Antibiotics are compounds that are used in prevention and treatment of microbial infections. Their method of action could be bacteriostatic or bacteriocidal in nature. Cephalosporins are a group of β‐lactam antibiotics that have antimicrobial activity due to their interactions with the outer membrane proteins. Proteobacteria producing β‐lactamases are an emerging problem in posing resistance to antibacterial agents. The primary objective of the study was to identify the isolates from Pune, India and to assess the resistance mechanism conferred by them to persist as multidrug resistant (MDR). Detailed genomic assay of the sequences in this study included confirmation of the microorganism’s identity, antibiotic resistance profile construction and review the mechanism most employed by these towards antibiotic resistance. DNA was isolated and sequenced which were then investigatedfor occurrence of antibiotic resistance genes of different classes of antibiotics using various bioinformatics tools. All the isolates in this study were confirmed to be MDR. The most common mechanism observed was high mutations in efflux pump genes that usually expel substances toxic to the bacteria such as antibiotics. Four antibiotic resistance genes were found common to each of the isolates that were adeL, macA, macB and rosB, all of them conferring resistance by way of efflux pumps. The highlight of this study was the two closely associated E. coli that could be identified onlywhen the strains were subjected to antibiotic stress. It was also observed that the antibiotic resistant strain of the two has a suspiciously large genome. These strains also showed larger content of mutated genes while building their antibiotic resistance profile. MDR is a major public health threat, and the presence of resistant organisms in environmental waters is an emerging concern around the world. The potential for this resistance to be transferred to native populations or other pathogenic species is largely unknown and therefore requires a further investigation