Abstract
Many Helicobacter pylori (Hp) strains carry cryptic plasmids of different size and gene content, the function of which is not well understood. A subgroup of these plasmids (e.g. pHel4, pHel12), contain a mobilisation region, but no cognate type IV secretion system (T4SS) for conjugative transfer. Instead, certain H. pylori strains (e.g. strain P12 carrying plasmid pHel12) can harbour up to four T4SSs in their genome (cag-T4SS, comB, tfs3, tfs4). Here, we show that such indigenous plasmids can be efficiently transferred between H. pylori strains, even in the presence of extracellular DNaseI eliminating natural transformation. Knockout of a plasmid-encoded mobA relaxase gene significantly reduced plasmid DNA transfer in the presence of DNaseI, suggesting a DNA conjugation or mobilisation process. To identify the T4SS involved in this conjugative DNA transfer, each individual T4SS was consecutively deleted from the bacterial chromosome. Using a marker-free counterselectable gene deletion procedure (rpsL counterselection method), a P12 mutant strain was finally obtained with no single T4SS (P12ΔT4SS). Mating experiments using these mutants identified the comB T4SS in the recipient strain as the major mediator of plasmid DNA transfer between H. pylori strains, both in a DNaseI-sensitive (natural transformation) as well as a DNaseI-resistant manner (conjugative transfer). However, transfer of a pHel12::cat plasmid from a P12ΔT4SS donor strain into a P12ΔT4SS recipient strain provided evidence for the existence of a third, T4SS-independent mechanism of DNA transfer. This novel type of plasmid DNA transfer, designated as alternate DNaseI-Resistant (ADR) mechanism, is observed at a rather low frequency under in vitro conditions. Taken together, our study describes for the first time the existence of three distinct pathways of plasmid DNA transfer between H. pylori underscoring the importance of horizontal gene transfer for this species.
Highlights
Helicobacter pylori is a highly motile, microaerophilic, Gramnegative bacterium, resident in the gastric mucus layer of about 50% of the human population
A remarkable feature of H. pylori is its panmictic population structure, reflected by an extreme genetic heterogeneity, possibly resulting from frequent recombination events after import of small pieces of foreign DNA from other H. pylori strains during persistent or transient mixed infections [3,4,5,6]. Such an efficient DNA exchange has been attributed to the natural transformation competence of H. pylori [7], mediated by the comB type IV transport system [8,9,10], which is stimulated by DNA damage to trigger genetic exchange [11]
A subgroup of H. pylori plasmids has been described that carries putative mobilisation genes [13,17,20,26], which might be involved in conjugative transfer between different H. pylori strains
Summary
Helicobacter pylori is a highly motile, microaerophilic, Gramnegative bacterium, resident in the gastric mucus layer of about 50% of the human population. A remarkable feature of H. pylori is its panmictic population structure, reflected by an extreme genetic heterogeneity, possibly resulting from frequent recombination events after import of small pieces of foreign DNA from other H. pylori strains during persistent or transient mixed infections [3,4,5,6]. Such an efficient DNA exchange has been attributed to the natural transformation competence of H. pylori [7], mediated by the comB type IV transport system [8,9,10], which is stimulated by DNA damage to trigger genetic exchange [11]. Chromosomal integration and excision of plasmid DNA after transfer from a donor into a recipient strain might be an alternative way to generate genome rearrangements [13]
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