Low levels of metabolic auxotrophy among environmental Pseudomonas isolates

Auxotrophy, the inability of bacteria to synthesize one or multiple essential metabolites (e.g. amino acids, vitamins, metabolites) is thought to be common among bacteria. However, studies often rely either on bioinformatic tools to predict auxotrophies from genome data or on experiments with low numbers of strains. Here, we combine experimental and bioinformatic approaches to assess amino acid auxotrophy levels among 315 co-isolated natural Pseudomonas strains from pond and soil habitats. Both approaches revealed that Pseudomonas isolates are predominantly prototrophs. We identified one single histidine auxotroph and five non-specific auxotrophs featuring complex growth phenotypes incompatible with single amino acid auxotrophies. While different bioinformatic pipelines vary in the extent to which auxotrophy is over- or underestimated, none of the pipelines could resolve the basis of non-specific auxotrophies. Our analysis further revealed the existence of multiple alternative biosynthesis pathways for methionine, proline, and phenylalanine, with significant enrichments of specific pathways among soil or pond strains. We conclude that combining experiments with bioinformatics is a powerful approach to assess the metabolic potential of environmental bacteria. Moreover, taxa like Pseudomonas can be predominantly prototrophic possibly owing to their generalist lifestyle, thus calling for nuanced ecological concepts predicting auxotrophy levels based on lifestyle and habitat.