title: Diagnosis of 25 genotypes of human papillomaviruses for their physical statuses in cervical precancerous/cancerous lesions: a comparison of E2/E6E7 ratio-based vs. multiple E1-L1/E6E7 ratio-based detection techniques.
abstract: BACKGROUND
  Cervical lesions caused by integrated human papillomavirus (HPV) infection are highly dangerous because they can quickly develop into invasive cancers. However, clinicians are currently hampered by the lack of a quick, convenient and precise technique to detect integrated/mixed infections of various genotypes of HPVs in the cervix. This study aimed to develop a practical tool to determine the physical status of different HPVs and evaluate its clinical significance.
  
  
  METHODS
  The target population comprised 1162 women with an HPV infection history of > six months and an abnormal cervical cytological finding. The multiple E1-L1/E6E7 ratio analysis, a novel technique, was developed based on determining the ratios of E1/E6E7, E2/E6E7, E4E5/E6E7, L2/E6E7 and L1/E6E7 within the viral genome. Any imbalanced ratios indicate integration. Its diagnostic and predictive performances were compared with those of E2/E6E7 ratio analysis. The detection accuracy of both techniques was evaluated using the gold-standard technique "detection of integrated papillomavirus sequences" (DIPS). To realize a multigenotypic detection goal, a primer and probe library was established.
  
  
  RESULTS
  The integration rate of a particular genotype of HPV was correlated with its tumorigenic potential and women with higher lesion grades often carried lower viral loads. The E1-L1/E6E7 ratio analysis achieved 92.7% sensitivity and 99.0% specificity in detecting HPV integration, while the E2/E6E7 ratio analysis showed a much lower sensitivity (75.6%) and a similar specificity (99.3%). Interference due to episomal copies was observed in both techniques, leading to false-negative results. However, some positive results of E1-L1/E6E7 ratio analysis were missed by DIPS due to its stochastic detection nature. The E1-L1/E6E7 ratio analysis is more efficient than E2/E6E7 ratio analysis and DIPS in predicting precancerous/cancerous lesions, in which both positive predictive values (36.7%-82.3%) and negative predictive values (75.9%-100%) were highest (based on the results of three rounds of biopsies).
  
  
  CONCLUSIONS
  The multiple E1-L1/E6E7 ratio analysis is more sensitive and predictive than E2/E6E7 ratio analysis as a triage test for detecting HPV integration. It can effectively narrow the range of candidates for colposcopic examination and cervical biopsy, thereby lowering the expense of cervical cancer prevention.
authors.affiliation: Department of Obstetrics and Gynecology, Fengxian Hospital, Southern Medical University, 6600 Nanfeng Road, Shanghai, 201499, China. rongzhang@163.com.
authors.forename: Rong
authors.initials: R
authors.lastname: Zhang
authors.affiliation: Department of Obstetrics and Gynecology, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, 160 Pujian Road, Shanghai, 200127, China. he_yifeng@hotmail.com.|Shanghai Key Laboratory of Gynecologic Oncology, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, 160 Pujian Road, Shanghai, 200127, China. he_yifeng@hotmail.com.
authors.forename: Yi-feng
authors.initials: YF
authors.lastname: He
authors.affiliation: Department of Gynecology, Obstetrics and Gynecology Hospital, Fudan University, 419 Fangxie Road, Shanghai, 200011, China. chenmo_nicy@163.com.
authors.forename: Mo
authors.initials: M
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University, 220 Handan Road, Shanghai, 200433, China. cmchen@fudan.edu.cn.
authors.forename: Chun-mei
authors.initials: CM
authors.lastname: Chen
authors.affiliation: Shanghai Key Laboratory of Gynecologic Oncology, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, 160 Pujian Road, Shanghai, 200127, China. zhuqiujingsun@163.com.
authors.forename: Qiu-jing
authors.initials: QJ
authors.lastname: Zhu
authors.affiliation: Department of Obstetrics and Gynecology, Fengxian Hospital, Southern Medical University, 6600 Nanfeng Road, Shanghai, 201499, China. luhuan1979@outlook.com.
authors.forename: Huan
authors.initials: H
authors.lastname: Lu
authors.affiliation: Department of Obstetrics and Gynecology, Fengxian Hospital, Southern Medical University, 6600 Nanfeng Road, Shanghai, 201499, China. 954078787@qq.com.
authors.forename: Zhen-hong
authors.initials: ZH
authors.lastname: Wei
authors.affiliation: Department of Obstetrics and Gynecology, Fengxian Hospital, Southern Medical University, 6600 Nanfeng Road, Shanghai, 201499, China. lanxueyudie8989@sina.com.
authors.forename: Fang
authors.initials: F
authors.lastname: Li
authors.affiliation: Department of Obstetrics and Gynecology, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, 160 Pujian Road, Shanghai, 200127, China. mxcz5293@163.com.
authors.forename: Xiao-xin
authors.initials: XX
authors.lastname: Zhang
authors.affiliation: Department of Gynecology, Obstetrics and Gynecology Hospital, Fudan University, 419 Fangxie Road, Shanghai, 200011, China. xucongjian@gmail.com.
authors.forename: Cong-jian
authors.initials: CJ
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University, 220 Handan Road, Shanghai, 200433, China. longyu@fudan.edu.cn.
authors.forename: Long
authors.initials: L
authors.lastname: Yu
mesh_terms: D000328:Adult; D000367:Age Factors; D001483:Base Sequence; D005260:Female; D018628:Gene Dosage; D005838:Genotype; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D027383:Papillomaviridae / Q000235:genetics*; D015203:Reproducibility of Results; D002583:Uterine Cervical Neoplasms / Q000473:pathology / Q000821:virology*; D016662:Virus Integration
chemical_list: D009856:Oncogene Proteins, Viral