RAPID FINGERPRINTING OF HELICOBACTER-PYLORI BY POLYMERASE CHAIN-REACTION AND RESTRICTION-FRAGMENT-LENGTH-POLYMORPHISM ANALYSIS

A simple and reliable technique was developed for differentiating Heli cobacter pylori strains by restriction fragment length polymorphism an alysis of polymerase chain reaction (PCR)-amplified DNAs. Oligonucleot ide primer pairs developed to the urease, 48-kDa stress protein (htrA) , and 26-kDa antigen-encoding genes were used to amplify fragments of the appropriate size from crude boiled cell preparations. The PCR-ampl ified products were digested with Sau3A, HaeIII, MspI, AluI, MluI, Hin fI, and XbaI restriction endonucleases. Restriction fragment length po lymorphisms were particularly evident within the urease and htrA genes and were easily detected by Sau3A, HaeIII, MspI, and AluI restriction endonuclease analysis. Double digestion of these separately amplified products or restriction analysis of multiple PCR-amplified fragments was found to discriminate 17 of 17 (100%) H. pylori strains which had unique genomic DNA fingerprints. Results of an investigation of multip le isolate sets obtained from patients before and after therapy was co nsistent with the hypothesis that treatment failures were due to the p ersistence of the same strain but did not discount the possibility tha t the patients were reinfected with a strain shared by family members or close contacts. The results indicate that the PCR-restriction endon uclease analysis method can be applied directly to biopsy samples, has the potential to fingerprint H. pylori isolates rapidly, and may perm it detailed epidemiological investigations on the transmission of this important pathogen.