a viable risk identification tool for ≥HSIL in HR-HPV–positive women, the present study analyzed type-specific viral loads of 14 HR-HPV types using the BMRT assay in a large-scale clinical trial.…”

Negative Predictive Value of Human Papillomavirus Testing: Implications for Anal Cancer Screening in People Living with HIV/AIDS

“…DNA was extracted from biopsy samples using QIAamp DNA mini kit (Qiagen, Germany) according to the manufacturer's instructions. PCR was performed using the fluorescence-based multiplex HPV DNA genotyping kit (Bioperfectus Ltd., China) that is designed to identify 19 high-risk or possibly carcinogenic HPV types (16,18,26,31,33,35,39,45,51,52,53,56,58,59, 66, 67, 68, 73, and 82) and 3 low-risk HPV types (6, 11, and 81) [16]. Results were analyzed using the Perfectus Software v1.0 (Bioperfectus Ltd., China).…”