Development and evaluation of a colorimetric PCR system for the detection and typing of human papillomaviruses.
In developing countries, the introduction of human papillomaviruses (HPV) DNA testing as an adjunct to cytological screening programs has been delayed due to the lack of high performance and cost effective diagnostic nucleic acid methods. In this study we report the development and evaluation of the L1HPVPCR, a PCR-based method for the detection and typing of five of the most prevalent high-risk HPV types. The L1HPVPCR assay combines amplification with the MY09/11 HPV consensus primer system, liquid hybridization of the PCR products with no radioactive probes and enzyme immunoassay analysis. The technique is a user-friendly system that allows accurate HPV DNA detection and typing with inexpensive instrumentation that could be performed with not sophisticated reagents in almost any laboratory. Different cutoff points for generic and specific HPV detection were determined using reproducibility analysis and receiver operating characteristic curves to ensure good analytical sensitivity and clinical effectiveness. We used the L1HPVPCR assay to estimate the prevalence of HPV infection in 127 women at risk of cervical cancer from the city of Rosario (Argentina), where no epidemiological data has been previously reported. Further, we explored the clinical utility of the L1HPVPCR assay respect the Pap smear using a combined diagnosis of cytology, histology and colposcopy as gold standard. In conclusion, our results indicate that the assay described here provides a tool for accurate HPV DNA testing and could be applied in regions where no commercial tests are available.
Chouhy
Diego
D
Virology Area, School of Biochemistry and Pharmaceutical Sciences, Rosario National University, 2000 Rosario, Argentina.
Gil
Lisandro Benítez
LB
Nocito
Ana L
AL
Wojdyla
Daniel
D
Ornella
Leonardo
L
Cittadini
Jorge
J
Gardiol
Daniela
D
Giri
Adriana A
AA
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D003124:Colorimetry / Q000379:methods; D017931:DNA Primers; D015344:DNA Probes, HPV; D004279:DNA, Viral / Q000032:analysis; D005260:Female; D006801:Humans; D008875:Middle Aged; D027383:Papillomaviridae / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D030361:Papillomavirus Infections / Q000175:diagnosis*; D016133:Polymerase Chain Reaction / Q000379:methods*; D002583:Uterine Cervical Neoplasms / Q000175:diagnosis* / Q000821:virology*
D017931:DNA Primers; D015344:DNA Probes, HPV; D004279:DNA, Viral