Helicobacter spp. are common causes of bacterial infection worldwide in both humans and animals.The canine Gram-negative Helicobacter bizzozeronii is one out of seven species in the group of H. heilmannii sensu lato (s. l.) also known as non-H. pylori gastric Helicobacter species (NPGHS) that are detected in 0.2-6% in gastric biopsies of human patients with gastritis. At present, H. bizzozeronii and H. felis are the only H. heilmannii s. l. species that have been successfully cultivated from human gastric biopsies. In contrast to human-to-human transmission described for H. pylori, the H. heilmannii sensu lato (s. l). are transmitted from animals to humans (zoonotic). To date, an understanding of the pathogenesis of non-pylori Helicobacter infections in humans and also of the mechanisms of their adaptation are lacking. In order to provide new insights into these topics, the present study focused on the molecular biology of H. bizzozeronii with particular interest in unravelling the bacterial features involved in host-jumping and adaptation to the human gastric mucosa. The study also investigated the molecular mechanisms of metronidazole resistance in H. bizzozeronii.
Using comparative genome analysis between H. pylori and a human isolate of H. bizzozeronii CIII-1, we investigated several molecular genetic features that could explain the zoonotic nature of NPGHS. In particular, we propose the hypothesis that the high metabolic versatility and the ability to react to a range of environmental signals, factors which differentiate H. bizzozeronii and other species belonging to H. heilmannii s. l. from H. pylori, are the molecular basis of the of the zoonotic nature of H. heilmannii s. l. infection in humans. Further we characterized the biosynthesis of lipopolysaccharide (LPS) of H. bizzozeronii in order to understand the factors involved in host adaptation during host jump from dogs to humans. As a major Gram-negative cell surface component, LPS has the potential to interact with certain surface components of the host s cells during infection, which modifies the inflammatory response and promotes progress of chronic infection. Our results showed that H. bizzozeronii expresses sialyl-LPS (containing the sialyl-lactoseamine epitope), which is a common characteristic among canine and human H. bizzozeronii strains. The expression of sialyl-glycans undergoes phase- and antigen variation and may influence the adaptation process of H. bizzozeronii during the transmission from dogs to humans.
Metronidazole, in combination with other antibiotics, is frequently used for the eradication of H. pylori and H. heilmannii s. l. infections but resistance to this antibiotic is a serious, increasing problem and a major cause of treatment failure. In this study, we performed comparative genomic analysis between antrum derived H. bizzozeronii populations that had been isolated from biopsy samples from a female Finnish patient who was suffering from chronic gastritis. The biopsies were taken three months before (T0) and six months after (T1) an unsuccessful eradication treatment. Our results that H. bizzozeronii had undergone genome diversification in order to colonize the patient s stomach successfully. We hypothesised that antimicrobial treatment induced a sudden decrease of H. bizzozeronii population size but effectively favoured the selection of a subpopulation, which subsequently acted as the founder of a new population that was characterized by higher numbers of fixed mutations. H. bizzozeronii isolates obtained after the treatment underwent a frame length extension of the oxygen-insensitive NAD(P)H-nitroreductase HBZC1_00960 (homologue of H. pylori RdxA) that is associated with the disruption of the C-terminal cysteine-containing conserved region (IACLXALGK). This disruption was the result of the extension (from C8 to C9) of a simple sequence cytosine repeat (SSCR) (homopolymeric tract) that is located in the 3' region of the rdxA gene. Further we investigated the role of rdxA gene in the molecular mechanisms of metronidazole in H. bizzozeronii. Our results showed that, in most of the in vitro spontaneous H. bizzozeronii metronidazole resistant mutants, the only observed mutation was that of the extension of the 3' SSCR of rdxA. Another major observation was that the MIC value of metronidazole for both spontaneous mutant isolates and an H. bizzozeronii ΔrdxA mutant strain were similar, which indicates that a loss of rdxA function is associated with the reduced susceptibility of H. bizzozeronii to metronidazole. Another key finding was that H. bizzozeronii acquires resistance to metronidazole at high mutation rates and that serial passages in vitro without selection induce reversibility of the resistant phenotype. These results suggest that the potential contingency nature of rdxA should be carefully considered if metronidazole is administered for the treatment of H. heilmannii-associated gastritis.
The results of the present studies contribute substantially in the understanding of the zoonotic nature of non-H. pylori gastric Helicobacter species and of the molecular mechanisms behind successful colonization and acquired antimicrobial resistance. Further studies are required in order to elucidate the specific roles of characterized factors, such as polysaccharide lyase and Sialyl-LPS, in the development of gastritis and MALT lymphoma.
|Tila||Julkaistu - 2013|
|OKM-julkaisutyyppi||G5 Tohtorinväitöskirja (artikkeli)|
- 413 Eläinlääketiede