Introduction: Blastocystis spp. is a protist found in humans. Although usually the most frequent protozoa found in stool samples of both symptomatic and healthy subjects, its pathogenic or rather opportunistic role is yet to be clearly elucidated. To attempt to fill this gap, a cross-sectional study was conducted to compare the frequency of Blastocystis spp. in HIV positive (HIV+) versus HIV negative (HIV-) individuals in four health facilities of the Center Region of Cameroon. Methodology: Stool samples were collected from 283 HIV positive and 245 HIV negative subjects and analyzed using direct diagnostic tests. Results: A total of 46 (8.7%) individuals were found infected with Blastocystis spp., including 6.7% HIV positive and 11.0% HIV negative. This species was more frequent in urban and semi-urban areas than in rural areas, but evenly distributed among genders and age groups as well as among all sectors of activity. The prevalence of Blastocystis spp. (11.3%) was higher in HIV+ patients with a CD4 count ≥ 500 cells / mm3, but no significant difference was found among HIV clinical stages. Likewise prevalence, the mean number of cysts per gram of stool was similar between HIV positive and HIV negative individuals. People infected with Blastocystis spp. showed diverse clinical signs, but only flatulence was significantly more prevalent. The frequencies of these clinical signs were not related to HIV status. Conclusion: No clear relationship links the infection with Blastocystis spp. to HIV, although its presence was associated with digestive disorder, suggesting that this parasite might not be opportunist.