The aim of this investigation was to perform an extensive screening using HPLC and I3C-NMR spectroscopy to disclose the spectrum of osmolytes produced by aerobic heterotrophic and anoxygenic phototrophic eubacteria. The most predominant solutes detected within a wide range of marine and halophilic micro-organisms were two recently discovered tetrahydropyrimidines ectoine and hydroxyectoine, which were synthesized in response to osmotic stress.

A new family, the Hafomonadaceae, has recently been proposed for members of the genera Deleya and Halornonas. The three strains investigated, Deleya haiuphila, Halomonas elongata and Flavobacterium halmephilum (reclassified as H. hahophila), are aerobic heterotrophic micro-organisms exhibiting an extreme salt tolerance. The major organic osmoregulatory solutes of these organisms were examined using 3C-nuclear magnetic resonance spectroscopy. The relative proportions of the solutes varied with respect to salt concentration, temperature and carbon source. The recently described amino acid ectoine was found to be a dominant solute. For the first time it could be shown for halophilic eubacteria that the intracellular concentration of solutes is sufficient to balance the osmotic pressure of the medium. Thus, there is no need to postulate a hypo-osmotic cytoplasm.

Many molecular details of the ecophysiology of halophilic bacteria that use compatible solutes to maintain osmotic equilibrium have been examined. We ask whether the details are consistent and complete enough to predict growth and osmoregulation in these bacteria by integrating this information in a mathematical model. Parameterized for the halophilic organism Halomonas elongata, the model predicts the substrate and salt dependence of growth, the uptake of potassium and ectoine and the synthesis of ectoine. It is shown that salt (NaCl) dependence of growth can be modelled by substrate inhibition kinetics. Osmoregulation is known to involve accumulation of both ectoine and potassium glutamate in H. elongata. Using published and newly determined parameters, osmoregulatory models using either direct turgor or two-step (turgor and potassium) signalling are compared. The results are consistent with a role for potassium as a second messenger for hyperosmotic stress. Simulations of osmotic upshifts show a transient overregulation of the intracellular solute levels, as has been previously observed in experiments. A possible adaptive value of this overregulation as 'pre-emptive' behaviour in an environment with frequent dry periods leading to steadily increasing osmolarity is proposed. As a result of growth parameter estimation, a maximum P : O value of 2 for H. elongata can be inferred. In conclusion, the model developed here reproduces essential aspects of growth and osmoregulation in halophilic bacteria with a minimal set of assumptions.

In the first column, the subheading for the third group of organisms should read Proteobacteria (y-subclass). p. 1632, column 2, line 12: for position 3 read position 5. p. 1634, column 1, line 2 : for glycosylglycerin read glycosylglycerol. p. 1635, column 2, line 16: for Firmacutes read Firmicutes.

Using high performance liquid chromatography and nuclear magnetic resonance techniques, the compatible solutes of some moderately halophilic bacteria were studied. The following accepted species of moderately halophilic bacteria were included: Volcaniella eurihalina and Deleya salina among Gram‐negative rods, and Salinicoccus roseus and Salinicoccus hispanicus among Gram‐positive cocci. Besides these strains we have also screened other new isolates, including Marinomonas species and Gram‐positive cocci and rods. The tetrahydropyrimidine carboxylic acid ‘ectoine’ was found to be the main compatible solute in the Gram‐negative strains tested when these were grown in glucose‐mineral medium. In addition, betaine was accumulated from complex media containing yeast extract. Among the Gram‐positive strains investigated, the solutes proline (bacillus 30, Salinicoccus) and hydroxyectoine (coccus 28) also played an important role, while alanine, glucose, glutamate, glutamine and trehalose occurred as minor components. We also detected two recently described compatible solutes: Nδ‐acetylornithine and a homologous compound, Nε‐acetyllysine. Representatives of distinct phenotypic groups of Gram‐positive cocci and rods were clearly distinguished by their solute pattern.