Detection and quantitation of HPV in genital and oral tissues and fluids by real time PCR
Tóm tắt
Human papillomaviruses (HPVs) remain a serious world health problem due to their association with anogenital/oral cancers and warts. While over 100 HPV types have been identified, a subset is associated with malignancy. HPV16 and 18 are the most prevalent oncogenic types, while HPV6 and 11 are most commonly responsible for anogenital warts. While other quantitative PCR (qPCR) assays detect oncogenic HPV, there is no single tube assay distinguishing the most frequent oncogenic types and the most common types found in warts. A Sybr Green-based qPCR assay was developed utilizing degenerate primers to the highly conserved HPV E1 theoretically detecting any HPV type. A single tube multiplex qPCR assay was also developed using type-specific primer pairs and TaqMan probes that allowed for detection and quantitation of HPV6,11,16,18. Each HPV type was detected over a range from 2 × 101 to 2 × 106copies/reaction providing a reliable method of quantitating type-specific HPV in 140 anogenital/cutaneous/oral benign and malignant specimens. 35 oncogenic and low risk alpha genus HPV types were detected. Concordance was detected in previously typed specimens. Comparisons to the gold standard detected an overall sensitivity of 89% (95% CI: 77% - 96%) and specificity of 90% (95%CI: 52% - 98%). There was good agreement between the ability of the qPCR assays described here to identify HPV types in malignancies previously typed using standard methods. These novel qPCR assays will allow rapid detection and quantitation of HPVs to assess their role in viral pathogenesis.
Tài liệu tham khảo
de Villiers EM, Fauquet C, Broker TR, Bernard HU, zur Hausen H: Classification of papillomaviruses. Virology 2004, 324: 17-27. 10.1016/j.virol.2004.03.033
zur Hausen H: Papillomaviruses and cancer: from basic studies to clinical application. Nat Rev Cancer 2002, 2: 342-50. 10.1038/nrc798
Palefsky J: Human papillomavirus and anal neoplasia. Curr HIV/AIDS Rep 2008, 5: 78-85. 10.1007/s11904-008-0013-5
Dianzani C, Calvieri S, Pierangeli A, Degener AM: Identification of human papilloma viruses in male dysplastic genital lesions. New Microbiol 2004, 27: 65-9.
Herrero R, Castellsague X, Pawlita M, Lissowska J, Kee F, Balaram P, Rajkumar T, Sridhar H, Rose B, Pintos J, et al.: Human papillomavirus and oral cancer: the International Agency for Research on Cancer multicenter study. J Natl Cancer Inst 2003, 95: 1772-83.
Schwartz SM, Daling JR, Doody DR, Wipf GC, Carter JJ, Madeleine MM, Mao EJ, Fitzgibbons ED, Huang S, Beckmann AM, et al.: Oral cancer risk in relation to sexual history and evidence of human papillomavirus infection. J Natl Cancer Inst 1998, 90: 1626-36. 10.1093/jnci/90.21.1626
D'Souza G, Kreimer AR, Viscidi R, Pawlita M, Fakhry C, Koch WM, Westra WH, Gillison ML: Case-control study of human papillomavirus and oropharyngeal cancer. N Engl J Med 2007, 356: 1944-56. 10.1056/NEJMoa065497
Munoz N, Bosch FX, de Sanjose S, Herrero R, Castellsague X, Shah KV, Snijders PJ, Meijer CJ: Epidemiologic classification of human papillomavirus types associated with cervical cancer. N Engl J Med 2003, 348: 518-27. 10.1056/NEJMoa021641
Swan DC, Tucker RA, Tortolero-Luna G, Mitchell MF, Wideroff L, Unger ER, Nisenbaum RA, Reeves WC, Icenogle JP: Human papillomavirus (HPV) DNA copy number is dependent on grade of cervical disease and HPV type. J Clin Microbiol 1999, 37: 1030-4.
Kreimer AR, Clifford GM, Snijders PJ, Castellsague X, Meijer CJ, Pawlita M, Viscidi R, Herrero R, Franceschi S: HPV16 semiquantitative viral load and serologic biomarkers in oral and oropharyngeal squamous cell carcinomas. Int J Cancer 2005, 115: 329-32. 10.1002/ijc.20872
Tucker RA, Unger ER, Holloway BP, Swan DC: Real-time PCR-based fluorescent assay for quantitation of human papillomavirus types 6, 11, 16, and 18. Mol Diagn 2001, 6: 39-47. 10.2165/00066982-200106010-00005
Swan DC, Tucker RA, Holloway BP, Icenogle JP: A sensitive, type-specific, fluorogenic probe assay for detection of human papillomavirus DNA. J Clin Microbiol 1997, 35: 886-91.
Peitsaro P, Johansson B, Syrjanen S: Integrated human papillomavirus type 16 is frequently found in cervical cancer precursors as demonstrated by a novel quantitative real-time PCR technique. J Clin Microbiol 2002, 40: 886-91. 10.1128/JCM.40.3.886-891.2002
Nagao S, Yoshinouchi M, Miyagi Y, Hongo A, Kodama J, Itoh S, Kudo T: Rapid and sensitive detection of physical status of human papillomavirus type 16 DNA by quantitative real-time PCR. J Clin Microbiol 2002, 40: 863-7. 10.1128/JCM.40.3.863-867.2002
Josefsson A, Livak K, Gyllensten U: Detection and quantitation of human papillomavirus by using the fluorescent 5' exonuclease assay. J Clin Microbiol 1999, 37: 490-6.
Gravitt PE, Peyton C, Wheeler C, Apple R, Higuchi R, Shah KV: Reproducibility of HPV 16 and HPV 18 viral load quantitation using TaqMan real-time PCR assays. J Virol Methods 2003, 112: 23-33. 10.1016/S0166-0934(03)00186-1
Broccolo F, Cocuzza CE: Automated extraction and quantitation of oncogenic HPV genotypes from cervical samples by a real-time PCR-based system. J Virol Methods 2008, 148: 48-57. 10.1016/j.jviromet.2007.10.003
Takacs T, Jeney C, Kovacs L, Mozes J, Benczik M, Sebe A: Molecular beacon-based real-time PCR method for detection of 15 high-risk and 5 low-risk HPV types. J Virol Methods 2008, 149: 153-62. 10.1016/j.jviromet.2007.12.009
Schmitz M, Scheungraber C, Herrmann J, Teller K, Gajda M, Runnebaum IB, Durst M: Quantitative multiplex PCR assay for the detection of the seven clinically most relevant high-risk HPV types. J Clin Virol 2009, 44: 302-7. 10.1016/j.jcv.2009.01.006
Roden R, Wu TC: How will HPV vaccines affect cervical cancer? Nat Rev Cancer 2006, 6: 753-63. 10.1038/nrc1973
Koutsky LA, Harper DM: Chapter 13: Current findings from prophylactic HPV vaccine trials. Vaccine 2006,24(Suppl 3):S3/114-21.
Sanchez JL, Storch GA: Multiplex, quantitative, real-time PCR assay for cytomegalovirus and human DNA. J Clin Microbiol 2002, 40: 2381-6. 10.1128/JCM.40.7.2381-2386.2002
D'Souza G, Fakhry C, Sugar EA, Seaberg EC, Weber K, Minkoff HL, Anastos K, Palefsky JM, Gillison ML: Six-month natural history of oral versus cervical human papillomavirus infection. Int J Cancer 2007, 121: 143-50. 10.1002/ijc.22667
Andrews E, Seaman WT, Webster-Cyriaque J: Oropharyngeal carcinoma in non-smokers and non-drinkers: a role for HPV. Oral Oncol 2009, 45: 486-91. 10.1016/j.oraloncology.2008.07.008
Watzinger F, Ebner K, Lion T: Detection and monitoring of virus infections by real-time PCR. Mol Aspects Med 2006, 27: 254-98. 10.1016/j.mam.2005.12.001
Markowitz LE, Sternberg M, Dunne EF, McQuillan G, Unger ER: Seroprevalence of human papillomavirus types 6, 11, 16, and 18 in the United States: National Health and Nutrition Examination Survey 2003-2004. J Infect Dis 2009, 200: 1059-67. 10.1086/604729
Kleter B, van Doorn LJ, ter Schegget J, Schrauwen L, van Krimpen K, Burger M, ter Harmsel B, Quint W: Novel short-fragment PCR assay for highly sensitive broad-spectrum detection of anogenital human papillomaviruses. Am J Pathol 1998, 153: 1731-9.
Gillison ML, Koch WM, Capone RB, Spafford M, Westra WH, Wu L, Zahurak ML, Daniel RW, Viglione M, Symer DE, et al.: Evidence for a causal association between human papillomavirus and a subset of head and neck cancers. J Natl Cancer Inst 2000, 92: 709-20. 10.1093/jnci/92.9.709