Adaptive Evolution of Geobacter sulfurreducens in Coculture with Pseudomonas aeruginosa

Interactions between microorganisms in mixed communities are highly complex, being either syntrophic, neutral, predatory, or competitive. Evolutionary changes can occur in the interaction dynamics between community members as they adapt to coexistence. Here, we report that the syntrophic interaction between and coculture change in their dynamics over evolutionary time. Specifically, sp. dominance increases with adaptation within the cocultures, as determined through quantitative PCR and fluorescence hybridization. This suggests a transition from syntrophy to competition and demonstrates the rapid adaptive capacity of spp. to dominate in cocultures with Early in coculture establishment, two single-nucleotide variants in the and genes emerged that were strongly selected for throughout coculture evolution with phenazine wild-type and phenazine-deficient mutants. Sequential window acquisition of all theoretical spectra-mass spectrometry (SWATH-MS) proteomics revealed that the variant cooccurred with the upregulation of an adenylate cyclase transporter, CyaE, and a resistance-nodulation-division (RND) efflux pump notably known for antibiotic efflux. To determine whether antibiotic production was driving the increased expression of the multidrug efflux pump, we tested -derived phenazine-1-carboxylic acid (PHZ-1-CA) for its potential to inhibit growth and drive selection of the and genetic variants. Despite its inhibitory properties, PHZ-1-CA did not drive variant selection, indicating that other antibiotics may drive overexpression of the efflux pump and CyaE or that a novel role exists for these proteins in the context of this interaction. and spp. cohabit many of the same environments, where spp. often dominate. Both bacteria are capable of extracellular electron transfer (EET) and play important roles in biogeochemical cycling. Although they recently in 2017 were demonstrated to undergo direct interspecies electron transfer (DIET) with one another, the genetic evolution of this syntrophic interaction has not been examined. Here, we use whole-genome sequencing of the cocultures before and after adaptive evolution to determine whether genetic selection is occurring. We also probe their interaction on a temporal level and determine whether their interaction dynamics change over the course of adaptive evolution. This study brings to light the multifaceted nature of interactions between just two microorganisms within a controlled environment and will aid in improv