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The role of strain heterogeneity in Helicobacter pylori colonisation, virulence and host adaptation
thesisposted on 09.02.2017, 05:31 by Grubman, Alexandra
The remarkable ability of Helicobacter pylori to colonise the mammalian gastric mucosa is facilitated by conserved strategies that promote strain diversification, allowing the bacteria to continually adapt to changing conditions encountered in this harsh niche. These mechanisms include rapid mutation, highly developed recombination pathways and an abundance of genes that can be regulated by slipped-strand mispairing, which all contribute to the ability of H. pylori to elicit vastly different pathologies in individual hosts. The aims of this study were to investigate how subtle diversity between closely related H. pylori strains affects virulence and adaptation of H. pylori to new hosts. We focused our study on the progenitor and derivatives of the “gold standard” mouse-colonising H. pylori SS1 strain, and investigated the phenotypic effects of transcriptional variation and genome sequence diversity. We determined that minor transcriptional differences between in vitro passaged derivative strains of H. pylori SS1 resulted in significant alterations to the capacity of H. pylori SS1 to colonise mice. Using macroarray comparisons, we identified two vitamin B6 biosynthesis enzymes, PdxA and PdxJ, as being downregulated in low-infectivity SS1 isolates. Significantly, this report demonstrates that besides its recognized metabolic functions, vitamin B6 is required for colonisation of mice by H. pylori, which is likely to be attributable to the role of Vitamin B6 in normal flagellar gly- cosylation and assembly, as well as for motility, a key virulence factor in H. pylori. Furthermore, we determined a role for vitamin B6 salvage pathways in H. pylori colonisation and characterised a novel multifunctional pyridoxal kinase, HP0844, which is involved in vitamin B6 salvage. Our data demonstrate, for the first time, an important in vivo role for pathogen-specific de novo vitamin B6 biosynthesis. In order to investigate the impact of genetic variability on H. pylori virulence, we performed next generation sequencing to compare H. pylori strains differing in their ability to induce inflammatory responses in human gastric cell lines. For this, we used the mouse-adapted H. pylori SS1 and its progenitor, clinical isolate 10700. We identified a total of 3366 single nucleotide polymorphisms (SNPs) as having arisen during host-adaptation of H. pylori 10700 to the murine gastric mu- cosa. We hypothesised that some of these SNPs may correlate with the impaired function in H. pylori SS1 of a key H. pylori virulence factor, the cag pathogenicity island (cagPAI)-encoded type 4 secretion system (T4SS). Importantly, two of these SNPs occurred within the non-cagPAI encoded proteins, ProV and HP0958, that we demonstrate to be essential for T4SS functionality. As loss of virulence factors in vivo potentially influences disease outcome in infected hosts, these findings also provide additional virulence markers for H. pylori strain typing. Moreover, our data demonstrate that mouse adaptation of H. pylori 10700 has resulted in specific intimate interactions with the murine gastric epithelium, independent of the cagPAI, that enable H. pylori SS1 to effectively persist within the host. Lastly, we identified the involvement of an alternative nuclear factor-κB (NF-κB) sub- unit, c-Rel, in murine-specific responses to H. pylori. Taken together, our data highlight the role of genome plasticity during adaptation in vivo allowing for host selection of strains that most efficiently overcome the challenges to colonisation of individual gastric environments.