Abstract:

The cyst nematodes of the Globodera genus are telluric organisms that exclusively parasite the Solanaceous botanic family. During compatible interaction, the second stage larvae hatch in the soil and move towards the apical parts of the roots where they penetrate. They then migrate intracellularly to the central cylinder. They elaborate an intimate relationship called syncytium which allows them to feed and complete their life cycle. This structure is the result of a genetic reprogramming of plant cells. The molecular bases of the plant/nematode interaction are relatively unknown but could largely improve our knowledge on the phytoparasitic nematodes. We first chose to characterise new pathogenicity genes. The description of the variability of these genes seemed to be necessary to understand their evolution. This approach should make it possible to identify the selective constraints to which they are subjected and which define to some extent the adaptive capacities of some species, populations or genera.

We isolated three new pathogenicity genes in Globodera pallida for which transcripts were localised in salivary glands, corresponding to proteins secreted by the nematode in the plant. Among these genes, we highlighted for the first time in G pallida a RanBPM homolog, Gp-rbp-1, specifically expressed at the pre parasitic and parasitic stages which lead to the development of the syncytium. Southern blot results suggest that this gene belongs to a multigenic family. The extent of the variability of this gene family was studied by screening a G. pallida cDNA library: 12 different RanBPM copies were sequenced and showed a 52.7% nucleotidic variability. Preliminary analysis of the substitutions positions for the three genes highlighted a high mutation rate on first and second codon position (~ 50%) suggesting a high proportion of non synonymous substitutions. However, the analyses will have to be carried out on a broader data set, as we did for two other genes for which we had more indications on their implication in parasitism.

Among many genes of the literature, we chose one gene involved in the early steps of parasitism, the pectate lyase gene, and one involved in the late stages, the cathepsin L gene. The elongation factor 1a, a priori not related to the parasitism, was used as a comparison. We amplified these three genes in 40 populations of cyst nematodes. After checking of the orthology of the sequences, we have shown that the same variability was observed whatever the gene considered, even if the calculated genetic distances showed an increase of the evolutionary tempo on parasitism genes. Substitutions observed in the first two positions of the codon were much more important for parasitism genes (50%) that for the housekeeping gene (20%). Sharp Analyses of the selection pressures made it possible to show that particular populations and species showed evolution rates more important that the others, even if we do not have any biological explanation for that. Moreover, we showed that the cathepsin L gene possesses a domain subjected to positive selection, indicating a strong evolutionary potential of this gene.

Thesis abstract

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