EXTENDED-spectrum β-lactamase (ESBL) producing Klebsiella pneumoniae strains can present resistance to many antibiotic groups due to resistant genes. This study conducted to detect and identify multi-drug resistant (MDR), ESBL producing K. pneumoniae strains from different clinical samples with detection and sequencing of both Temoneira (TEM) and sulfhydryl variable (SHV) genes and using Glycyrrhizic acid nanoparticle as an antimicrobial agent for ESBL producing K. pneumoniae strains. One hundred and fifty clinical specimens were processed. ESBL producing K. pneumoniae strains were detected by double disk synergy test. TEM and SHV genes responsible for MDR in K. pneumoniae were detected by polymerase chain reaction (PCR) and sequence alignment was done using DNA sequencing. The effect of different concentrations of Nano Glycyrrhizic acid was determined. K. pneumoniae was detected in 53.3% of the total collected samples (80/150). Seventy one percent (57/80) of them were found to be multi-drug resistant strains and 63% (36/57) also found to contain the ESBL enzymes. Males were highly infected than females. TEM gene was detected in 52.8% of the ESBL isolates while SHV gene was detected in 72.2%. Twenty Five percent of the ESBL producing K. pneumoniae was found to contain both TEM and SHV genes. Nucleic acid sequence alignment of both genes showed some mutations. Chloramphenicol was found to be the drug of choice to overcome ESBL producing K. pneumoniae with inhibition of 97.2%. The antibacterial activity of Nano Glycyrrhizic acid revealed that 10μg/ml was found to be the minimum bactericidal concentration (MBC) against ESBL producing K. pneumoniae isolates.