Blastocystis is a common intestinal parasitic protist infecting birds and mammals. Blastocystis comprises at least 17 subtypes (ST), of which ST1-ST9 have been detected in humans. Significant correlation between certain subtypes and pathogenicity remains to be established. Nevertheless, some studies suggest a potential linkage between subtypes (inter- and intra-subtype variation) and clinical manifestations. The aim of this study was to identify intra-subtype genetic variation of subtypes of Blastocystis in stools samples submitted by diarrheic and non-diarrheic patients. A 550-bp fragment of the nuclear small subunit ribosomal rRNA gene was amplified from 58 culture-positive samples isolated from diarrheic and non-diarrheic Iranian patients. PCR products were sequenced and sequences subjected to phylogenetic analysis. Intra-and inter-subtype variation was calculated. Based on comparison with reference sequences in GenBank, ST1, ST2 and ST3 were found in 18 (31.03%), 21 (36.22%), and 19 (32.75%) of the samples, respectively. Diarrheic stools were observed in eight (44.44%), 10 (47.61%), and nine (47.36%) patients with ST1, ST2, and ST3, respectively. No statistically significant correlation was found between subtypes and diarrhea (P = 1.000). Multiple sequence alignment exhibited a within-subtype similarity of 98.76%, 97.17%, and 99.78% in ST1, ST2, and ST3, respectively. Highest similarity was seen among ST3 isolates, while lowest similarity was seen among ST2 isolates. Phylogenetic analysis did not suggest any correlation between diarrhea and intra-subtype variation. Inter- and intra-subtype variation in SSU rRNA gene appears not to reflect differences in the clinical outcome of Blastocystis carriage. (C) 2017 Elsevier B.V. All rights reserved.