The microbiological assessment of the air in operating theatres is critical to control hospital-acquired infections. Regular surveillance is an important tool to evaluate the quality of air and find areas requiring intervention. In this context, the present study is undertaken to assess and compare the microbial contamination levels in operation theatre by active and passive methods. All the environmental surfaces and equipment of OTs and ICU at tertiary care hospital in Vijayapur, included in the study. This study used three sampling procedures: active, passive methods for air sampling, and swabing method for surfaces and equipment. Out of 15 OTs air sampling, the passive method showed more bacterial air contamination than the active method. Statistically, a significant difference was observed with the passive method compared to the active method with p-value of 0.0336 for both bacteria and fungus growth assessment. Out of total 90 swabs collected from all the OTs surfaces and instruments, Pseudomonas species (40%), Bacillus species (40%), Klebsiella species (20%) were the common species isolated. From the 50 swabs collected from in ICUs surfaces and instruments, culture positivity was 16% for pathogenic bacteria; Pseudomonas aeruginosa (62%), Klebsiella pneumonia (25%), and Escherichia coli (13%). The present study showed that the passive method is a better monitoring tool than the active method. So we recommend using passive air sampling method compared to active method, which is easy, cheap, and no instrument is needed for sampling the air.