bioRxiv2026
Bacterial adaptation to fluctuations in salinity includes the intracellular accumulation of organic compounds called compatible solutes (CS) such as the amino acid derivatives ectoine and 5-hydroxyectoine. These compounds also play a less appreciated role as readily available nutrients, scavenged from dissolved organic matter in both marine and terrestrial environments. Vibrio diabolicus is a marine bacterium originally isolated from deep-sea hydrothermal vents and later shown to have worldwide distribution. In this work, we demonstrated the biosynthesis and uptake of CS ectoine and glycine betaine under high osmotic stress conditions, but not in unstressed V. diabolicus cells. A region on chromosome 1 of V. diabolicus strain 3098 encoded homologues of genes for ectoine and 5-hydroxyectoine catabolism (eutDE, ssd_atf_eutBCA), regulation (asnC, enuR), and transport (ectoine TRAP-type uehP, uehQM). Our data showed that ectoine was used as a high energy yielding sole carbon source and the eutD gene was essential for ectoine consumption. Phylogenetics based on EutD (DoeA) and gene neighborhood analyses showed that a catabolism cluster was present in Proteobacteria, Thermosulfobacteriota, Bacillota, Actinomycetota, and Archaea. The cluster had a limited phylogenetic distribution in Gammaproteobacteria and Betaproteobacteria and was widespread in Alphaproteobacteria. Phylogenetic reconstruction was consistent with vertical inheritance with gene loss with repeated horizontal acquisitions of the pathway across lineages. The ectoine catabolism pathway was vertically inherited in Halomonadaceae and Vibrionaceae, with patterns of gene and pathway loss. Betaproteobacteria Burkholderia, Caballeronia, and Paraburkholderia EutD proteins clustered together and EutD from most Pseudomonas species shared a most recent common with this group. EutD from Alphaproteobacteria branched in eight divergent clusters with long branch lengths but showed a remarkable conservation of synteny. Catabolism and transporter genes in this group were contiguous and contained either a TRAP-type UehPQM or an ABC-type EhuABCD ectoine transporter. Gram-positive bacteria and Archaea were not previously shown to consume ectoines, however, we identified putative ectoine catabolism clusters among Bacilli, Clostridia, Actinomycetes, and Halobacteria. IMPORTANCEEctoine is a well-established CS used to overcome osmotic stress, produced by a wide range of bacteria. The demonstration of ectoine biosynthesis and catabolism in V. diabolicus showed that it is conditionally utilized as an osmoprotectant or a nutrient source depending on environmental cues. The conservation of large syntenic blocks of ectoine catabolism, transport, and regulatory genes suggested strong selective pressure to maintain this trait. EutD (DoeA) phylogeny patterns largely followed taxonomy with evidence of horizontal transfer in specific clades, and showed ectoine consumption has a broad taxonomic spread and is lineage enriched. In our dataset, Alphaproteobacteria contained the largest diversity of EutD lineages; Gammaproteobacteria from marine environments formed a strong secondary group; and EutD from Betaproteobacteria were the least diverse. Many species that contained EutD are associated with saline, marine, and plant-associated niches, where ectoine can be scavenged as a nutrient source. The identification of a putative ectoine catabolism pathway in Gram-positive bacteria and Archaea needs to be experimentally confirmed and suggests undiscovered diversity to be revealed by future genome sequencing.