The molecular mechanism of H. pylori resistance to clarithromycin is associated with A-to-G substitution within the peptidyltransferase-encoding region of the 23S rRNA gene [7,8]. The clarithromycin MICs ranged from 2 to >256 μg ml^-1 for the 52 strains included in this study. Either the A2142 mutation or the A2143G mutation were present in 94.2% of clarithromycin-resistant H. pylori strains examined. The majority of isolates (82.7%) contained the A2142G mutation, whereas the A2143G was present in 11.5% (Table ​(Table1).1). Additionally, when an MIC of ≥ 64 μg ml^-1 was used to define a high level of clarithromycin resistance, differences in prevalence of each mutation were observed (Table ​(Table1).1). There was a relationship between the presence of the A2142G mutation and the highest MIC values (p = 0.01).

Some authors have shown that the A2143G mutation was the most frequently detected mutation [19,20], however, our data and those of others demonstrate that the A2142G mutation is more prevalent in Brazil [8,21]. Although the presence of the A2143G mutation was rare in the present study and the A2142G mutation demonstrated the highest frequency observed, the clinical relevance of this distinction has a limited value, since the presence of either of these mutations is able to confer clarithromycin resistance [22]. A high level of resistance to clarithromycin has been associated with the presence of the A2142G mutation in H. pylori [13,23]. According to our results, a greater effect upon MIC was observed among strains possessing the A2142G mutation. Thus, this mutation may incur a greater probability of treatment failure in populations infected by H. pylori [13].

In three isolates, the PCR-RFLP method was unable to detect these substitutions. The sequence analysis of these isolates, from the internal fragment that comprises the majority of point mutations related to clarithromycin resistance, revealed only a T-to-C transition at position 2245 (Based on GenBank accession no. U27270). To determine whether the T2245C mutation observed was responsible for clarithromycin resistance, the 26695 cla^S H. pylori strain was transformed with genomic DNA and specific PCR products. For this purpose, the 23S rRNA was amplified using primers, 18 and 21 [8], while internal primers, Cla1995 and Cla2274, created a 280 bp fragment that only contained the T2245C substitution. The transformants were selected on BHI-YE plates supplemented with clarithromycin. Although the T2245C substitution was present in these three resistant strains, clarithromycin-resistant transformants containing the T2245C substitution were only observed after transformation with genomic DNA, but not after transformation with 23S rRNA PCR fragments. While transformants were found after transformation with DNA and PCR products from the BZ586 cla^R strain, none of the transformants were selected after transformation with DNA from the cla^S strain 26695 or TE. The MIC values of clarithromycin of 10 randomly selected resistant transformants was determined by agar dilution and were identical to those of donor strains.