Blastocystis is the most prevalent eukaryotic microbe colonizing the human gut, infecting approximately 1 billion individuals worldwide. Although Blastocystis has been linked to intestinal disorders, its pathogenicity remains controversial because most carriers are asymptomatic. Here, the genome sequence of Blastocystis subtype (ST) 1 is presented and compared to previously published sequences for ST4 and ST7. Despite a conserved core of genes, there is unexpected diversity between these STs in terms of their genome sizes, guanine-cytosine (GC) content, intron numbers, and gene content. ST1 has 6,544 protein-coding genes, which is several hundred more than reported for ST4 and ST7. The percentage of proteins unique to each ST ranges from 6.2% to 20.5%, greatly exceeding the differences observed within parasite genera. Orthologous proteins also display extreme divergence in amino acid sequence identity between STs (i.e., 59%-61% median identity), on par with observations of the most distantly related species pairs of parasite genera. The STs also display substantial variation in gene family distributions and sizes, especially for protein kinase and protease gene families, which could reflect differences in virulence. It remains to be seen to what extent these inter-ST differences persist at the intra-ST level. A full 26% of genes in ST1 have stop codons that are created on the mRNA level by a novel polyadenylation mechanism found only in Blastocystis. Reconstructions of pathways and organellar systems revealed that ST1 has a relatively complete membrane-trafficking system and a near-complete meiotic toolkit, possibly indicating a sexual cycle. Unlike some intestinal protistan parasites, Blastocystis ST1 has near-complete de novo pyrimidine, purine, and thiamine biosynthesis pathways and is unique amongst studied stramenopiles in being able to metabolize α-glucans rather than β-glucans. It lacks all genes encoding heme-containing cytochrome P450 proteins. Predictions of the mitochondrion-related organelle (MRO) proteome reveal an expanded repertoire of functions, including lipid, cofactor, and vitamin biosynthesis, as well as proteins that may be involved in regulating mitochondrial morphology and MRO/endoplasmic reticulum (ER) interactions. In sharp contrast, genes for peroxisome-associated functions are absent, suggesting Blastocystis STs lack this organelle. Overall, this study provides an important window into the biology of Blastocystis, showcasing significant differences between STs that can guide future experimental investigations into differences in their virulence and clarifying the roles of these organisms in gut health and disease.

Biosciences University of Exeter Exeter United Kingdom

Canadian Institute for Advanced Research CIFAR Program in Integrated Microbial Biodiversity Toronto Canada

Centre for Comparative Genomics and Evolutionary Bioinformatics Dalhousie University Halifax Nova Scotia Canada

CNRS UMR 7257 Aix Marseille University Marseille France

College of Life and Environmental Sciences University of Exeter Exeter United Kingdom

Department of Biochemistry and Molecular Biology Dalhousie University Halifax Nova Scotia Canada

Department of Biological Sciences King Abdulaziz University Jeddah Saudi Arabia

Department of Biology and Ecology Faculty of Science University of Ostrava Ostrava Czech Republic

Department of Cell Biology University of Alberta Edmonton Alberta Canada

Faculty of Infectious and Tropical Diseases London School of Hygiene and Tropical Medicine London United Kingdom

Faculty of Life and Environmental Sciences Prefectural University of Hiroshima Nanatsuka 562 Shobara Hiroshima Japan

INRA USC 1408 AFMB Marseille France

Université Côte d'azur INRA ISA Sophia Antipolis France

Université des Sciences et Technologies de Lille Unité de Glycobiologie Structurale et Fonctionnelle UMR8576 CNRS USTL Cité Scientifique Villeneuve d'Ascq Cedex France

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