Exo-Metabolome of Pseudovibrio sp. FO-BEG1 Analyzed by Ultra-High Resolution Mass Spectrometry and the Effect of Phosphate Limitation

Abstract

Oceanic dissolved organic matter (DOM) is an assemblage of reduced carbon compounds, which results from biotic and abiotic processes. The biotic processes consist in either release or uptake of specific molecules by marine organisms. Heterotrophic bacteria have been mostly considered to influence the DOM composition by preferential uptake of certain compounds. However, they also secrete a variety of molecules depending on physiological state, environmental and growth conditions, but so far the full set of compounds secreted by these bacteria has never been investigated. In this study, we analyzed the exo-metabolome, metabolites secreted into the environment, of the heterotrophic marine bacterium Pseudovibrio sp. FO-BEG1 via ultra-high resolution mass spectrometry, comparing phosphate limited with phosphate surplus growth conditions. Bacteria belonging to the Pseudovibrio genus have been isolated worldwide, mainly from marine invertebrates and were described as metabolically versatile Alphaproteobacteria. We show that the exo-metabolome is unexpectedly large and diverse, consisting of hundreds of compounds that differ by their molecular formulae. It is characterized by a dynamic recycling of molecules, and it is drastically affected by the physiological state of the strain. Moreover, we show that phosphate limitation greatly influences both the amount and the composition of the secreted molecules. By assigning the detected masses to general chemical categories, we observed that under phosphate surplus conditions the secreted molecules were mainly peptides and highly unsaturated compounds. In contrast, under phosphate limitation the composition of the exo-metabolome changed during bacterial growth, showing an increase in highly unsaturated, phenolic, and polyphenolic compounds. Finally, we annotated the detected masses using multiple metabolite databases. These analyses suggested the presence of several masses analogue to masses of known bioactive compounds. However, the annotation was successful only for a minor part of the detected molecules, underlining the current gap in knowledge concerning the biosynthetic ability of marine heterotrophic bacteria.

Phosphate Limitation Induces Drastic Physiological Changes, Virulence-Related Gene Expression, and Secondary Metabolite Production in Pseudovibrio sp. Strain FO-BEG1

“…S6 in the supplemental material). We previously showed that P i limitation drastically affected both the amount and composition of compounds secreted by the strain during growth (20). Among these compounds, in the present work we identified the potent antibiotic TDA, which is a broad-spectrum antibiotic for which stable resistant mutants have been found to be difficult to select (69).…”

“…Strain FO-BEG1 was cultivated under ϪP i and ϩP i conditions by using the carbohydrate-mineral (CM) medium, as described previously (20). The medium contained glucose (10 mmol liter Ϫ1 ) as the only carbon source, ammonia (10 mmol liter Ϫ1 ) as the only nitrogen source, and final P i concentrations of 1.4 mmol liter Ϫ1 and 0.1 mmol liter Ϫ1 under ϩP i and ϪP i conditions, respectively.…”

Inefficient microbial production of refractory dissolved organic matter in the ocean

“…On the basis of the presence of molecular formulae, the experimental DOM was almost undistinguishable from oceanic RDOM within o2 months of incubation. This is most remarkable, because among the almost 8,000 molecular formulae identified (with the same instrumentation) in the exometabolome of a marine bacterium, a Pseudovibrio strain in monoculture, o2% were identical to DOM from the Pacific water reference sample 28 . We attribute the high number of deep-sea molecular formulae in our experiment to the high microbial diversity and the multitude of possible interactions in the mesocosms.…”

Deciphering associations between dissolved organic molecules and bacterial communities in a pelagic marine system

“…The 'species' level provided the highest canonical correlations for the DNA/DOM and the RNA/DOM analyses, which is a promising basis for further in-depth studies to elucidate functional and mechanistic relationships between these features. Transcriptomic or proteomic analyses of individual bacteria and communities (Teeling et al, 2012;Ottesen et al, 2014) alongside the identification of DOM compounds in the exometabolome (Romano et al, 2014;Schwedt et al, 2015) and marine DOM, prospectively on a structural level, are most suitable approaches for such investigations.…”