title: Evaluation of PCR-RFLP in the Pre-S Region as Molecular Method for Hepatitis B Virus Genotyping.
abstract: BACKGROUND
  Hepatitis B virus (HBV) infection is a public health problem in developing countries. HBV genotypes play major role in the evolution of infection since they were involved in different clinical presentations and response to treatment.
  
  
  OBJECTIVE
  This study was conducted to evaluate the efficiency of restriction fragment length polymorphism (RFLP) analysis for HBV genotyping.
  
  
  METHODS
  We investigated 98 samples collected from patients chronically infected with HBV. HBV genotypes were determined by analysis of patterns obtained after amplification in Pre-S region and digestion of the amplicon by two endonucleases AvaII and DpnII. Obtained results were confirmed by partial sequencing in the same region.
  
  
  RESULTS
  Two different HBV genotypes were detected in this study, Genotype D (in 95. 9%) and Genotype A (in 4.1%). Seventy-four samples (75.5%) were successfully genotyped with RFLP analysis and all classified as genotype D. The remaining 24 samples (24.5%) which were un-genotyped by RFLP analysis, were classified by partial sequencing of the pre-S region as HBV genotype D (20 samples, 20.4%) and genotype A (4 samples, 4.1%). Atypical profiles were significantly associated with advanced liver disease (P = 0.001) as well as older age (P < 0.05).
  
  
  CONCLUSIONS
  Several previous studies used PCR-RFLP to genotype HBV; however, we showed the high risk to obtain atypical profiles, especially in advanced stages of chronic infection, with as results difficulties to genotype the virus. These profiles resulted from the accumulation of mutations during natural course of infection resulting in a modification in restriction sites for enzymes. So, we recommended completing the investigation by partial sequencing to confirm obtained results.
authors.affiliation: Laboratory of Clinical Virology, Institute Pasteur de Tunis, Tunis, Tunisia.
authors.forename: Rim
authors.initials: R
authors.lastname: Ouneissa
authors.affiliation: Laboratory of Clinical Virology, Institute Pasteur de Tunis, Tunis, Tunisia.
authors.forename: Olfa
authors.initials: O
authors.lastname: Bahri
authors.affiliation: Laboratory of Clinical Virology, Institute Pasteur de Tunis, Tunis, Tunisia.
authors.forename: Ahlem
authors.initials: A
authors.lastname: Ben Yahia
authors.affiliation: Laboratory of Clinical Virology, Institute Pasteur de Tunis, Tunis, Tunisia.
authors.forename: Henda
authors.initials: H
authors.lastname: Touzi
authors.affiliation: Department of Gastroenterology, Hospital of Tahar Maamouri, Nabeul, Tunisia.
authors.forename: Mohamed Msaddak
authors.initials: MM
authors.lastname: Azouz
authors.affiliation: Department of Gastroenterology, Hospital La Rabta, Tunis, Tunisia.
authors.forename: Nabyl
authors.initials: N
authors.lastname: Ben Mami
authors.affiliation: Laboratory of Clinical Virology, Institute Pasteur de Tunis, Tunis, Tunisia.
authors.forename: Henda
authors.initials: H
authors.lastname: Triki
keywords: Direct Sequencing; Genotype; Hepatitis B Virus; Restriction Fragment Length Polymorphism