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

Abstract Introduction Human papillomavirus (HPV) genotype predicts cervical cancer risk, and genotyping could help guide 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 controls/samples in one hour plus preparation time. A novel format with pre-aliquoted reagents and an anti-contamination component (Zebra BioDome™) could simplify the HPV testing process by substantially reducing the assay preparation time and the chances of post-amplification contamination. We validated Zebra BioDome formulation prior to its clinical use. Methods 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. Results 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%). Conclusions 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. Potential application of Zebra BioDome technology to other DNA amplification assays should be considered.