Quasispecies of genotype 4 of hepatitis C virus genomes in Saudi patients managed with interferon alfa and ribavirin therapy

Many patients with hepatitis C virus [HCV] infection do not respond to antiviral treatment, possibly due to viral quasispecies. We aimed to investigate whether the quasispeices population could be used as a predictor of response to therapy in our patients. The quasispecies of HCV genotype 4 [HCV-4] were studied in 25 naive Saudi patients at zero, three, and six months following interferon alfa and ribavirin combination therapy. Hypervariable region 1 within the E2/NS1 gene of the virus was analyzed by the single-strand conformation polymorphism [SSCP] technique after amplification. Pretreatment DNA bands by SSCP [2-7 bands] were detected in all patients. In those who achieved a complete virological response within six months [viral load <0.2 Meq/mL; n=7], bands ranged from 2-6 [mean = 3.71 +/- 1.25]. In six of these seven patients, the number of SSCP bands remained either the same or decreased sequentially. In those patients who did not respond [viral load >0.2 Meq/mL; n=18], the bands also ranged from 2-7; mean 3.77 +/- 1.73. In six of these non-responding patients, the SSCP bands remained the same or decreased sequentially. There was no significant difference between pretreatment quasispecies composition and response [P=.53]. Two of the four patients with pretreatment high viral load and the same or decreased composition of quasispecies bands responded to the therapy. Quasispecies in our studied patients cannot be used to predict responsiveness to treatment, but may offer an explanation for failure of most HCV-4 patients to respond to interferon alfa and ribavirin therapy

comparative study on the application of 3 molecular methods in epidemiological typing of bacterial isolates using MRSA as a prototype

To evaluate and compare 3 widely used molecular techniques, namely, restriction endonuclease analysis of plasmid deoxyribonucleic acid [REAP], randomly amplified polymorphic DNA [RAPD] and pulsed-field gel electrophoresis [PFGE] for their suitability and usefulness in the typing and fingerprinting of bacterial isolates. Twenty-four epidemiologically unrelated methicillin-resistant Staphylococcus aureus [MRSA] isolates were used to evaluate the molecular typing methods [REAP, RAPD and PFGE]. The study was conducted at the Research and Diagnostic Laboratories of King Faisal Specialist Hospital and Research Center from January 2002 through January 2003. Only 20.8% of all isolates studied were of the same genotypes by all 3 methods. Two major clusters of strains each representing 33% of the total number of isolates were identified by REAP analysis. Each of RAPD and PFGE however, identified one major cluster represented by 54% and 83% of the total number of isolates, All 3 typing methods, therefore, showed the clonal genetic relatedness among distant MRSA isolates. However inter-strain comparison of fingerprint data generated from each method revealed differences in clonal representation of the MRSA isolates. Although a variety of molecular assays are available for typing of bacterial species, there is no single standardized protocol for routine analysis. Reproducibility and interpretation of genotypic data are therefore, highly dependent on methodologies employed by the individual laboratory. Our findings illustrate the importance of using a combination of methods in typing schemes of bacterial isolates. In terms of reproducibility and typeability we found that PFGE is superior to REAP and RAPD and, therefore, more suitable for routine, standardized tracing of nosocomial bacterial isolates