The main aim of the current research is to achieve active targeting of antimicrobials in secondary urinary tract infections using the direction of renal blood flow. The objective is further extended to maintain a constant internal environment and to deliver an extended maintenance dose using Poly lactic acid (PLLA) electrospun nanofibers. The medicated nanofibers used to evaluate their physical characteristics and drug release behaviour. Therapeutic efficacy of the developed formulation was achieved by organ distribution, pharmacokinetic and challenge study. A major fraction of drug was recovered from the kidney after 32 hours results in a significant reduction in colony forming unit (CFU) count establish the therapeutic efficacy of developed formulation. Polyanionic nature of PLLA electrospun nanofibers assists as an excellent carrier for intravaginal delivery of antimicrobial in urinary tract infections.