Metabolomic signatures of pathogen suppression effect of Baltic eelgrass meadows in surrounding seawater

Organic molecules exuded into water column by marine organisms represent a significant portion of marine dissolved organic matter (DOM) that modulates biochemical interactions. Secreted allelochemicals have been suggested to be involved in regulation of pathogen abundance in seagrass meadows, however, seagrass exometabolome has remained unstudied. We aimed to identify seagrass exometabolites, within and outside meadows, and explore their potential involvement in pathogen suppression under varying environmental conditions. We collected seawater (SW) samples from eelgrass (Zostera marina)-vegetated (V) and non-vegetated (NV) areas across 5 locations spanning 270 km of coastline along the German Baltic Sea. Comparative LC-MS/MS-based untargeted computational metabolomics combined with statistical analyses and machine learning tools were employed to pinpoint (exo)metabolomic signatures of eelgrass leaves. Simultaneously, we measured abiotic parameters and the abundance of three common pathogenic taxa in seawater, and investigated spatiotemporal variations. Here we show the correlation of pathogen biomass and eelgrass pathogen reduction effect with increasing seawater temperature, eutrophication and anthropogenic influences. Exometabolomics studies revealed that eelgrass exudates contributed significantly to overall seawater DOM at molecular level, while SW overlying eelgrass meadows contained many chemical features unique to the eelgrass leaf metabolome. We identified four flavone aglycones as key biomarkers distinguishing SW-V and SW-NV samples. Their drastically increased concentrations correlated with the lowest pathogen biomass, suggesting their role in pathogen regulation. These combined analytical and microbiological approaches indicate that flavones are defensive allelochemicals released into eelgrass meadows upon environmental stress and serve as potential bioindicators of eelgrass' sanitation effect.