PMC:3091627 / 6394-10727 JSONTXT

{"project":"2_test","denotations":[{"id":"20537153-14573679-10781733","span":{"begin":995,"end":997},"obj":"14573679"},{"id":"20537153-16425389-10781734","span":{"begin":1212,"end":1214},"obj":"16425389"},{"id":"20537153-14742532-10781735","span":{"begin":4329,"end":4331},"obj":"14742532"}],"text":"Non-ubiquitous gene distribution in relation to associated diseases\nHybridization results for the 120 studied DNAs used as a probe and the home-made macroarrays derived from the reference strain 26695 are presented in Additional file 1 (data based on the binary presence/absence analyses) and Figure 1 (data based on the multidimensional analysis of continuous values, see material and methods). Both presentations illustrate the distribution of each of the 254 genes (213 non-ubiquitous, and 41 ubiquitous, used for normalization) with respect to associated diseases. Each strain hybridization profile (Figure 1) is represented by a series of vertically aligned bar charts, whereas the horizontal lines represent each of the 254 genes. Each strain exhibited a unique profile. The most striking features were related to the distribution of the cagPAI genes: almost all H. pylori strains associated with metaplasia harbored a complete cagPAI, a result consistent with findings by Nilsson et al. [26]. However, a complete cagPAI was present in 70% of duodenal ulcer strains, and in 50% of chronic gastritis and of MALT lymphoma strains, confirming previously published findings for isolates collected in the West [27].\nFigure 1 Hybridization reactions on a DNA macroarray membrane containing 254 PCR products that are representative of H. pylori strain 26695 (41 ubiquitous genes + 213 non-ubiquitous or strain-specific genes). Bacterial DNAs from 120 isolates involved in various diseases, including chronic gastritis (yellow), intestinal metaplasia (pink), duodenal ulcer (blue) and gastric MZBL (green), were tested by hybridization. Isolates are listed on the horizontal axis, and the genes tested, on the vertical axis. Clustering (genesis software) was carried out using the continuous values from 120 heterologous hybridization experiments, where each value corresponds to the (log26695-logheterol.strain) value for each tested gene (see materials \u0026 methods). Colors of the line range from blue, if the gene is present, to red, if absent. The range of intermediate colors reflects the degree of hybridization and thus homology, but also the redundancy of the tested genes. This figure represents the clustering based on the complete set of 254 genes. Hierarchical clustering of the continuous values derived from the hybridization experiments of 120 French clinical isolates presenting different disease characteristics was performed (Figure 1). This allowed us to visualize a branch clustering almost exclusively isolates associated with MALT lymphoma. Furthermore, principal component analysis allowed us to identify a combination of 48 genes (Additional file 1), which proved to be the most informative during multidimensional analysis. We then performed hierarchical clustering based on the values of these 48 genes (Figure 2). Two main branches were detected, one consisting of a distinct cluster of 20 isolates, all totally deprived of the cagPAI. Eighteen of the isolates were associated with MALT lymphoma and two with gastritis. Interestingly, none of the peptic ulcer or metaplasia isolates clustered in this branch. The second branch splits into two main clusters, one corresponding to isolates that totally or partially lack cagPAI genes mostly associated with gastritis and the other clustering isolates associated with other diseases.\nFigure 2 Hybridization reactions on a DNA macroarray membrane: clustering based on the 48 most discriminatory genes identified as key combinations of variables (genes/axes) from Principal Component Analysis. These 48 genes are labeled in Addional file 1. To clarify the genetic determinism of the MALT lymphoma strains, we selected one strain that was representative of the MALT lymphoma cagPAI minus branch and determined its genome sequence. We selected strain B38, which was isolated from a 62-year-old man suffering from MALT lymphoma. It fulfilled various requirements: i) it belonged to the hpEurope phylogenetic branch according to MLST analysis (Suerbaum, personal communication), a property that was consistent with the five Helicobacter genome sequences previously published (26695, J99, HPAG1, P12, and G27); ii) it was genetically transformable; iii) it was plasmid free, and iv) it was capable of colonizing the mouse gastric mucosa. Its vacA status was s2m2 [18]."}