© 2016 The Author(s). Open Access.Background: We investigated whether the carriage of Blastocystis in IBS patients was associated with differences in the faecal microbiota. Forty patients with diarrhoea-predominant IBS (26 Blastocystis-positive and 14 Blastocystisnegative) and 57 healthy controls (HC) (42 Blastocystis-positive and 15 Blastocystis-negative) submitted faecal samples for metataxonomic analysis of the 16S ribosomal RNA gene. Differences in the relative abundance of bacteria in these IBS and HC groups were evaluated from phylum to genus level. Results: Significant changes were observed in two dominant phyla in IBS patients, regardless of Blastocystis infection status, namely a rise in Firmicutes and a statistically significant reduction in relative abundance of Bacteroidetes (with a threefold increase in the Firmicutes to Bacteoridetes ratio). Significant differences at genus level in IBS subjects compared to HC were also observed for many bacterial species. However, further clinical subgroup analysis of Blastocystis-positive and Blastocystis-negative subjects, regardless of symptoms, showed no significant differences at the phylum or genus level in IBS-P compared to IBS-N. Conclusions: Significant differences in the faecal microbiota between diarrhoea-predominant IBS patients and healthy controls were confirmed, but the carriage of Blastocystis did not significantly alter the faecal microbiota. If Blastocystispositive patients represent a separate clinical subtype of IBS, this group is not identified by changes in the microbiota. Abbreviations: DNA, Deoxyribonucleic acid; HC, Healthy control subjects; HC-N, Healthy control subjects negative for Blastocystis carriage; HC-P, Healthy control subjects positive for Blastocystis carriage; IBS, Irritable bowel syndrome; IBS-C, Constipation-predominant IBS; IBS-D, Diarrhoea-predominant IBS; IBS-M, Mixed bowel habit IBS; IBS-N, Irritable bowel syndrome patients negative for Blastocystis carriage; IBS-P, Irritable bowel syndrome patients positive for Blastocystis carriage; PCoA, Principal Coordinates Analysis; PCR, Polymerase chain reaction; rRNA, Ribosomal ribonucleic acid; XIVC, Xenic in vitro culture.