Phosphorelay changes and plasticity underlie the life history evolution of Bacillus subtilis sporulation and germination in serial batch culture

Abstract

Bacterial endospores facilitate survival in extreme and unpredictably fluctuating environments. However, under abundant nutrient conditions, the production of endospores is quickly reduced or lost. We hypothesized that endospore-forming bacteria exposed to frequent cycling of nutrient availability would evolve reduced sporulation efficiency. We employed replicated batch culture for 11 transfers to test the effects of rapid nutrient cycles on the evolution of the life history traits of sporulation, germination and growth in Bacillus subtilis. We periodically measured total cell and endospore densities during the period between transfers. Replicates evolved in parallel behaviourally and genetically. By the fourth transfer, we saw a reduction in endospore production, which continued to decline throughout the experiment. Our results support a decreased likelihood of sporulation being driven by frequent nutrient renewal. The proportion of endospores germinating after transfer increased significantly by the end of the experiment through the effects of plasticity alone. Every evolved replicate culture displayed colony dimorphism: the dominant morphology being translucent with reduced sporulation ability and the rarer being opaque with accelerated sporulation and highly efficient germination. Colony dimorphism was reflected in the genomes, with all isolates with reduced sporulation having mutations in elements of the sporulation phosphorelay, particularly kinA. Some opaque colonies had no mutations, indicating that those adaptive changes occurred through plasticity. These results suggest that our selection conditions of nutrient cycling resulted in the parallel evolution of communities of ecologically diverse strains, where most reduced sporulation while a smaller proportion accelerated it.