Pathogenic superbugs are the root cause of untreatable complex infections with limited or no treatment options. These infections are becoming more common as clinical antibiotics have lost their effectiveness over time. Therefore, the development of novel antibacterial agents is ur-gently needed to counter these microbes. Antimicrobial peptides (AMPs) are a viable treatment op-tion due to their bactericidal potency against multiple microbial classes. AMPs are naturally selected physiological microbicidal agents that are found in all forms of organisms. In the present study, we developed two tilapia piscidin 2 (TP2)-based AMPs for antimicrobial application. Unlike the parent peptide, the redesigned peptides showed significant antimicrobial activity against multidrug-re-sistant bacterial species. These peptides also showed minimal cytotoxicity. In addition, they were significantly active in the presence of physiological salts, 50% human serum and elevated tempera-ture. The designed peptides also showed synergistic activity when combined with clinical antibiot-ics. The current approach demonstrates a fruitful strategy for developing potential AMPs for anti-microbial application. Such AMPs have potential for progression to further trials and drug development investigations.