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Genomic analysis of a metronidazole resistant  H. bizzozeronii  strain revealed a frame length extension of the oxygen-insensitive NAD(P)H-nitroreductase HBZC1_00960 (RdxA), associated with the disruption of the C-terminal cysteine-containing conserved region (IACLXALGK). This was the result of the extension (from C 8  to C 9 ) of a simple sequence cytosine repeat (SSCR) located in the 3’ of the gene. A 3' SSCR is also present in the  rdxA  homolog of  H. heilmannii  sensu stricto, but not in  H. pylori . We showed that in the majority of in vitro spontaneous  H. bizzozeronii  metronidazole resistant mutants, the extension of the 3′ SSCR of  rdxA  was the only mutation observed. In addition, we observed that  H. bizzozeronii  Δ rdxA  mutant strain showed the same MIC value of metronidazole observed in the spontaneous mutants. These data indicate that loss of function mutations in  rdxA  and in particular the disruption of the conserved region IACLXALGK is associated with reduced susceptibility to metronidazole in  H. bizzozeronii . Slipped-strand mispairing of the SSCR located in the 3′ of the  H. bizzozeronii rdxA  appears to be the main mechanism. We also observed that  H. bizzozeronii  acquires resistance to metronidazole at high mutation rate, and that serial passages in vitro without selection induced an increased level of susceptibility. In conclusion, contrary to what was previously described in  H. pylori , the  H. bizzozeronii rdxA  appears to be a contingency gene which undergoes phase variation. The contingency nature of  rdxA  should be carefully considered when metronidazole is used in the treatment of  H. heilmannii -associated gastritis.

Contingency nature of Helicobacter bizzozeronii oxygen-insensitive NAD(P)H-nitroreductase (HBZC1_00960) and its role in metronidazole resistance