BackgroundThe human gastric pathogen Helicobacter pylori is a paradigm for chronic bacterial infections. Its persistence in the stomach mucosa is facilitated by several mechanisms of immune evasion and immune modulation, but also by an unusual genetic variability which might account for the capability to adapt to changing environmental conditions during long-term colonization. This variability is reflected by the fact that almost each infected individual is colonized by a genetically unique strain. Strain-specific genes are dispersed throughout the genome, but clusters of genes organized as genomic islands may also collectively be present or absent.ResultsWe have comparatively analysed such clusters, which are commonly termed plasticity zones, in a high number of H. pylori strains of varying geographical origin. We show that these regions contain fixed gene sets, rather than being true regions of genome plasticity, but two different types and several subtypes with partly diverging gene content can be distinguished. Their genetic diversity is incongruent with variations in the rest of the genome, suggesting that they are subject to horizontal gene transfer within H. pylori populations. We identified 40 distinct integration sites in 45 genome sequences, with a conserved heptanucleotide motif that seems to be the minimal requirement for integration.ConclusionsThe significant number of possible integration sites, together with the requirement for a short conserved integration motif and the high level of gene conservation, indicates that these elements are best described as integrating conjugative elements (ICEs) with an intermediate integration site specificity.

Helicobacter pylori, the causative agent of type B gastritis, peptic ulcers, gastric adenocarcinoma and MALT lymphoma, uses the Cag type IV secretion system to induce a strong proinflammatory response in the gastric mucosa and to inject its effector protein CagA into gastric cells. CagA translocation results in altered host cell gene expression profiles and cytoskeletal rearrangements, and it is considered as a major bacterial virulence trait. Recently, it has been shown that binding of the type IV secretion apparatus to integrin receptors on target cells is a crucial step in the translocation process. Several bacterial proteins, including the Cag-specific components CagL and CagI, have been involved in this interaction. Here, we have examined the localization and interactions of CagI in the bacterial cell. Since the cagI gene overlaps and is co-transcribed with the cagL gene, the role of CagI for type IV secretion system function has been difficult to assess, and conflicting results have been reported regarding its involvement in the proinflammatory response. Using a marker-free gene deletion approach and genetic complementation, we show now that CagI is an essential component of the Cag type IV secretion apparatus for both CagA translocation and interleukin-8 induction. CagI is distributed over soluble and membrane-associated pools and seems to be partly surface-exposed. Deletion of several genes encoding essential Cag components has an impact on protein levels of CagI and CagL, suggesting that both proteins require partial assembly of the secretion apparatus. Finally, we show by co-immunoprecipitation that CagI and CagL interact with each other. Taken together, our results indicate that CagI and CagL form a functional complex which is formed at a late stage of secretion apparatus assembly.

Key Points• As little as 5% of kindlin-3 is sufficient to maintain basal platelet and leukocyte functions.• Platelets and neutrophils contain stoichiometric quantities of kindlin-3 and talin-1.Hematopoietic cells depend on integrin-mediated adhesion and signaling, which is induced by kindlin-3 and talin-1. To determine whether platelet and polymorphonuclear neutrophil (PMN) functions require specific thresholds of kindlin-3, we generated mouse strains expressing 50%, 10%, or 5% of normal kindlin-3 levels. We report that in contrast to kindlin-3-null mice, which die perinatally of severe bleeding and leukocyte adhesion deficiency, mice expressing as little as 5% of kindlin-3 were viable and protected from spontaneous bleeding and infections. However, platelet adhesion and aggregation were reduced in vitro and bleeding times extended. Similarly, leukocyte adhesion, extravasation, and bacterial clearance were diminished. Quantification of protein copy numbers revealed stoichiometric quantities of kindlin-3 and talin-1 in platelets and neutrophils, indicating that reduction of kindlin-3 in our mouse strains progressively impairs the cooperation with talin-1. Our findings show that very low levels of kindlin-3 enable basal platelet and neutrophil functions, whereas in stress situations such as injury and infection, platelets and neutrophils require a maximum of functional integrins that is achieved with high and stoichiometric quantities of kindlin-3 and talin-1. (Blood. 2015;126(24):2592-2600

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