PCR-BASED RESTRICTION-FRAGMENT-LENGTH-POLYMORPHISM TYPING OF HELICOBACTER-PYLORI

We applied a molecular typing approach for Helicobacter pylori that uses restriction fragment length polymorphism (RFLP) analyses of an 820-bp PCR-amplified portion of the ureC gene in H. pylori. The PCR products were digested with restriction enzyme HhaI, MboI, or MseI, and the fragments generated were analyzed by agarose electrophoresis. Among 25 independent clinical isolates, each showed a different pattern when a combination of the three RFLP patterns was used. Using this method, we studied isolates from the antrum or the body of the stomach of 14 patients before and after antibiotic therapy. Before treatment, successful isolation of H. pylori from the two sites of the stomach was possible for 12 of the 14 patients. For 10 of these 12 patients, each pair of isolates had identical RFLP profiles. For the other two patients (16.7%), however, isolates from the antrum and the body of the stomach had different RFLP profiles. Treatment was successful for 6 of the 14 patients; of the 8 patients with treatment failures, 5 had identical isolate pairs. In each case, the isolates found posttreatment were the same as the pretreatment isolates. For one of the patients who was colonized with two different isolates pretreatment, one of the isolates was identified at both sites after unsuccessful treatment. We also studied six long-term follow-up patients who had sequential biopsies at intervals of up to 5 months. Each follow-up isolate from each patient had the same RFLP profile as the initial isolate. This typing method provides a reliable and reproducible typing scheme for the study of H. pylori infections and indicates that infection with more than one H. pylori isolate is not rare.