In silico proteomic and phylogenetic analysis of the outer membrane protein repertoire of gastric Helicobacter species.

Helicobacter (H.) pylori is an important risk factor for gastric malignancies worldwide. Its outer membrane proteome takes an important role in colonization of the human gastric mucosa. However, in zoonotic non-H. pylori helicobacters (NHPHs) also associated with human gastric disease, the composition of the outer membrane (OM) proteome and its relative contribution to disease remain largely unknown. By means of a comprehensive survey of the diversity and distribution of predicted outer membrane proteins (OMPs) identified in all known gastric Helicobacter species with fully annotated genome sequences, we found genus- and species-specific families known or thought to be implicated in virulence. Hop adhesins, part of the Helicobacter-specific family 13 (Hop, Hor and Hom) were restricted to the gastric species H. pylori, H. cetorum and H. acinonychis. Hof proteins (family 33) were putative adhesins with predicted Occ- or MOMP-family like 18-stranded ß-barrels. They were found to be widespread amongst all gastric Helicobacter species only sporadically detected in enterohepatic Helicobacter species. These latter are other members within the genus Helicobacter, although ecologically and genetically distinct. LpxR, a lipopolysaccharide remodeling factor, was also detected in all gastric Helicobacter species but lacking as well from the enterohepatic species H. cinaedi, H. equorum and H. hepaticus. In conclusion, our systemic survey of Helicobacter OMPs points to species and infection-site specific members that are interesting candidates for future virulence and colonization studies.

A novel isolation protocol and probe-based RT-PCR for diagnosis of gastric infections with the zoonotic pathogen Helicobacter suis.

BACKGROUND: Helicobacter suis is a very fastidious microorganism associated with gastritis, gastric ulcers, and mucosa-associated lymphoid tissue lymphoma in humans. In vitro isolation of this agent from human patients has so far been unsuccessful. MATERIALS AND METHODS: A probe-based real-time PCR (RT-PCR) for the rapid detection of H. suis in gastric biopsies was developed. Secondly, a mouse-passage-based protocol was optimized for isolation of low numbers of viable H. suis bacteria. Mice were inoculated with different numbers of viable H. suis (102 -108 ) and kept for 4 weeks to allow multiplication of this pathogen. RESULTS: The probe-based real-time PCR (RT-PCR) exhibited a high degree of diagnostic specificity and analytical sensitivity, high linear correlations (r2 between 0.995 and 0.999), and high amplification efficiencies (>90%) for H. suis. No cross-reactivity was detected with human, porcine, non-human primate, and murine DNA nor with DNA from other bacteria including Helicobacter spp. and Campylobacter spp. H. suis was successfully re-isolated from the stomach of mice inoculated with at least 104 viable H. suis, using a biphasic medium (pH 5), consisting of Brucella agar with Brucella broth on top, both supplemented with vitox supplement, Campylobacter-selective supplement, amphotericin (5 µg/mL), HCl (0.05%), fetal bovine serum (20%), and linezolid (5 µg/mL). Linezolid was necessary to inhibit proliferation of contaminants, including lactobacilli. CONCLUSION: The methods described above can be implemented for detection or isolation of H. suis from human gastric biopsies.

Divergence between the Highly Virulent Zoonotic Pathogen Helicobacter heilmannii and Its Closest Relative, the Low-Virulence "Helicobacter ailurogastricus" sp. nov.

Helicobacter heilmannii naturally colonizes the stomachs of dogs and cats and has been associated with gastric disorders in humans. Nine feline Helicobacter strains, classified as H. heilmannii based on ureAB and 16S rRNA gene sequences, were divided into a highly virulent and a low-virulence group. The genomes of these strains were sequenced to investigate their phylogenetic relationships, to define their gene content and diversity, and to determine if the differences in pathogenicity were associated with the presence or absence of potential virulence genes. The capacities of these helicobacters to bind to the gastric mucosa were investigated as well. Our analyses revealed that the low-virulence strains do not belong to the species H. heilmannii but to a novel, closely related species for which we propose the name Helicobacter ailurogastricus. Several homologs of H. pylori virulence factors, such as IceA1, HrgA, and jhp0562-like glycosyltransferase, are present in H. heilmannii but absent in H. ailurogastricus. Both species contain a VacA-like autotransporter, for which the passenger domain is remarkably larger in H. ailurogastricus than in H. heilmannii. In addition, H. ailurogastricus shows clear differences in binding to the gastric mucosa compared to H. heilmannii. These findings highlight the low-virulence character of this novel Helicobacter species.