Irritable bowel syndrome (IBS) is a common intestinal disorder that includes continuous or recurrent intestinal pain and discomfort and altered bowel habits. The pathophysiology of IBS is incompletely understood, but it may involve an altered intestinal microbiota. The aim of the present study was to compare the composition and temporal stability of faecal microbiota of IBS patients and healthy controls by applying culture-based techniques and PCR-DGGE analysis. No difference in the prevalence or mean culturable numbers of bacteroides, bifidobacteria, spore-forming bacteria, lactobacilli, enterococci or yeasts were observed between the IBS and control groups, whereas slightly higher numbers of coliforms as well as an increased aerobe:anaerobe ratio was observed in the IBS group. PCR-DGGE revealed more temporal instability in the predominant bacterial population of IBS subjects than in controls. In 9 out of 21 IBS subjects and 5 out of 17 controls the PCR-DGGE profiles obtained from the samples of the same individual on different occasions (sampling points 0, 3 and 6 months) were clearly different. However, the instability in some of the IBS subjects could partly be explained by antibiotic consumption during the study. The present study suggests that instability of intestinal microbiota may be involved in IBS. However, further studies are needed to associate the instability with specific IBS symptoms or with specific bacterial groups and species.