Introduction:Staphylococcus aureus bacteria is a common cause of hospital infections.Since most conventional antibiotics cannot treat MRSA infections, which pose substantial hazards to hospitals and populations worldwide, the rise of strains resistant to methicillin and other antimicrobials has become a major worry, especially in hospital settings. Aim and Objectives: To Study the prevalence, antimicrobial susceptibility testing and the molecular characterization of Mec A gene in MRSA isolates obtained from various clinical samples Material and Methods:This was a Cross sectional study carried out in the Department of Microbiology at a tertiary care centre, for a period of 1 year i.e, May 2023 to May 2024. A total of 750 clinical isolates was studied out of which 200 isolates of S. aureus were identified using the biochemical test from the clinical samples. The different Phenotypic Methods including Cefoxitin and Oxacillin Disc Diffusion test and the genotypic method including MecA gene detection for Methicillin Resistant Staphylococcus aureus isolates was performed. The DNA was extracted using the DNA extraction Qiagen Kit and the resistant Mec A gene was detected by the conventional PCR method. Results: In the present study a total of 50 MRSA isolates were identified by CX, OX, and E-test test phenotypically. The prevalence of MRSA and the Mec A gene was observed to be 25% . The maximum number of isolates was obtained from OPD ward 38.5% followed by the medicine ward 28%, surgery ward 25% and least for NICU with 8.5%. The ratio of the males 36 (70%) was more as compared to the females with 14 (28%) with the maximum age of 41-50 years being affected the most followed by 31-40 years and least in the age group of 61years and above. The pus 50% was the most common isolate followed by the blood with 26% and least for urine and the body fluids with 2%. All the MRSA isolates were found 100% sensitive to linezolid, Teicoplanin, vancomycin, however all the isolates were recorded 100% resistant with Cefoxitin and Oxacillin. The presence of MecA gene was observed in all the 50 isolates of MRSA with the prevalence rate of 25 % for Mec A gene. The presence of MecA gene was confirmed by the PCR followed by sequencing assay. Conclusion: The study offers precise instructions for measuring and diagnosing MRSA infections in hospitals and communities. The data generated by this inquiry may be used in future studies to detect MRSA isolates. The management of antibiotic resistance necessitates the implementation of effective control strategies, continuous surveillance, knowledge of MRSA incidence, and maintenance of hygienic standards, which have proven necessary to lower MRSA infections.