EVALUATION OF 2 SIMPLIFIED METHODS FOR GENOTYPING HEPATITIS-C VIRUS

A number of different approaches have been used for genotyping hepatit is C virus (HCV). Two simplified methods were evaluated, both of which used polymerase chain reaction (PCR) to amplify products from the 5' non-coding region of HCV: non-isotopic restriction fragment length pol ymorphism (RFLP) analysis and type-specific PCR. Sixty-four viraemic p atients suffering from chronic HCV infection were studied using these two techniques; 25/64 samples were further tested with a commercial se rotyping ELISA based on synthetic NS4 antigen (Murex, U.K.). The resul ts of the three typing methods were generally in agreement with each o ther. When only the predominant genotype identified by each method was analysed, the 3 methods had 100% agreement. RFLP did not detect any m ixed infections and it was unsuccessful in 16/64 (25%) samples. Both t ype-specific PCR and serotyping ELISA detected mixed infections. Howev er, serotyping ELISA did not give typeable results in 7/25 (28%) sampl es, whereas type-specific PCR gave typeable results in all 64 samples. Type-specific PCR detected more mixed infections than serotyping ELIS A. Direct sequencing of four PCR products with indeterminate RFLP conf irmed changes in restriction enzyme recognition sites. The sequences a lso confirmed the validity of the predominant genotype in cases of app arent mixed infections. It is possible that some of these cases were a rtefacts as a result of quasispecies. (C) 1997 Wiley-Liss, Inc.