Fluoroquinolone resistance in Helicobacter pylori: role of mutations at position 87 and 91 of GyrA on the level of resistance and identification of a resistance conferring mutation in GyrB.
OBJECTIVE
Fluoroquinolone-containing regimens have been suggested as an alternate to standard triple therapy for the treatment of Helicobacter pylori infections. To determine the relationship between fluoroquinolone resistance and mutations of GyrA and GyrB in H. pylori, we exchanged the mutations at positions 87 and 91 of GyrA among fluoroquinolone-resistant clinical isolates. GyrB of a strain with no mutations in GyrA was also analyzed to identify mechanisms of resistance to norfloxacin.


METHODS
Natural transformation was performed using the amplified fragment of the gyrA and gyrB gene as donor DNA. The amino acid sequences of GyrA and GyrB were determined by DNA sequencing of the gyrA and gyrB genes.


RESULTS
Norfloxacin-resistant strains which had mutations at position 87 and 91 became susceptible when the mutations were converted to the wild type. When the mutation from Asp to Asn at position 91 was exchanged to the mutation from Asn to Lys at position 87, the MIC to levofloxacin, gatifloxacin, and sitafloxacin increased. Norfloxacin-resistant strain TS132 with no mutations in GyrA but had a mutation at position 463 in GyrB. Transformants obtained by natural transformation using gyrB DNA of TS132 had a mutation at position 463 of GyrB and revealed resistant to norfloxacin and levofloxacin.


CONCLUSIONS
Mutation from Asn to Lys at position 87 of GyrA confers higher resistance to levofloxacin and gatifloxacin than does mutation from Asp to Asn at position 91. We propose that mutation at position 463 in GyrB as a novel mechanism of fluoroquinolone resistance in H. pylori.
Department of Microbiology, School of Pharmacy, Tokyo University of Pharmacy and Life Science, Horinouchi, Tokyo, Japan.
Emiko
E
Rimbara
Norihisa
N
Noguchi
Takashi
T
Kawai
Masanori
M
Sasatsu
D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology / Q000627:therapeutic use; D001426:Bacterial Proteins / Q000235:genetics*; D027081:DNA Gyrase / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D024841:Fluoroquinolones / Q000494:pharmacology / Q000627:therapeutic use*; D016481:Helicobacter Infections / Q000188:drug therapy* / Q000382:microbiology; D016480:Helicobacter pylori / Q000187:drug effects / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D016415:Sequence Alignment
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D024841:Fluoroquinolones; D027081:DNA Gyrase