Experimental Evolution in Plant-Microbe Systems: A Tool for Deciphering the Functioning and Evolution of Plant-Associated Microbial Communities

Abstract

In natural environments, microbial communities must constantly adapt to stressful environmental conditions. The genetic and phenotypic mechanisms underlying the adaptive response of microbial communities to new (and often complex) environments can be tackled with a combination of experimental evolution and next generation sequencing. This combination allows to analyse the real-time evolution of microbial populations in response to imposed environmental factors or during the interaction with a host, by screening for phenotypic and genotypic changes over a multitude of identical experimental cycles. Experimental evolution (EE) coupled with comparative genomics has indeed facilitated the monitoring of bacterial genetic evolution and the understanding of adaptive evolution processes. Basically, EE studies had long been done on single strains, allowing to reveal the dynamics and genetic targets of natural selection and to uncover the correlation between genetic and phenotypic adaptive changes. However, species are always evolving in relation with other species and have to adapt not only to the environment itself but also to the biotic environment dynamically shaped by the other species. Nowadays, there is a growing interest to apply EE on microbial communities evolving under natural environments. In this paper, we provide a non-exhaustive review of microbial EE studies done with systems of increasing complexity (from single species, to synthetic communities and natural communities) and with a particular focus on studies between plants and plant-associated microorganisms. We highlight some of the mechanisms controlling the functioning of microbial species and their adaptive responses to environment changes and emphasize the importance of considering bacterial communities and complex environments in EE studies.

Keywords: evolutionary adaptation; experimental evolution; holobiont; interaction network; microbiota; synthetic community.

FIGURE 1

Comparison of the potential outcomes from experimental evolution (EE) studies starting with (A) a single clone or (D) a microbial community. (A) Example of an experimental evolutionary design based on a single clone serially cultivated in a simple medium, a stressful or a changing environment; those experiments enable to track phenotypic and genetic changes over time. (B) Theoretical Muller diagram depicting the distinct genotypes frequencies over microbial generations that could be observed in EE study described in A. (C) Theoretical phylogenetic tree built from genotypes that can be sampled from different time points throughout the EE described in A and sampled at the same generation times. (D) Example of an experimental evolutionary design based on an inoculum made up of several microorganisms (synthetic or natural communities); those experiments make it possible to monitor changes in population levels (E), and genetic, transcriptomic or metabolomic changes over time (F). (E) Theoretical bar graph illustrating the composition of a microbial community showing differences in taxa among both the inoculum (ancestral mix) and different experimental cycles. (F) At each cycle, approaches could be implemented at the scale of the whole community (i.e., meta-omics) or of individual cells after cell-sorting (single-cell sequencing, transcriptomic).

FIGURE 2

Relationships between plant and microbial components. (A) Plants interact with a wide diversity of microorganisms (microbiota) in the different compartments of the plant system. The rhizosphere corresponds to the soil zone where the roots impact the soil organization and microbial functioning, while the rhizoplane is the interface between the root surface and the soil. The phyllosphere corresponds to the surface of all the aerial organs of the plant. Root exudates attract or repel soil microorganisms toward roots and increase the growth of myriads of microorganisms that will interact between each other (positive, negative or neutral biotic interactions). (B) Major biotic interactions in the rhizosphere include plant-microorganism interactions and microorganism/microorganism interactions, involving the exchange of different classes of molecules (hormones, toxins, virulence factors, signals, etc.).