Research PaperMetabolic responses of endophytic Nicotiana benthamiana plants experiencing water stress

•Endophytes cause differential accumulation of metabolites in plants undergoing water stress.•Endophyte infection accumulates certain compounds in plants regardless of water status.•Fungal endophytes increase plant root mass and relative water content under stress.

Endophytic fungal colonization may influence how plants respond to environmental stress. Two promising fungal isolates, one resembling Cladosporium cladosporioides and another unidentified ascomycetous fungus, isolated from wild N. benthamiana plants in northern Australia were inoculated to plants of the research accession of N. benthamiana (RA-4). Inoculated seedlings were grown under adequate or water deficit conditions. We examined leaf metabolites using gas chromatography-mass spectrometry (GC-MS) to compare levels of sugars, sugar alcohols, amino acids and other metabolites at various stages of plant growth and stress application. Ninety-three metabolites were detected in leaves, including 20 sugars, 13 sugar alcohols, 21 amino acids, 29 organic and fatty acids and ten other compounds. Endophyte colonization caused significantly differential accumulation of 17-21 metabolites when the plants were grown under well-watered condition. The presence of endophytes under water stress conditions caused differential accumulation of cytosine, diethylene glycol, galactinol, glycerol, heptadecanoate, mannose, oleic acid, proline, rhamnose, succinate, and urea. Accumulation of these metabolites suggests that fungal endophytes influence plants to accumulate certain metabolites under water-stress. Further, plants colonised by the two different endophytes tested, showed some differences in the metabolites they accumulated. Colonization with endophytic fungi significantly increased root dry mass and relative water content in plants under severe water stress condition, suggestive of a symbiotic relationship between these fungi and N. benthamiana plants, a species adapted to the hot and unpredictable soil moisture conditions of northern Australia. We reveal that endophyte colonization triggers reprogramming of host metabolism and indices changes in host development. This study sheds lights on the mechanisms underlying increased tolerance to water stress in plants conferred by fungal endophytes. Fungal endophytes have the potentials for application to increase the inherent water stress tolerance of crops.