Automated high throughput DNA isolation for detection of human papillomavirus in oral rinse samples

Background: Oral HPV infection elevates risk of oropharyngeal cancer, but its natural history is unknown. Natural history studies necessitate validation of an automated, high-throughput method for HPV genomic DNA detection in oral rinse samples (ORS).

Objectives: To compare agreement of oral HPV detection in ORS processed by a magnetic-bead based automated platform to a previous gold-standard, manual protein-precipitation method. Agreement was compared to that of repeat sampling and repeat HPV testing.

Study design: HIV-infected individuals (n=100) provided two ORS collected 15 min apart. DNA was isolated from equal aliquots by either a protein-precipitation based (Puregene, Qiagen) or magnetic bead-based (QIAsymphony™ SP, Qiagen) method. HPV DNA was detected and type-specified by consensus primer PCR and reverse line blot hybridization. The kappa statistic was used to assess overall agreement (OA) and agreement on a positive test (Ps+).

Results: The DNA purification methods had very high agreement for categorizing an individual as HPV infected (OA=0.95; Ps+=0.94) as well as for detection of HPV type-specific infection (OA=0.99; Ps+=0.88) in ORS. Agreement for detection of HPV type-specific infection was greater than that observed with repeat oral rinse sampling (OA=0.99, Ps+=0.76) but comparable to inter-assay agreement (OA=1.00, Ps+=0.90).

Conclusions: HPV detection in ORS processed with a magnetic-bead based automated platform will facilitate large natural history studies of oral HPV infection necessary to evaluate the potential use of oral HPV detection in oral cancer screening.