Perforation peritonitis is treated with surgery and antibiotics. This study was conducted to identify bacterial and fungal microorganisms responsible for peritonitis in patients with hollow viscus perforation and to examine the influence of these microorganisms on the outcome. A prospective study was conducted from May 2005 to September 2006 involving 84 consecutive patients with spontaneous gastrointestinal perforation peritonitis, who were referred for surgery. Peritoneal fluid was analyzed by microbial culture and biochemical tests for bacteria and fungi. The Jabalpur Prognostic Score was calculated. Forty-two of the 84 patients had positive peritoneal fluid cultures. Escherichia coli was the most common bacterium (n=26) and Candida (n=13) the most common fungus isolated. Bacterial isolates were largely sensitive to amikacin while all the Candida isolates were sensitive to fluconazole. Mortality was significantly higher in patients with positive peritoneal cultures (15/42) compared with those with negative peritoneal cultures (0/42, p<0.001), and in patients with mixed bacterial and fungal-positive cultures (10/13) compared with those with isolated bacterial cultures (5/29, p<0.001). Using the Jabalpur Prognostic Score, positive fungal cultures were found to be associated with a significantly higher than expected mortality. Patients with gastrointestinal perforations and positive peritoneal cultures have a poor prognosis, which is significantly worsened by the association of positive fungal cultures. Early recognition and treatment of fungal infection is advisable.