Tuberculosis (TB) is one of the most highly infectious disease in Bangladesh and secondary bacterial infection along with TB may delay the curing period of tuberculosis resulting in arises of various complication like Multi Drug Resistance (MDR). In present study, a total of 450 TB suspected patients were examined during September to December 2012 period. Among those, 100 samples were cultured for isolating secondary bacterial infection of newly detected pulmonary TB (PTB) patients whose were already treated by TB drugs. From these culture samples, 22 were isolated as Klebsiella spp. and 10 were isolated as Staphylococcus aureus. From antibiotic sensitivity study, Amoxacillin, Cephalothin and Cefotaxim showed 100% resistance whereas Ciprofloxacin, Gentamycin and Nalidixic acid showed 100% sensitive to these isolated microbes.