The Commelina forskaolii Vahl (Commelinaceae) is an edible herb with wide therapeutic value. The present study aimed to explore the preliminary phytochemical screening, GC-MS spectrometry analysis, FTIR, and evaluate the antimicrobial activity of C. forskaolii Vahl leaves using various solvent extracts with different polarities tested for their inhibitory property against most prevalent aquatic pathogenic bacteria (Gram-positive and Gramnegative) and fungi by the agar diffusion method. The results of phytochemical and GC-MS analysis of the different solvent extracts revealed the occurrence of secondary metabolites and totally 35 biologically active compounds. The FTIR results confirmed the presence of alkanes, amines, alcohol, aryl disulfides, ethers, hydroxy group, vinylidene, chloro, bromo, and iodo compounds. The in vitro antimicrobial activity of five solvent extracts of C. forskaolii Vahl was investigated against Streptococcus agalactiae and Streptococcus iniae, Aeromonas hydrophila, Pseudomonas aeruginosa, Vibrio cholerae, Aspergillus niger and Aspergillus flavus. Among all extracts, aqueous and chloroform extract showed maximum antibacterial activity against all bacterial pathogens based on the different concentrations, which inhibited in the range of 11 mm to 18 mm (aqueous), and 11 mm to 27 mm (chloroform) followed by inhibition range of other extracts 9 mm to 12 mm (ethanol), 11 mm to 23 mm (methanol), and 12 mm to 16 mm (acetone). In addition, the highest antifungal activity was reported in chloroform extract against Aspergillus niger and aqueous extract against Aspergillus flavus with inhibition of 16 mm followed by 17 mm at 75 µl concentration. In the present investigation, C. forskaolii Vahl leaves were examined and reported for the first time and the study suggests that leaves of C. forskaolii Vahl could be potentially used as a natural source of antimicrobial agents in aquaculture and fisheries industries.