Molecular prevalence and genetic diversity of Blastocystis spp. in patients with irritable bowel syndrome and inflammatory bowel disease: a case–control study in southwestern Iran

The clinical significance of Blastocystis spp. in irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD) remains uncertain. This study investigated its molecular prevalence, subtype (ST) distribution, and phylogenetic characteristics among patients with IBS or IBD and healthy controls in southwestern Iran. In this hospital-based case–control study, 181 stool samples were collected from 85 patients (33 with IBS and 52 with IBD) and 96 healthy controls in Shiraz, Iran. Blastocystis spp. was detected by polymerase chain reaction (PCR) targeting the small subunit ribosomal RNA ( SSU rRNA ) gene. Positive amplicons were characterized by single-direction Sanger sequencing, and edited sequences were used for ST assignment and Maximum Likelihood phylogenetic analysis. Group differences were evaluated using appropriate categorical tests, and the association between disease status and Blastocystis spp. detection was expressed as an unadjusted odds ratio (OR) with a 95% confidence interval (CI). Blastocystis spp. DNA was detected in 27/181 (14.9%) participants, including 16/85 (18.8%) patients and 11/96 (11.5%) controls. Detection was more frequent among patients, but the difference was not statistically significant (unadjusted OR = 1.79, 95% CI: 0.78–4.11; P = 0.165). Four STs were identified: ST1, ST2, ST3, and ST7. ST2 and ST3 were the most frequent, each accounting for 10/27 (37.0%) isolates. ST3 predominated among controls, whereas ST2 was most frequent among patients. ST7 was identified in three patients and no controls; because of the small number of isolates, this observation should be considered descriptive and hypothesis-generating. Phylogenetic analysis supported the assigned STs, with study sequences showing 98–100% nucleotide identity to reference sequences. Blastocystis spp. was detected more frequently among patients with IBS or IBD than among healthy controls, but no statistically significant association was demonstrated. ST-specific findings were exploratory and should not be interpreted as evidence of disease association. Larger, adequately powered studies using higher-resolution sequencing are needed to clarify the clinical significance of Blastocystis spp. in these disorders.