Dark septate endophytes (DSE) protect host plants against a variety of environmental stresses, however our knowledge about the roles of DSE in improving drought tolerance of crops is poor. In this study, sorghum (Sorghum bicolor L. Moench) was inoculated with a DSE strain (Exophiala pisciphila GM25) under two different soil water conditions (well-watered (WW), -0.11 MPa; drought-stressed (DS), -0.69 MPa) for one month. At the end of this experiment, sorghum roots were obviously colonized by DSE with 50.5%-62.5% colonization rate. When compared with non-inoculated seedlings under both WW and DS conditions, E. pisciphila-inoculated sorghum had greater plant height, collar diameter, shoot dry weight, net photosynthetic rate (Pn), stomatal conductance (g s ), transpiration rate (E), maximal photochemical efficiency of PSII photochemistry (Fv/Fm) and actual quantum yield (PSII), and lower intercellular CO 2 concentration (Ci). In addition, in comparison to noninoculation under DS conditions, E. pisciphila inoculation also improved the root dry weight, non-photochemical quenching values (NPQ), photochemical quenching values (qP), increased the content of related secondary metabolites including anthocyanin, polyphenol and flavonoid and enhanced the enzymatic activities related to secondary metabolism, such as cinnamyl alcohol dehydrogenase (CAD), phenylalanine ammonia-lyase (PAL), guaiacol peroxidase (G-POD) in sorghum seedlings. Our results demonstrated that the drought resistance of sorghum seedlings were positively improved by E. pisciphila inoculation with better plant growth, gas exchange, photosynthesis, chlorophyll fluorescence, secondary metabolites and enzyme activities related to secondary metabolism. Inoculation with E. pisciphila is an efficient strategy to survive for sorghum in drought environment.