Abstract
ABSTRACTBeneficial plant root-associated microorganisms carry out a range of functions that are essential for plant performance. Establishment of a bacterium on plant roots, however, requires overcoming several challenges, including competition with neighboring microorganisms and host immunity. Forward and reverse genetics have led to the identification of mechanisms that are used by beneficial microorganisms to overcome these challenges, such as the production of iron-chelating compounds, the formation of strong biofilms, or the concealment of characteristic microbial molecular patterns that trigger the host immune system. However, how such mechanisms arose from an evolutionary perspective is much less understood. To study bacterial adaptation in the rhizosphere, we employed experimental evolution to track the physiological and genetic dynamics of root-dwelling Pseudomonas protegens in the Arabidopsis thaliana rhizosphere under axenic conditions. This simplified binary one plant/one bacterium system allows for the amplification of key adaptive mechanisms for bacterial rhizosphere colonization. We identified 35 mutations, including single-nucleotide polymorphisms, insertions, and deletions, distributed over 28 genes. We found that mutations in genes encoding global regulators and in genes for siderophore production, cell surface decoration, attachment, and motility accumulated in parallel, underlining the finding that bacterial adaptation to the rhizosphere follows multiple strategies. Notably, we observed that motility increased in parallel across multiple independent evolutionary lines. All together, these results underscore the strength of experimental evolution in identifying key genes, pathways, and processes for bacterial rhizosphere colonization and a methodology for the development of elite beneficial microorganisms with enhanced root-colonizing capacities that can support sustainable agriculture in the future.
Highlights
Erqin Li and Hao Zhang contributed to this article
We previously studied five experimental-evolutionary populations, referred to as lines, of CHA0 evolving in the rhizosphere of Arabidopsis in a gnotobiotic system
Independent populations were introduced on the roots, and after 4 weeks of plant growth, the populations were transferred to new plants
Summary
Erqin Li and Hao Zhang contributed to this article. The author order was determined by contributions to writing the manuscript. Bacterial genes that are involved in the colonization of plant roots can be revealed by identifying beneficial mutations that evolve during adaption of bacteria to the rhizosphere environment and have positive effects on root colonization. As the frequency of each mutation (Fig. 3; Table 2) was determined from only six bacterial colonies that were isolated and sequenced from each evolutionary line at cycles 2, 4, and 6, we further investigated and accurately measured the populationlevel frequency at the end of each experimental cycle.
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