Role of Hpn and NixA of Helicobacter pylori in susceptibility and resistance to bismuth and other metal ions.
BACKGROUND
Helicobacter pylori produces Hpn, a 60-amino acid, histidine-rich protein that avidly binds nickel and zinc ions, and NixA, a high-affinity nickel transporter in the cytoplasmic membrane. We tested the hypothesis that Hpn and NixA govern susceptibility to metal ions in H. pylori.


METHODS
Hpn-negative mutants of four H. pylori strains were constructed by standard allelic exchange techniques to yield isogenic Hpn+/Hpn-deficient pairs. A metal concentration that inhibited growth by 50% (IC50) was calculated for Ni2+, Zn2+, Cu2+, and Co2+ by comparing OD600 of cultures in metal-supplemented and control media.


RESULTS
Among all four pairs of isogenic strains, the tolerance for Ni2+ was reduced significantly (p <.001) in the Hpn mutants; the mean IC50 value for wild-type strains was 1.9 mM; for the mutant, it was 0.8 mM. In contrast, growth inhibition by Zn2+ was identical within the fours pairs, as was Cu2+ and Co2+ tolerance in one pair tested. We also found that deletion of the hpn gene increases susceptibility to therapeutic forms of bismuth by testing a mutant and wild-type pair with ranitidine bismuth citrate, bismuth citrate, and four antibiotics. Minimal inhibitory concentrations of ranitidine bismuth citrate dropped from 9.2 to 2.3 microg/ml, and those of bismuth citrate dropped from 7.4 to 3.2 microg/ml (p <.05 for both comparisons), while susceptibility to the antibiotics was unaffected. Disruption of the nixA gene encoding the specific Ni2+ transport protein of H. pylori did not change susceptibility to bismuth.


CONCLUSIONS
We concluded that bacteria lacking Hpn, cultured in vitro, are more susceptible than is the wild type to bismuth and Ni2+.
Mobley
H L
HL
Department of Microbiology and Immunology, University of Maryland School of Medicine, Baltimore 21201, USA. hmobley@umaryland.edu
Garner
R M
RM
Chippendale
G R
GR
Gilbert
J V
JV
Kane
A V
AV
Plaut
A G
AG
D001426:Bacterial Proteins; D001729:Bismuth / Q000494:pharmacology*; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D027682:Cation Transport Proteins; D004352:Drug Resistance, Microbial; D016481:Helicobacter Infections / Q000382:microbiology; D016480:Helicobacter pylori / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D019216:Metals, Heavy / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D009942:Organometallic Compounds / Q000494:pharmacology; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011899:Ranitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D000099053:Histidine-Rich Glycoprotein
D001426:Bacterial Proteins; D002352:Carrier Proteins; D027682:Cation Transport Proteins; D008565:Membrane Proteins; D019216:Metals, Heavy; C095257:NixA protein, Helicobacter pylori; D009942:Organometallic Compounds; D011506:Proteins; D000099053:Histidine-Rich Glycoprotein; C073340:ranitidine bismuth citrate; D011899:Ranitidine; C002791:bismuth tripotassium dicitrate; D001729:Bismuth