Heart rot disease, caused by Lasiodiplodia theobromae, is destructive for date palms and other woody plants. The disease was reported in several oases in Egypt, and the pathogen was found in association with infected trees suffering dieback and rachis blight. Seven phylogenetically distinct fungal isolates were selected, and their pathogenicity was confirmed on date palms. The isolates exhibited variable degrees of virulence on inoculated leaves, which confirms the variation. We examined the antifungal effect of microbial bioagents and plant extracts on heart rot disease. The isolates of Trichoderma spp. gave moderate reduction of the pathogen’s linear growth (40 to 60%), whereas their exudates were ultimately ineffective. Bacillus spp. isolates, except for B. megaterium, were more effective against spore germination, giving 80 to 90% reduction on average. Among the examined plant extracts, garlic sap gave 98.67% reduction of linear growth followed by artemisia (15.5%) and camphor (24.8%). The extraction methods greatly influenced the antifungal efficiency of each extract because exposure to organic solvents significantly decreased the efficiency of all extracts, whereas hot water extraction negatively affected garlic sap only. Successful bioagents and plant extracts were further assayed for the suppression of heart rot disease on date palms. Both T. album and T. harzianum gave comparable degrees of suppression as by commercial fungicides. In addition, treatment before or during pathogen inoculation was the most effective because it significantly enhanced the expression of defense-related enzymes. Our findings suggest biopesticides possess a dual role in disease suppression and defense boosters for date palms suffering heart rot disease.