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Xavier Nesme

Publications and source records attributed to Xavier Nesme.

6 recordsLinked to original sources
PubMed2006

Identification of genomic species in Agrobacterium biovar 1 by AFLP genomic markers.

Biovar 1 of the genus Agrobacterium consists of at least nine genomic species that have not yet received accepted species names. However, rapid identification of these organisms in various biotopes is needed to elucidate crown gall epidemiology, as well as Agrobacterium ecology. For this purpose, the AFLP methodology provides rapid and unambiguous determination of the genomic species status of agrobacteria, as confirmed by additional DNA-DNA hybridizations. The AFLP method has been proven to be reliable and to eliminate the need for DNA-DNA hybridization. In addition, AFLP fragments common to all members of the three major genomic species of agrobacteria, genomic species G1 (reference strain, strain TT111), G4 (reference strain, strain B6, the type strain of Agrobacterium tumefaciens), and G8 (reference strain, strain C58), have been identified, and these fragments facilitate analysis and show the applicability of the method. The maximal infraspecies current genome mispairing (CGM) value found for the biovar 1 taxon is 10.8%, while the smallest CGM value found for pairs of genomic species is 15.2%. This emphasizes the gap in the distribution of genome divergence values upon which the genomic species definition is based. The three main genomic species of agrobacteria in biovar 1 displayed high infraspecies current genome mispairing values (9 to 9.7%). The common fragments of a genomic species are thus likely "species-specific" markers tagging the core genomes of the species.

Bacterial Typing Techniques↗
PubMed2006

Development and validation of a prototype 16S rRNA-based taxonomic microarray for Alphaproteobacteria.

The microarray approach has been proposed for high throughput analysis of the microbial community by providing snapshots of the microbial diversity under different environmental conditions. For this purpose, a prototype of a 16S rRNA-based taxonomic microarray was developed and evaluated for assessing bacterial community diversity. The prototype microarray is composed of 122 probes that target bacteria at various taxonomic levels from phyla to species (mostly Alphaproteobacteria). The prototype microarray was first validated using bacteria in pure culture. Differences in the sequences of probes and potential target DNAs were quantified as weighted mismatches (WMM) in order to evaluate hybridization reliability. As a general feature, probes having a WMM > 2 with target DNA displayed only 2.8% false positives. The prototype microarray was subsequently tested with an environmental sample, which consisted of an Agrobacterium-related polymerase chain reaction amplicon from a maize rhizosphere bacterial community. Microarray results were compared to results obtained by cloning-sequencing with the same DNA. Microarray analysis enabled the detection of all 16S rRNA gene sequences found by cloning-sequencing. Sequences representing only 1.7% of the clone library were detected. In conclusion, this prototype 16S rRNA-based taxonomic microarray appears to be a promising tool for the analysis of Alphaproteobacteria in complex ecosystems.

Alphaproteobacteria↗
PubMed2006

Potential of a 16S rRNA-based taxonomic microarray for analyzing the rhizosphere effects of maize on Agrobacterium spp. and bacterial communities.

Bacterial diversity is central to ecosystem sustainability and soil biological function, for which the role of roots is important. The high-throughput analysis potential of taxonomic microarray should match the breadth of bacterial diversity. Here, the power of this technology was evidenced through methodological verifications and analysis of maize rhizosphere effect based on a 16S rRNA-based microarray developed from the prototype of H. Sanguin et al. (Environ. Microbiol. 8:289-307, 2006). The current probe set was composed of 170 probes (41 new probes in this work) that targeted essentially the Proteobacteria. Cloning and sequencing of 16S rRNA amplicons were carried out on maize rhizosphere and bulk soil DNA. All tested clones that had a perfect match with corresponding probes were positive in the hybridization experiment. The hierarchically nested probes were reliable, but the level of taxonomic identification was variable, depending on the probe set specificity. The comparison of experimental and theoretical hybridizations revealed 0.91% false positives and 0.81% false negatives. The microarray detection threshold was estimated at 0.03% of a given DNA type based on DNA spiking experiments. A comparison of the maize rhizosphere and bulk soil hybridization results showed a significant rhizosphere effect, with a higher predominance of Agrobacterium spp. in the rhizosphere, as well as a lower prevalence of Acidobacteria, Bacteroidetes, Verrucomicrobia, and Planctomycetes, a new taxon of interest in soil. In addition, well-known taxonomic groups such as Sphingomonas spp., Rhizobiaceae, and Actinobacteria were identified in both microbial habitats with strong hybridization signals. The taxonomic microarray developed in the present study was able to discriminate and characterize bacterial community composition in related biological samples, offering extensive possibilities for systematic exploration of bacterial diversity in ecosystems.

Cloning, Molecular↗
PubMed2002

A mathematical method for determining genome divergence and species delineation using AFLP.

The delineation of bacterial species is presently achieved using direct DNA-DNA relatedness studies of whole genomes. It would be helpful to obtain the same genomically based delineation by indirect methods, provided that descriptions of individual genome composition of bacterial genomes are obtained and included in species descriptions. The amplified fragment length polymorphism (AFLP) technique could provide the necessary data if the nucleotides involved in restriction and amplification are fundamental to the description of genomic divergences. Firstly, in order to verify that AFLP analysis permits a realistic exploration of bacterial genome composition, the strong correspondence between predicted and experimental AFLP data was demonstrated using Agrobacterium strain C58 as a model system. Secondly, a method is proposed for determining current genome mispairing and evolutionary genome divergences between pairs of bacteria, based on arbitrary sampling of genomes by using AFLP. The measure of current genome mispairing was validated by comparison with DNA-DNA relatedness data, which itself correlates with base mispairing. The evolutionary genome divergence is the estimated rate of nucleotide substitution that has occurred since the strains diverged from a common ancestor. Current genome mispairing and evolutionary genome divergence were used to compare members of Agrobacterium, used as a model of closely related genomic species. A strong and highly significant correlation was found between calculated genome mispairing and DNA-DNA relatedness values within genomic species. The canonical 70% DNA-DNA hybridization value used to delineate genomic species was found to correspond to a range of current genome mispairing of 13-13.6%. These limits correspond to 0.097 and 0.104 nucleotide substitutions per site, respectively. In addition, experimental data showed that the large Ti and cryptic plasmids of Agrobacterium had little effect on the estimation of genome divergence. Evolutionary genome divergence was used for phylogenetic inferences. Data showed that members of the same genomic species clustered consistently, as supported by bootstrap resampling. On the basis of these results, it is proposed that the genomic delineation of bacterial species could be based, in future, on phylogenetic groups supported by bootstraps and genome descriptions of individual strains, obtained by AFLP analysis, recorded in accessible databases; this approach might eventually replace DNA-DNA hybridization studies.

Biological Evolution↗
PubMed2002

Relationships between Staphylococcus aureus genetic background, virulence factors, agr groups (alleles), and human disease.

The expression of most Staphylococcus aureus virulence factors is controlled by the agr locus, which encodes a two-component signaling pathway whose activating ligand is an agr-encoded autoinducing peptide (AIP). A polymorphism in the amino acid sequence of the AIP and of its corresponding receptor divides S. aureus strains into four major groups. Within a given group, each strain produces a peptide that can activate the agr response in the other member strains, whereas the AIPs belonging to different groups are usually mutually inhibitory. We investigated a possible relationship between agr groups and human S. aureus disease by studying 198 S. aureus strains isolated from 14 asymptomatic carriers, 66 patients with suppurative infection, and 114 patients with acute toxemia. The agr group and the distribution of 24 toxin genes were analyzed by PCR, and the genetic background was determined by means of amplified fragment length polymorphism (AFLP) analysis. The isolates were relatively evenly distributed among the four agrgroups, with 61 strains belonging to agr group I, 49 belonging to group II, 43 belonging to group III, and 45 belonging to group IV. Principal coordinate analysis performed on the AFLP distance matrix divided the 198 strains into three main phylogenetic groups, AF1 corresponding to strains of agr group IV, AF2 corresponding to strains of agr groups I and II, and AF3 corresponding to strains of agr group III. This indicated that the agr type was linked to the genetic background. A relationship between genetic background, agr group, and disease type was observed for several toxin-mediated diseases: for instance, agr group IV strains were associated with generalized exfoliative syndromes, and phylogenetic group AF1 strains with bullous impetigo. Among the suppurative infections, endocarditis strains mainly belonged to phylogenetic group AF2 and agr groups I and II. While these results do not show a direct role of the agr type in the type of human disease caused by S. aureus, the agr group may reflect an ancient evolutionary division of S. aureus in terms of this species' fundamental biology.

Alleles↗