Most of the Urinary Tract Infections (UTIs) are caused by multidrug-resistant bacteria mostly E. coli which produce Extended Spectrum β-Lactamase (ESBL). The prevalence of UTI infection caused by ESBL producing bacteria is much worldwide and associated with rising health care costs. To identify the pattern of multidrug resistance amongst the E. coli isolates which were obtained from the patients of UTI. We found antibiotic resistance rates as: ampicillin (100%), amoxicillin (100%), cefotaxime (85.7%), tetracycline (88.5%), amikacin (57.1%), nitrofurantoin (65.7%), gentamicin (71.4%), ciprofloxacin (82.8%), chloramphenicol (94.2%), norfloxacin (77.1%), ceftazidime (62.8) and amoxicillin/clavulanic acid (0%). ESBL production was found to be 34.28% by using clavulanic acid as a β-lactamase inhibitor in the phenotypic. The ESBL-encoding genes were characterized for CTX-M, TEM, and SHV genes, CTX-M was found to be 85.4%, TEM 12.8% and SHV 6.5% in isolates of E. coli. Apart from this, we also developed a highly sensitive and specific LAMP assay for ESBL producing E. coli. The assay is rapid (results can be obtained in less than 1 hour), cost-effective easy to perform, and it will be very easy to adapt in small-scale industries, hospitals, and testing laboratories.