The present study aimed at exploring mahua (Madhuca indica) oil cake as a novel substrate for the production of biosurfactant by Serratia rubidaea SNAU02 under solid-state fermentation (SSF). Response surface methodology showed followings as the optimal conditions for the production of biosurfactant: mahua oil cake 7.48 g, 2.5 ml inoculum size (1 x 10(8) cells/ml), and pH 7.22 and 31 degrees C temperature. The characterization of the biosurfactant by TLC, FT-IR and GC-MS revealed the presence of rhamnolipid. The presence of rhamnosyl transferase gene responsible for biosynthesis of rhamnolipid was identified. The strain SNAU02 exhibited antifungal activity and demonstrated no toxicity against the seeds of Brassica oleracea and Artemia salina employed as a bio-indicator. The present findings indicated the potential of mahua oil cake as suitable substrate for the production of rhamnolipids in SSF by S. rubidaea SNAU02 and application potential of the biosurfactant produced as biocontrol agent against plant pathogens. (C) 2014 Elsevier Ltd. All rights reserved.