To find out whether intestinal microflora in colicky infants is different from that in noncolicky controls, stool samples were collected from colicky infants during colic (n = 55) and at the age of 3 months (n = 46) and compared with samples from age-matched controls (n = 49 and n = 45, respectively). The samples were cultured on several selective and unselective aerobic and anaerobic culture agars, and gas-liquid chromatography of bacterial cellular fatty acids was used to produce fatty-acid profiles of the stool samples. In quantitative bacterial cultures, no differences were found between the colicky and control groups in the amounts of each bacterium. The colicky infants were more frequently colonized with Clostridium difficile during the time of colic than were the age-matched controls. This difference disappeared by age 3 months. The fatty-acid profiles did not differ between the colicky and control groups as a whole at the time of colicky symptoms. At age 3 months, a difference in fatty-acid profiles was found between the colicky infants who had suffered from severe colic and the control infants. The fatty-acid profiles were also influenced by the age of the infant, the mode of delivery, antimicrobial drugs taken by the mother during delivery, and breast-feeding and type of feeding. In conclusion, no difference in intestinal microflora was found between the colicky infants at the time of colic and the controls. However, a difference in bacterial cellular fatty-acid profiles at the age of 3 months was found that correlated with severe infantile colic. This difference may contribute to the cause(s) of colic, or it may be secondary to the colic, which may influence the microbial environment of the intestine.