Rebecca J Hall (@drrebeccajhall.bsky.social)

Metabolite toxicity as a driver of bacterial metabolite externalization Some microbes externalize costly biosynthetic precursors in sufficient quantities to sustain a recipient population through cross-feeding. However, it is unclear whether metabolites are externalized purely for a reciprocal benefit or if metabolite externalization also plays a physiological role for the producer. Here we focus on adenine, a metabolite externalized by some strains of the phototrophic bacterium Rhodopseudomonas palustris at sufficient levels to support Escherichia coli growth. In 10 long-term monocultures and 22 cocultures pairing R. palustris with E. coli, extracellular adenine externalized by all 140 isolates screened was 1.7 - 3.4-fold higher than that by the ancestor, suggesting that there was selective pressure for adenine externalization. We hypothesized that adenine is toxic to R. palustris. The CGA0092 growth rate decreased by half in the presence of about 0.3 mM external adenine. This inhibitory effect increased by an order of magnitude when we over-expressed adenine phosphoribosyltransferase to overcome a bottleneck in the purine salvage pathway, suggesting that toxicity stems from a metabolite derived from adenine. To assess whether adenine tolerance is connected to adenine externalization, we surveyed 12 evolved isolates and 49 environmental strains that externalized different levels of adenine, revealing a significant positive correlation. Our data suggests a physiological role for externalization of costly-metabolites like adenine at the origin of cross-feeding. In addition to cross-feeding, resulting metabolic interactions could be negative, considering that even a biosynthetic precursor like adenine can be inhibitory. ### Competing Interest Statement The authors have declared no competing interest. United States Army Research Office, https://ror.org/05epdh915, W911NF-14-1-0411, W911NF-17-1-0159 U.S. National Science Foundation, MCB-1749489 https://www.biorxiv.org/content/10.64898/2026.01.17.700106v1