Simulated drought with Polyethylene-Glycol (PEG) decreases above-ground performance and increases nodulation in the legume Medicago lupulina

Under climate change, understanding how plants and crops respond to drought is essential for basic research in ecology and evolution, and improving agricultural resilience. One common method of simulating drought in experimental conditions is by applying polyethylene glycol (PEG) to plants. We investigated drought growth responses in Medicago lupulina (black medic) using PEG to simulate drought stress. We grew Medicago lupulina plants inoculated with Sinorhizobium meliloti in Magenta boxes under controlled conditions and randomly assigned them to one of three treatments: a control, PEG applied to the bottom (PEG added to the bottom-watering container of a magenta box), or PEG applied from the top (PEG poured over the growth media). After 60 days, we measured true leaf number, nodule count, and below- and above-ground dry biomass. PEG treatments significantly reduced above-ground growth, including total biomass and leaf number, but unexpectedly increased nodulation. Our results suggest that while PEG effectively simulates drought stress on above-ground growth parameters, it may not accurately simulate drought effects on rhizobial symbiosis. PEG treatments had no effect on below-ground biomass, suggesting that increased nodulation is not a result of increased plant investment in below-ground growth under simulated drought. We hypothesize that PEG, as a persistent liquid that plants do not absorb, created conditions favorable for nodulation. Overall, these results highlight the importance of interpreting PEG-simulated drought experiments with caution when assessing mutualistic interactions.