Background: Among chronic bacterial infections in humans is Urinary tract infection particularly when these infections are caused through MDR isolates which resist various traditional treatment especially antibiotic-based methods. Objectives: In this article we were isolate then biochemical and molecular identification of most potent MDR isolates cause UTIs in Egyptian patients' males and females. Methodology: different urine samples were obtained from male and female patients, used as a source for bacterial pathogen isolates, which purified on nutrient agar plates, then isolates cultivated on blood agar to determine their hemolysis activity, then these all-bacterial isolates were tested through agar disc diffusion method against twelve different antibiotics, then primarily identified through gram reaction then through VITEK2, followed by molecular identification through 16S rDNA sequencing. Results: in our study we obtained 22 different bacterial isolates which all are gram negative belonging to different four genera based on their growth on Uriselect into E. coli, Klebsiella sp, Pseudomonas sp., and Acinetobacter sp., they have beta-hemolytic activity, beside that they exhibit MDR behavior against twelve antibiotics, according to growth on Macconkey agar, 17 isolates with lactose fermenting (16 Klebsiella sp and one E. coli) and five were non-lactose fermenting isolates. The most potent bacterial isolates were Klebsiella pneumonia A061 and Klebsiella pneumonia A031 with accession numbers OP811040 and OP811041 respectively. Conclusion: MDR Klebsiella pneumonia one of the most threat of Urinary tract in humans as they resist traditional treatment with antibiotics. For that a new strategy should be considered to eliminate these bacterial pathogens