Mutations in the 16S rRNA Genes ofHelicobacter pyloriMediate Resistance to Tetracycline

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Abstract
Low-cost and rescue treatments forHelicobacter pyloriinfections involve combinations of several drugs including tetracycline. Resistance to tetracycline has recently emerged inH.pylori. The 16S rRNA gene sequences of two tetracycline-resistant clinical isolates (MIC = 64 μg/ml) were determined and compared to the consensusH.pylori16S rRNA sequence. One isolate had four nucleotide substitutions, and the other had four substitutions and two deletions. Natural transformation with the 16S rRNA genes from the resistant organisms conferred tetracycline resistance on susceptible strains. 16S rRNA genes containing the individual mutations were constructed and tested for the ability to confer resistance. Only the 16S rRNA gene containing the triple mutation, AGA965-967TTC, was able to confer tetracycline resistance onH.pylori26695. The MICs of tetracycline for the transformed strains were equivalent to those for the original clinical isolates. The two original isolates were also metronidazole resistant, but this trait was not linked to the tetracycline resistance phenotype. Serial passage of severalH.pyloristrains on increasing concentrations of tetracycline yielded mutants with only a very modest increase in tetracycline resistance to a MIC of 4 to 8 μg/ml. These mutants all had a deletion of G942 in the 16S rRNA genes. The mutations in the 16S rRNA are clearly responsible for tetracycline resistance inH.pylori.