Enhanced nutrient uptake is sufficient to drive emergent cross-feeding between bacteria in a synthetic community

Interactive microbial communities are ubiquitous, influencing biogeochemical cycles and host health. One widespread interaction is nutrient exchange, or cross-feeding, wherein metabolites are transferred between microbes. Some cross-fed metabolites, such as vitamins, amino acids, and ammonium (NH₄⁺), are communally valuable and impose a cost on the producer. The mechanisms that enforce cross-feeding of communally valuable metabolites are not fully understood. Previously we engineered a cross-feeding coculture between N₂-fixing Rhodopseudomonas palustris and fermentative Escherichia coli. Engineered R. palustris excretes essential nitrogen as NH₄⁺ to E. coli, while E. coli excretes essential carbon as fermentation products to R. palustris. Here, we sought to determine whether a reciprocal cross-feeding relationship would evolve spontaneously in cocultures with wild-type R. palustris, which is not known to excrete NH₄⁺. Indeed, we observed the emergence of NH₄⁺ cross-feeding, but driven by adaptation of E. coli alone. A missense mutation in E. coli NtrC, a regulator of nitrogen scavenging, resulted in constitutive activation of an NH₄⁺ transporter. This activity likely allowed E. coli to subsist on the small amount of leaked NH₄⁺ and better reciprocate through elevated excretion of fermentation products from a larger E. coli population. Our results indicate that enhanced nutrient uptake by recipients, rather than increased excretion by producers, is an underappreciated yet possibly prevalent mechanism by which cross-feeding can emerge.