Experimental Evolution of a Plant Pathogen into a Legume Symbiont

Marta Marchetti,Carine Gris,Catherine Masson-Boivin,Béatrice Chane-Woon-Ming,Jacques Batut,Ton Timmers,Luz B Gilbert,Claudine Médigue,Stéphane Cruveiller,Philipp Heeb,Delphine Capela,Véréna Poinsot,Michelle Glew

doi:10.1371/journal.pbio.1000280

Abstract

Author SummaryMost leguminous plants can form a symbiosis with members of a group of soil bacteria known as rhizobia. On the roots of their hosts, some rhizobia elicit the formation of specialized organs, called nodules, that they colonize intracellularly and within which they fix nitrogen to the benefit of the plant. Rhizobia do not form a homogenous taxon but are phylogenetically dispersed bacteria. How such diversity has emerged is a fascinating, but only partly documented, question. Although horizontal transfer of symbiotic plasmids or groups of genes has played a major role in the spreading of symbiosis, such gene transfer alone is usually unproductive because genetic or ecological barriers restrict evolution of symbiosis. Here, we experimentally evolved the usually phytopathogenic bacterium Ralstonia solanacearum, which was carrying a rhizobial symbiotic plasmid into legume-nodulating and -infecting symbionts. From resequencing the bacterial genomes, we showed that inactivation of a single regulatory gene allowed the transition from pathogenesis to legume symbiosis. Our findings indicate that following the initial transfer of symbiotic genes, subsequent genome adaptation under selection in the plant has been crucial for the evolution and diversification of rhizobia.

Highlights

Bacteria known as rhizobia have evolved a mutualistic endosymbiosis of major ecological importance with legumes that contributes ca. 25% of global nitrogen cycling
We show that a recipient genome—that is not immediately converted to a rhizobium upon transfer of a symbiotic plasmid— could rapidly evolve two specific symbiotic traits, i.e., nodulation and intracellular infection, under plant selection pressure
Nitrogen fixation—and mutualism—was not achieved and evolved clones could be considered as cheaters [28], evolution of nodulation and infection capacities is the first step in the evolutionary process of reciprocal cooperation [29]

Summary

Introduction

Bacteria known as rhizobia have evolved a mutualistic endosymbiosis of major ecological importance with legumes that contributes ca. 25% of global nitrogen cycling. To isolate clones expressing symbiotic potential, we took advantage of specific traits of the rhizobium–legume symbiosis, (i) legume plants act as a trap by selecting rare, nodulationproficient mutants in an otherwise non-nodulating population [17], (ii) a single bacterium enters and multiplies within the nodule [18], which implies that a rare nodulation-conferring mutation in a population is rapidly fixed, and (iii) nodulation, infection, and nitrogen fixation, are phenotypically clear-cut symbiotic stages Both the original chimera CBM124 and a gentamicin-resistant derivative, CBM124GenR, were used to repeatedly inoculate sets of ca. A few extracellular bacteria were found in nodules formed by the hrpG chimeric mutant and CBM212 and CBM349 clones (Figure 3B), which is never seen with C. taiwanensis In these cases, no plant cell wall thickening could be observed in proximity to extracellular bacteria, suggesting that they did not induce plant defence reactions. Identification of the gene(s) downstream of hrpG controlling intracellular infection should shed light to this key, but still obscure, step of the symbiotic interaction

Conclusion

Findings

Materials and Methods