Abstract Introduction: Infective factor seems to be a major cause of formation of gallstones. Moynihan’s aphorism that “gall stone is a tomb stone erected in the memory of the organism with in it” is true today [1]. Evidence in favour of infection includes isolation of E. coli, bacterium typhosum, streptococcus from the gallbladder bile and from the center of the gallstones. Methodology: All patients who were subjected to laparoscopic cholecystectomy had their Gallbladder retrieved in a plastic endobag and bile collected in a culture bottle under strict aseptic precautions and transferred at room temperature in an hour to the laboratory for culture. Results: The organisms grown from bile culture were E-Coli 45.5%(41/90), Klebsiella 27.7% (25/90), Enterobacter 8.8% (8/90), Pseudomonas 7.7% (7/90), Staphyl-ococcus 2.2%(2/90), Non-fermenting gram negative bacilli 2.2%(2/90), and Citrobacter freundii 1.1%(1/90). Conclusion: Fluroquinolones (Levofloxacin/ Ofloxacin) may be used as first line drug for preoperative prophylaxis in gallstone diseases undergoing surgery.