Diabetic foot ulcer infections are the main causes of hospitalization in diabetics. The present study aimed to develop vancomycin and imipenem/cilastatin loaded core-shell nanofibers to facilitate the treatment of diabetic foot ulcers. Therefore, novel core-shell nanofibers composed of polyethylene oxide, chitosan, and vancomycin in shell and polyvinylpyrrolidone, gelatin, and imipenem/cilastatin in core compartments were prepared using the electrospinning technique. The nanofibers were characterized using scanning electron microscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, tensile test, and drug release. The antibacterial activity of drug-loaded nanofibers in different drugs concentrations was evaluated against Methicillin-resistant Staphylococcus aureus (MRSA), Escherichia coli, and Pseudomonas aeruginosa by disk diffusion method. Furthermore, the cytotoxicity of fibers was investigated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay. The obtained results showed that the prepared nanofibers were smooth having a core-shell structure with almost no cytotoxicity. The nanofibrous mats exhibited significant antibacterial activity against S. aureus and MRSA with the inhibition zones of 2.9 and 2.5 cm and gram-negative bacteria species of E. coli and P. aeruginosa with the inhibition zones of 1.9 and 2.8 cm, respectively. With respect to the significant antibacterial activities of these nanofibrous mats, they might be used as suitable drug delivery devices not only for diabetic foot ulcer infections but also for other chronic wounds.