The effect of inactivation of the 5'-GATC-3' methylase HpyIIIM in Helicobacter pylori (H. pylori) on mismatch repair, adherence, and in vitro fitness was examined. Chromosomal DNA from 90 H. pylori strains was isolated, and restriction enzyme digestion indicated all strains examined possess HpyIIIM. Wild-type H. pylori and a strain with an inactive HpyIIIM were found to have rifampicin mutation frequencies of 2.93 x 10(-7) and 1.05 x 10(-7) (P > 0.05), respectively, indicating that HpyIIIM does not appear to be important in mismatch repair. Adherence of H. pylori in an in vitro model cell system was also unaffected by inactivation of HpyIIIM. Inactivation of HpyIIIM did not result in a decrease in fitness, as determined by liquid in vitro competition experiments.