Rewinding the tape: experimental evolution of resistance to glyphosate

Modern agriculture relies on chemical weed control to secure crop yields, but extensive and prolonged herbicide use has imposed strong selective pressures, driving the evolution of resistance in weeds and contaminating non-target ecosystems. Understanding the evolutionary dynamics and outcomes of herbicide selection is critical for addressing thesechallenges. Microbial model organisms, such as the green alga Chlamydomonas reinhardtii, provide an ideal system to study resistance evolution in a controlled laboratory setting, and represents an important group of nontarget organisms exposed to herbicide pollution.This thesis examines the effects of glyphosate selection on adaptation in C. reinhardtii. Specifically, the study focused on the evolutionary dynamics, fitness, and genomic changes associated with exposure of C. reinhardtii to this herbicide, linking resistance phenotypes to genomic changes under contrasting selection regimes. While most research has focused on sudden exposure to high herbicide doses, gradual dose increases can occur due to the accumulation of herbicides in the environment, with potentially distinct evolutionary impacts. Using experimental evolution and whole genome sequencing, adaptive responses were investigated under two ecological scenarios. The first experiment involved a simple scenario with a single rapid glyphosate dose increase, while the second tested a complex scenario with variable changes in dose rates (rapid, intermediate, and slow rates of change).The findings from these experiments revealed that rapid dose increases augmented variability and delayed resistance evolution, whereas gradual changes imposed fitness costs. Genome-wide variant detection indicated that glyphosate resistance was not associated with mutations or copy number variation in the target enzyme 5 enolpyruvylshikimate-3-phosphate synthase (EPSPS). Instead, evidence suggests that glyphosate resistance in C. reinhardtii is driven by non-target-site mechanisms, regardless of selection history. Furthermore, limited overlap in resistance-associated loci between treatments and replicates suggests a complex, and potentially independent, evolutionary response to glyphosate selection. These insights contribute to our understanding of the evolutionary and genomic mechanisms driving herbicide resistance