The genome sequence of Helicobacter pylori suggests that this bacterium possesses several Fe acquisition systems, including both Fe2+‐ and Fe3+‐citrate transporters. The role of these transporters was investigated by generating insertion mutants in feoB, tonB, fecA1 and fecDE. Fe transport in the feoB mutant was ≈ 10‐fold lower than in the wild type (with 0.5 μM Fe), irrespective of whether Fe was supplied in the Fe2+ or Fe3+ form. In contrast, transport rates were unaffected by the other mutations. Complementation of the feoB mutation fully restored both Fe2+ and Fe3+ transport. The growth inhibition exhibited by the feoB mutant in Fe‐deficient media was relieved by human holo‐transferrin, holo‐lactoferrin and Fe3+‐dicitrate, but not by FeSO4. The feoB mutant had less cellular Fe and was more sensitive to growth inhibition by transition metals in comparison with the wild type. Biphasic kinetics of Fe2+ transport in the wild type suggested the presence of high‐ and low‐affinity uptake systems. The high‐affinity system (apparent Ks = 0.54 μM) is absent in the feoB mutant. Transport via FeoB is highly specific for Fe2+ and was inhibited by FCCP, DCCD and vanadate, indicating an active process energized by ATP. Ferrozine inhibition of Fe2+ and Fe3+ uptake implied the concerted involvement of both an Fe3+ reductase and FeoB in the uptake of Fe supplied as Fe3+. Taken together, the results are consistent with FeoB‐mediated Fe2+ uptake being a major pathway for H. pylori Fe acquisition. feoB mutants were unable to colonize the gastric mucosa of mice, indicating that FeoB makes an important contribution to Fe acquisition by H. pylori in the low‐pH, low‐O2 environment of the stomach.

Mycobacterium tuberculosis (Mtb) is an obligate aerobe that is capable of long-term persistence under conditions of low oxygen tension. Analysis of the Mtb genome predicts the existence of a branched aerobic respiratory chain terminating in a cytochrome bd system and a cytochrome aa3 system. Both chains can be initiated with type II NADH:menaquinone oxidoreductase. We present a detailed biochemical characterization of the aerobic respiratory chains from Mtb and show that phenothiazine analogs specifically inhibit NADH:menaquinone oxidoreductase activity. The emergence of drug-resistant strains of Mtb has prompted a search for antimycobacterial agents. Several phenothiazines analogs are highly tuberculocidal in vitro, suppress Mtb growth in a mouse model of acute infection, and represent lead compounds that may give rise to a class of selective antibiotics. Mycobacterium tuberculosis ͉ respiratory chainT he World Health Organization estimates that two billion people are infected with Mycobacterium tuberculosis (Mtb), and two million people die of the disease each year (1). Most individuals infected with the organism are latent carriers who have a 2-23% lifetime risk of developing reactivation tuberculosis (TB). The risk dramatically increases if the carrier's immune system is suppressed. Also, drug resistance is a serious concern; the isoniazid (INH)-resistance rate is Ϸ10%, and the rifampicin (RIF) resistance rate is Ϸ1%, with lower numbers in countries with effective TB programs and higher numbers in countries with deficient TB programs. The World Health Organization declared TB infections to be a global public health emergency (1), and the need to identify targets for antimicrobial therapy remains urgent.Mtb is capable of establishing persistent infection in the host by using a complex interplay between the host immune system and bacterial survival mechanisms. In the persistent infection, Mtb adapt to depletion of available oxygen and nutrients and enter a stage of nonreplicating persistence (NRP) in granulomatous or necrotic lesions. NRP Mtb are resistant to INH, ethambutol, and RIF, but they become sensitive to metronidazole in vitro (2). Given the critical role of oxygen in the generation of cellular energy and bacterial long-term survival, there is surprisingly little information on oxidative phosphorylation in Mtb. Clearly, oxidative phosphorylation is a central component in the production of ATP and the subsequent growth and pathogenesis of Mtb. Here, we characterize the aerobic respiratory pathway and show that NADH:menaquinone oxidoreductase (Ndh) is a key target for TB agents. Materials and MethodsMedia and Strains. Mtb H 37 R v was a gift from C. Imperatrice (Clinical Infectious Diseases, Hospital of the University of Pennsylvania) and Mycobacterium smegmatis Mc 2 155 was obtained from V. Mizrahi (National Health Laboratory Service, Johannesburg). Bacteria were cultured in 7H9 broth supplemented with 10% oleic acid-albumin-dextrose catalase͞0.5% glycerol͞0.05% Tween 80. Solid agar (15 g͞liter) was ad...

The lipopolysaccharide (LPS) of Helicobacter pyloriexpresses the Lewis x (Lex) and/or Ley antigen. We have shown previously that H. pylori LPS displays phase variation whereby an Lex-positive strain yields variants with different LPS serotypes, for example, Lex plus Ley or nonfucosylated polylactosamine. H. pylori has two α3-fucosyltransferase genes that both contain poly(C) tracts. We now demonstrate that these tracts can shorten or lengthen randomly, which results in reversible frameshifting and inactivation of the gene products. We provide genetic and serological evidence that this mechanism causes H. pylori LPS phase variation and demonstrate that the on or off status of α3-fucosyltransferase genes determines the LPS serotypes of phase variants and clinical isolates. The role of the α3-fucosyltransferase gene products in determining the LPS serotype was confirmed by structural-chemical analysis of α3-fucosyltransferase knockout mutants. The data also show that the two α3-fucosyltransferase genes code for enzymes with different fine specificities, and we propose the names futA and futB to designate the orthologs of the H. pylori 26695 α3-fucosyltransferase genes HP0379 and HP0651, respectively. The data also show that the α3-fucosylation in H. pylori precedes α3-fucosyltransferase, an order of events opposite to that which prevails in mammals. Finally, the data provide an understanding at the molecular level of the mechanisms underlying LPS diversity in H. pylori, which may play an important role in adaptation to the host.

Ever since the realization that Helicobacter pylori was intimately associated with the development of gastritis and peptic ulcer disease in humans, there has been a need for a simple animal model in which modes of pathogenicity, transmission, immunization, and chemotherapeutic intervention can be evaluated. Whereas small animals such as mice and rats are particularly well suited as experimental hosts for many infections, early studies suggested that H. pylori had a very narrow host range that did not extend to these species. Although many attempts to infect small laboratory animals with H. pylori were apparently made, these proved generally unsuccessful (1,2) and the view became established rapidly that "H. pylori will not colonize many of the usual laboratory animal species, including conventionally reared rats, mice, rabbits, guinea pigs, specific-pathogen-free pigs, colostrum-deprived piglets, and gnotobiotic rats and mice" (3). An apparent exception was the claim that H. pylori would colonize. Mongolian gerbils particularly after gastric lesions were produced by indomethacin (4); however, this work has never been substantiated nor followed up. Instead, most attention was paid subsequently to the use of naturally occurring Helicobacter mustelae infections of ferrets (5,6), experimental challenge with H. pylori in gnotobiotic piglets (7) and the important development of the Helicobacter felis model in mice and rats (8,9).

Infection with mycobacterial species, including