Biocontrol of okra-rot-causing Cochliobolus spicifer-CSN-20 using secondary metabolites of endophytic fungi associated with Solenostemma arghel

Abstract

Rot disease is responsible for serious economic losses related to okra (Abelmoschus esculentus) crops cultivated in Upper Egypt. Colonies with a consistent morphology were isolated from the infected okra stems and leaves and subjected to morphological and molecular examinations. The causal pathogen was identified as Cochliobolus spicifer based on morphological fungus descriptions, as well as on the amplified 28S rDNA and internal transcribed spacer region (ITS) sequences, which showed 99%–100% similarity to the sequences of C. spicifer-CSN-20 strains. The volatile and non-volatile organic compounds (VOCs and n-VOCs, respectively) produced by the endophytic fungi that are associated with the medicinal plant Solenostemma arghel, namely Fusarium solani-F4-1007, Penicillium verrucosum-F2-1006, and Aspergillus terreus-F5-1008, inhibited the growth of the C. spicifer pathogen by 34.2%,31.4%, and 30.5%, respectively. In total,27 VOCs were identified by GC/MS, among which eight were specific to A. terreus-F5-1008, eight to P. verrucosum-F2-1006, and three to F. solani-F4-1007, whereas nine VOCs were commonly produced by the three endophytic fungi. Moreover, F. solani-F4-1007-produced VOCs and n-VOCs exhibited the highest antifungal activity, with 37.27% and 37.1% inhibition against C. spicifer colony growth, respectively. The potent antifungal VOCs produced by F. solani-F4-1007 were identified as 3,4-dihydro-2 h-1,5-(3″-t-butyl) benzodioxepine, 4-(2-hydroxyethyl) phenol, and phenylethyl alcohol using GC/MS. Therefore, F. solani-F4-1007 was tested as a potential biocontrol agent against C. spicifer-CSN-20 using an in-planta assay. Okra plants treated with endophytic F. solani-F4-1007did not show any disease symptoms, whereas those that were not treated with F. solani-F4-1007 exhibited severe disease symptoms when challenged with inoculation of the C. spicifer pathogen. Our results demonstrated the contribution of the endophytic fungus F. solani-F4-1007 as a potential biocontrol agent against the C. spicifer pathogen, to improve okra growth.