Fifty nine isolates belonging to six species of Enterococci namely, Enterococcus faecalis, Enterococcus faecium, Enterococcus raffinosus, Enterococcus durans, Enterococcus mundtii and Enterococcus avium (n = 35, 15, 4, 3, 1 and 1 isolates, respectively) were obtained from different clinical specimens including urine, pus, blood, wound, sputum and synovial fluid. The highest numbers of enterococci were recorded from the pus (20 isolates, 33.90%) followed by urine (12 isolates, 20.34%) while the lowest frequency was observed with synovial fluid samples (2 isolates, 3.39%). These isolates showed different multidrug resistant patterns with the lowest resistant for linezolid (n = 5, 8.48%), followed by teicoplanin (n = 14, 23.73%) and vancomycin (n = 20, 33.90%) while they exhibited the highest resistant against penicillin (n = 53, 89.83%), oxacillin (n = 50, 84.75%), erythromycin (n = 49, 83.05%) and streptomycin (n = 47, 79.66 %). On the other hand, a free living marine bacterium under isolation code ESRAA3010 was isolated from seawater samples obtained from the fishing area Masturah, Red Sea, Jeddah, Saudi Arabia. The phenotypic, chemotaxonomic, 16S rRNA gene analyses and phylogenetic data proved that isolate ESRAA3010 is very close to Bacillus subtilis and then it was designated as Bacillus subtilis ESRAA3010. It gave the highest antagonistic activity against all clinical Enterococcus faecalis, Enterococcus faecium, Enterococcus raffinosus, Enterococcus durans, Enterococcus mundtii and Enterococcus avium isolates under study with minimum inhibitory concentration (MIC) ranged from 4 to 56 µg/mL, 4 to 12 µg/mL, 4 to 8 µg/mL, 4 to 8 µg/mL, 8 µg/mL and 4 µg/mL, respectively as well as minimum bactericidal concentration (MBC) (8 to 64 µg/mL, 4 to 16 µg/mL, 4 to 12 µg/mL, 4 to 16 µg/mL, 12 µg/mL and 8 µg/mL, respectively). Moreover it showed anti-proliferative activity against colon (HCT-116), liver (HepG-2), breast (MCF-7) and lung (A-549) carcinomas with IC50 equal to 39, 50, 75 and 19 µg/mL, respectively which indicates its prospective usage in the upcoming decades.