Abstract: A total of 150 Gallstones samples were collected from patients undergoing cholecystectomy at the general teaching Hilla
Hospital from November 2011 to June 2012. All samples were surveyed and examined for the presences of the Escherichia coliand
differentiate it from other bacterial species. The Bacterial isolates were identified by using morphological and biochemical diagnostic
investigations methods; Samples were cultured on Eosin Methylene Blue Agar (EMB), incubated at 37C? for 24 hrs, and then cultured
on MacConkey Agar (MAC).50 bacterial isolates were identified as Escherichia coli. By using traditional biochemical tests. The
Bacterial isolates were further identified by using Single Polymerase Chain Reaction (SPCR) technique for the presence or absence of 2
genes (astA, uidA) that encode for main virulence factors to diagnose E. coli isolated from Gallstones by using specific primers for these
genes. Appearance of ustA and uidA in E.coli that isolated from gallstone confirmed the importance of these genes in gallstone
formation through their ability to encoded B-glucuronidase.