title: Precore/basal core promoter mutants quantification throughout phases of hepatitis B virus infection by Simpleprobe.
abstract: OBJECTIVE
  To investigate precore/basal core promoter (PC/BCP) mutants throughout hepatitis B virus (HBV) infection and to determine their relationship to hepatitis B early antigen (HBeAg) titers.
  
  
  METHODS
  We enrolled 191 patients in various stages of HBV infection at the Huashan Hospital and the Taizhou Municipal Hospital from 2010 to 2012. None of the patients received antiviral therapy. HBV DNA from serum, was quantified by real-time PCR. The HBV genotype was determined by direct sequencing of the S gene. We used the Simpleprobe ultrasensitive quantitative method to detect PC/BCP mutants in each patient. We compared the strain number, percentage, and the changes in PC/BCP mutants in different phases, and analyzed the relationship between PC/BCP mutants and HBeAg by multiple linear regression and logistic regression.
  
  
  RESULTS
  Patients with HBV infection (n = 191) were assigned to groups by phase: Immune tolerance (IT) = 55, Immune clearance (IC) = 67, Low-replicative (LR) = 49, and HBeAg-negative hepatitis (ENH) = 20. Of the patients (male, 112; female, 79) enrolled, 122 were HBeAg-positive and 69 were HBeAg-negative. The median age was 33 years (range: 18-78 years). PC and BCP mutation detection rates were 84.82% (162/191) and 96.86% (185/191), respectively. In five HBeAg-negative cases, we detected double mutation G1896A/G1899A. The logarithm value of PC mutant quantities (log10 PC) significantly differed in IT, IC, and LR phases, as well as in the ENH phase (F = 49.350, P < 0.001). The logarithm value of BCP mutant quantities (log10 BCP) also differed during the four phases (F = 25.530, P < 0.001). Log10 PC and log10 BCP values were high in the IT and IC phases, decreased in the LR phase, and increased in the ENH phase, although the absolute value at this point remained lower than that in the IT and IC phases. PC mutant quantity per total viral load (PC%) and BCP mutant quantity per total viral load (BCP%) differed between phases (F = 20.040, P < 0.001; F = 10.830, P < 0.001), with PC% and BCP% gradually increasing in successive phases. HBeAg titers negatively correlated with PC% (Spearman's rho = -0.354, P < 0.001) and BCP% (Spearman's rho = -0.395, P < 0.001). The negative correlation between PC% and HBeAg status was significant (B = -5.281, P = 0.001), but there was no such correlation between BCP% and HBeAg status (B = -0.523, P = 0.552).
  
  
  CONCLUSIONS
  PC/BCP mutants become predominant in a dynamic and continuous process. Log10 PC, log10 BCP, PC% and BCP% might be combined to evaluate disease progression. PC% determines HBeAg status.
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Wen-Hui
authors.initials: WH
authors.lastname: Tu
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Lv
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Yong-Mei
authors.initials: YM
authors.lastname: Zhang
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Hou
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Jin-Yu
authors.initials: JY
authors.lastname: Wang
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Yi-Jun
authors.initials: YJ
authors.lastname: Zhang
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Hong-Yan
authors.initials: HY
authors.lastname: Liu
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Hao-Xiang
authors.initials: HX
authors.lastname: Zhu
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Yan-Li
authors.initials: YL
authors.lastname: Qin
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Ri-Cheng
authors.initials: RC
authors.lastname: Mao
authors.affiliation: Wen-Hui Tu, Ying Lv, Yong-Mei Zhang, Jin-Yu Wang, Yi-Jun Zhang, Hong-Yan Liu, Hao-Xiang Zhu, Yan-Li Qin, Ri-Cheng Mao, Ji-Ming Zhang, Department of Infectious Disease, Huashan Hospital, Fudan University, Shanghai 200040, China.
authors.forename: Ji-Ming
authors.initials: JM
authors.lastname: Zhang
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D015415:Biomarkers / Q000097:blood; D002681:China; D004252:DNA Mutational Analysis / Q000379:methods*; D004279:DNA, Viral / Q000097:blood; D018450:Disease Progression; D005260:Female; D005838:Genotype; D006509:Hepatitis B / Q000097:blood / Q000175:diagnosis / Q000821:virology*; D006512:Hepatitis B Core Antigens / Q000235:genetics*; D006513:Hepatitis B e Antigens / Q000097:blood; D006515:Hepatitis B virus / Q000235:genetics* / Q000254:growth & development / Q000276:immunology; D006801:Humans; D016014:Linear Models; D016015:Logistic Models; D008297:Male; D008875:Middle Aged; D009154:Mutation; D010641:Phenotype; D060888:Real-Time Polymerase Chain Reaction; D012189:Retrospective Studies; D014758:Viral Core Proteins / Q000235:genetics*; D019562:Viral Load; D055815:Young Adult
chemical_list: D015415:Biomarkers; D004279:DNA, Viral; D006512:Hepatitis B Core Antigens; D006513:Hepatitis B e Antigens; D014758:Viral Core Proteins
keywords: Basal core promoter mutant; Hepatitis B early antigen titers; Hepatitis B virus; Precore mutant; Quantification