Despite the major impact of Blastocystis sp. in terms of prevalence in human and animal populations and the risk of zoonotic transmission, no epidemiological survey has yet been conducted in cattle herds in France. The aim of this study was thus to assess the prevalence and molecular diversity of Blastocystis sp. and associated factors in dairy cattle from the north of France. A total of 1581 fecal samples were collected from 1246 animals reared in 20 farms. Molecular detection of the protozoan was performed by real-time PCR and indicated an overall prevalence of Blastocystis sp. reaching 54.8% in the study population. Important inter-herd variation (from 22.2% to 76.5%) of Blastocystis sp. prevalence was also reported. Sequence analysis of 159 positive samples highlighted a very large predominance of ST10 (36/159) and ST14 (64/159), and ST2 was only found in 2 samples. Mixed subtype infections were common, representing 35.8% of sequenced samples (57/159). A putative correlation between Blastocystis sp. colonization and various animal and herd characteristics or feed intake was subsequently investigated. The protozoan was less prevalent in cows that have recently calved but Blastocystis sp. carriage was not significantly related to age. Blastocystis sp. colonization also decreased with high beet pulp and pasture grass consumption and increased with corn silage intake. Finally, the only significant association between Blastocystis sp. STs and animal and herd characteristics was the number of lactations of cows, with a predominance of ST14 in cows that calved once only.