title: Melting curve analysis for the screening of hepatitis B virus genotypes A, D and F in patients from a general hospital in southern Brazil.
abstract: BACKGROUND
  Hepatitis B virus (HBV) can cause fulminant hepatitis, cirrhosis and hepatocellular carcinoma, and is one of the most common causes of acute and chronic liver failure. The genetic variants of HBV can be decisive for the evolution of these diseases as well as for the election of therapy.
  
  
  OBJECTIVE
  The aim of this study was to evaluate and standardize an in house methodology based on the analysis of the melting curve polymerase chain reaction (PCR) of real-time (qPCR) to screen for genotypes A, D and F of HBV in patients from a hospital in Rio Grande do Sul, Brazil.
  
  
  METHODS
  We evaluated 104 patients presumably with HBV chronic infection. Viral DNA was extracted from plasma and viral genotypes and different mutations were determined using PCR-based protocols.
  
  
  RESULTS
  A PCR-based methodology was standardized for the analysis of genotypes A, D and F of HBV. The technique was based in a nested PCR with the final step consisting of a multiplex real-time PCR, using the melting curve as a tool for the differentiation of fragments. A higher frequency of genotype D (44.4%), followed by genotype A (22.2%) and genotype F (3.7%) was observed.
  
  
  CONCLUSIONS
  The standardized assay, a nested PCR-multiplex qPCR using specific primers, provides a rapid and accurate method for the differentiation of HBV genotypes that are more frequent in Southern Brazil - A, D and F. This method can be applied in the clinical practice.
authors: Universidade Federal de Ciências da Saúde de Porto Alegre, Programa de Pós-Graduação em Medicina: Hepatologia.
authors: Carlos Eduardo
authors: CE
authors: Becker
authors: Nelson Alexandre
authors: NA
authors: Kretzmann
authors: Angelo Alves de
authors: AA
authors: Mattos
authors: Ana Beatriz Gorini da
authors: AB
authors: Veiga
mesh_terms: D001938:Brazil; D004279:DNA, Viral / Q000032:analysis; D005260:Female; D005838:Genotype; D006515:Hepatitis B virus / Q000235:genetics*; D019694:Hepatitis B, Chronic / Q000821:virology*; D006769:Hospitals, General; D006801:Humans; D008297:Male; D008875:Middle Aged; D060888:Real-Time Polymerase Chain Reaction; D044366:Transition Temperature
chemical_list: D004279:DNA, Viral