title: How best to interpret mixed human papillomavirus genotypes in high-grade cervical intraepithelial neoplasia lesions.
abstract: OBJECTIVE
  This study aimed to determine human papillomavirus (HPV) genotypes present in biopsy sections from young women of vaccine eligible age living in Victoria, Australia, with confirmed cervical intraepithelial neoplasia grade 3 (CIN3) or adenocarcinoma in situ (AIS) using laser capture microdissection (LCM).
  
  
  METHODS
  Histologically confirmed CIN3 or AIS positive biopsies from vaccine eligible women (born after 30th June 1981, n=169), between May 2011 and March 2013, were identified. CIN3 or AIS lesions were isolated from biopsy material using LCM, and the HPV genotypes present in whole tissue sections (WTS) as well as LCM-isolated lesion tissue were determined by a sensitive reverse hybridisation assay; RHA kit HPV SPF10-LiPA25, version 1 (Labo Bio-medical Products, Rijswijk, The Netherlands).
  
  
  RESULTS
  One hundred and sixty-eight cases were shown to be HPV positive (99%), of which 20 (12%) had more than one HPV genotype detected using WTS-PCR. Evaluation by LCM of individual biopsies with mixed infections showed 18 cases (90%) had only one HPV genotype associated with each CIN3 lesion. HPV 16 was the most common HPV type, found in 95/168 cases (57%).
  
  
  CONCLUSIONS
  LCM-PCR allowed us to confirm the presence of a single HPV genotype associated with each biologically separate CIN3 lesion, supporting the theory that only one virus type causes each independent CIN lesion. LCM will provide an important tool in assessing vaccine effectiveness in HPV vaccine programs.
authors.affiliation: The University of Melbourne, Grattan Street, Parkville, Melbourne 3010, VIC, Australia; Murdoch Childrens Research Institute, Parkville, VIC, Australia. Electronic address: emmatc@student.unimelb.edu.au.
authors.forename: Emma T
authors.initials: ET
authors.lastname: Callegari
authors.affiliation: Regional HPV Reference Laboratory, Department of Microbiology and Infectious Diseases, The Royal Women's Hospital, Parkville, VIC, Australia; Murdoch Childrens Research Institute, Parkville, VIC, Australia; Department of Obstetrics and Gynaecology, University of Melbourne, Melbourne, VIC, Australia. Electronic address: Sepehr.Tabrizi@thewomens.org.au.
authors.forename: Sepehr N
authors.initials: SN
authors.lastname: Tabrizi
authors.affiliation: Department of Anatomical Pathology, Royal Women's Hospital, 5th Floor, 20 Flemington Road, Parkville, Melbourne 3052, VIC, Australia. Electronic address: Jan.Pyman@thewomens.org.au.
authors.forename: Jan
authors.initials: J
authors.lastname: Pyman
authors.affiliation: VCS Incorporated, 265 Faraday St, Carlton, Melbourne 3053, VIC, Australia. Electronic address: msaville@vcs.org.au.
authors.forename: Marion
authors.initials: M
authors.lastname: Saville
authors.affiliation: Regional HPV Reference Laboratory, Department of Microbiology and Infectious Diseases, The Royal Women's Hospital, Parkville, VIC, Australia; Murdoch Childrens Research Institute, Parkville, VIC, Australia. Electronic address: alyssa.cornall@mcri.edu.au.
authors.forename: Alyssa M
authors.initials: AM
authors.lastname: Cornall
authors.affiliation: VCS Incorporated, 265 Faraday St, Carlton, Melbourne 3053, VIC, Australia; School of Population and Global Health, University of Melbourne, Melbourne, VIC, Australia. Electronic address: jbrother@vcs.org.au.
authors.forename: Julia M L
authors.initials: JM
authors.lastname: Brotherton
authors.affiliation: Regional HPV Reference Laboratory, Department of Microbiology and Infectious Diseases, The Royal Women's Hospital, Parkville, VIC, Australia; Murdoch Childrens Research Institute, Parkville, VIC, Australia; Department of Obstetrics and Gynaecology, University of Melbourne, Melbourne, VIC, Australia. Electronic address: suzanne.garland@thewomens.org.au.
authors.forename: Suzanne M
authors.initials: SM
authors.lastname: Garland
mesh_terms: D065311:Adenocarcinoma in Situ / Q000821:virology*; D000293:Adolescent; D000328:Adult; D001315:Australia; D004279:DNA, Viral / Q000302:isolation & purification; D005260:Female; D005838:Genotype; D006801:Humans; D061346:Laser Capture Microdissection; D027383:Papillomaviridae / Q000145:classification*; D030361:Papillomavirus Infections / Q000821:virology*; D055815:Young Adult; D002578:Uterine Cervical Dysplasia / Q000821:virology*
chemical_list: D004279:DNA, Viral
keywords: AIS; CIN3; Cervical cancer; Human papillomavirus; Laser capture microdissection; Multiple infections