Title: Human Papillomavirus: A Study of Type Specific Antibody Positivity among College Age Women
Students: Kent Buck, Valerie White
Team: 4
Faculty Mentor: Scott Wright
Date: 6 October, 2004

Abstract:
Emerging information about the high prevalence of sexually transmitted HPV and the ability of certain subtypes to predispose infected women to cervical cancer suggests the need for information relating type specificity of antibodies to HPV subtypes as determined by serological testing.  We propose to screen college age women for the virus and if subjects exhibit positivity, determine the subtype of the virus through PCR methodology.  The same patients will then be screened for type specific antibodies in the blood and saliva.

Introduction:
Human Papillomaviruses (HPV’s) produce squamous epithelial tumors.  Certain types of HPV cause the common wart of the fingers, and others cause Condylomata acuminata in the anogenital region (genital warts), some of which can become severely dysplastic and progress to invasive carcinoma (Eron, Sherwood et. al., 1998).  Other types cause common warts such as Plantar wars, Flat warts, Butchers warts, Oral focal epithelial hyperplasia, and other common and not so common epithelial tumors (Eron, Sherwood et. al., 1998).
  
Over 100 subtypes of HPV have been identified; approximately 35 infect the anogenital area through sexual transmission (Koutsky & Kiviat, 1999). Of these types i.e., HPV-16, HPV-18, HPV-30, HPV-31, HPV-33, HPV-35, HPV-39, HPV-45, HPV-51, HPV-52, HPV-56, have been shown to dispose the infected female to Cervical dysplasia and carcinoma as well as bowenoid papulosis (Eron, Sherwood et. al., 1998).

Objective:
Objectives of this study include; Testing to determine the prevalence of HPV’s in college age women at Weber State University and, among those who test positive, the prevalence of subtypes of the virus that predispose the subject to cervical cancer.  The last objective of the study is to determine whether type specific antibodies can be discovered in the serum and/or saliva and consider the possibility of a serological blood test that could determine infection with a subtype that could predispose one to Cervical Cancer.

Methods:
The method used to test for HPV is polymerase chain reaction (PCR) on the vaginal swab specimen and serological testing on the plasma specimen for HPV antibodies. The testing will be done by Impact Diagnostics (Salt Lake City, UT) personnel. The PCR positive specimens are then genotyped by SBT to determine the exact viral type(s) present in the sample.

Real Time Polymerase Chain Reaction (R-PCR) technology is a nucleic acid amplification testing (NAT) technology that uses fluorescent detection. Using R-PCR allows HPV detection to be 17% more sensitive than the most common HPV test. Positive R-PCR samples are then reflexed to proprietary genotyping assay. SBT uses primers specific for the PCR positive samples to uniquely identify the genotype of over 18 viral types.

The ELISA assay is still experimental and proof of concept is part of this study. Basically, HPV peptides are bound to a 96-well plate, Patient serum is then added to the plate and any HPV antibodies present can bind to antigens on the peptides. After binding, excess serum is washed away and a secondary antibody that will bind to the fc portions of the HPV antibodies is added to the plate. Finally, substrate (TMB turbo) is added to the plate and positive samples turn blue. The positive signal strengths are measured in a plate reader.

Materials:
We are considering applying for a grant for the purpose of providing incentives for participants to donate specimens, such as lunch or a gift certificate for food at a local restaurant. Impact Diagnostics will provide specimen collection kits including swabs, blood collection tubes and needles, reagents and testing materials at an estimated cost of $1000.

References:
Matthews, Dawn D., Sexually Transmitted Diseases Sourcebook, Second Edition, Detroit, Omigraphics, 2001.
Gorback, Sherwood L., M.D., Bartlett, John G., M.D., Blacklow, Neil, R., MD, Infectious
        Diseases, Second Edition, Philadelphia, WB Saunders, 1992.
Mays, Rose M, PhD, Rn, et. al., “Human Papillomavirus, Genital Warts, Pap Smears, and
Cervical Cancer: Knowledge and Beliefs of Adolescent and Adult Women”. Healthcare for Women International, Vol 21, p. 361-374, 2000.
Rosenthal, Dorothy L., MD, FIAC, “Cervical disease screening and detection: emergin
        techniques in molecular diagnostic assays”., MLO, September 2004.

Reviewers: