The use of medicinal plants in marginal communities was for the treatment of various ailments for centuries. Nevertheless, the potential of endophytic fungi (EF) associated with bioprospecting medicinal plants remains understudied. Research on the diversity of EF associated with various Egyptian medicinal plants remains limited. Therefore, our study conducted an analysis and comparison of the colonization frequency (CF), richness, and diversity indices of EF communities that inhabit nine different medicinal plants located in two different areas: the Protected Area of Wadi Degla in Maadi and the Natural Cultivated Area in Helwan, Egypt. These plants were Agathophora alopecuroides, Anabasis setifera, Atriplex halimus, Halocnemum strobitaceum, Lantana camara, Mesembryanthemum forsskaollii, Raphanus raphanistrum, Suaeda vermiculata, and Zygophyllum coccineum. Also, the antimicrobial and antioxidant potential of isolated EF has been investigated. A total of 39 morphospecies EF were isolated and identified, belonging to fifteen genera. Aspergillus spp. and Penicillium spp. were the dominant genera identified in the selected plants. A. setifera and S. vermiculata plants had the highest numbers of EF isolates, followed by M. forsskaollii and R. raphanistrum. Furthermore, these plants had a significant diversity index and species richness compared to other plants investigated. The most predominant EF was Aspergillus sp.3, which had the highest occurrence rate. Among all EF ethyl acetate extracts (EAEs), Aspergillus sp.3 demonstrated the highest antimicrobial activities against different human pathogenic bacteria, yeasts, and fungi. Furthermore, it showed the highest 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity. Therefore, this isolate was reidentified molecularly as Aspergillus terreus AUMC16223 with accession number PP491988. Moreover, EAE of A. terreus endophyte showed cytotoxicity potential activity with the significant IC value of 41.75 ± 1.83 µg/mL for the human lung carcinoma cell line (A549) and a nontoxic effect on the normal cell line (WI 38) with the significant CC value of 196.2 ± 3.74 µg/mL. Our results indicated diverse EF communities associated with different Egyptian medicinal plants, showing A. terreus endophyte extract as the most significant antimicrobial, antioxidant, and cytotoxic agent.