title: A method for rapid detection and genotype identification of hepatitis C virus 1-6 by one-step reverse transcription loop-mediated isothermal amplification.
abstract: OBJECTIVE
  Hepatitis C virus (HCV) is probably the leading cause of liver cirrhosis and hepatocellular carcinoma globally. Diagnostic tools conventionally used for the detection and identification of HCV infection are technically demanding, time-consuming, and costly for resource-limited environments. This study reports the development of the first rapid loop-mediated reverse transcription isothermal amplification assay that rapidly detects and identifies HCV genotypes in blood components.
  
  
  METHODS
  RNA extracted from donor plasma and serum specimens was applied to a one-step reverse transcription loop-mediated isothermal amplification reaction performed with HCV-specific oligonucleotides. Reactions were conducted at 63.5 °C for 30-60 min. The diagnostic characteristics of the assay were investigated and validated with clinical specimens.
  
  
  RESULTS
  Electrophoretic analysis of amplification revealed detection and identification of HCV genotypes 1-6. Positive amplification revealed unique ladder-like banding patterns that identified each HCV genotype. The assay demonstrated a sensitivity of 91.5% and specificity of 100%. Rapid naked-eye detection of HCV infection was facilitated by observation of an intense fluorescent glow of amplified targets under UV illumination.
  
  
  CONCLUSIONS
  These diagnostic characteristics highlight the potential utility of this assay for the rapid detection and genotype identification of HCV infection in field and point-of-care settings in endemic regions and resource-limited environments.
authors.affiliation: Division of Emerging and Transfusion-Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, 10903 New Hampshire Ave, Silver Spring, MD 20993, USA. Electronic address: dnyan@doctor.com.
authors.forename: Dougbeh-Chris
authors.initials: DC
authors.lastname: Nyan
authors.affiliation: Department of Biology, Morgan State University, Baltimore, Maryland, USA.
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Swinson
mesh_terms: D005838:Genotype; D016174:Hepacivirus / Q000235:genetics / Q000302:isolation & purification*; D006526:Hepatitis C / Q000175:diagnosis*; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012367:RNA, Viral / Q000302:isolation & purification; D048348:Reverse Transcription; D012680:Sensitivity and Specificity
chemical_list: D012367:RNA, Viral
keywords: Detection; Genotype identification; Hepatitis C virus; Isothermal amplification; Plasma; Serum