The genome of Encephalitozoon cuniculi, a eukaryotic parasite, was sequenced and analyzed, revealing a genome of approximately 2.9 Mb with 1,997 potential protein-coding genes. The genome is highly compact, with reduced intergenic regions and short protein sequences compared to eukaryotic orthologues. This compactness reflects the parasite's strong host dependence, as it lacks genes for certain biosynthetic pathways and the tricarboxylic acid cycle. Phylogenetic analysis supports a fungal affiliation for microsporidia, despite their lack of mitochondria. The genome contains genes related to mitochondrial functions, suggesting that microsporidia may have retained a mitochondria-derived organelle. The genome consists of 11 chromosomes, with the smallest chromosome sequenced. The genome is organized with a 'unique sequences' core region flanked by divergent regions containing ribosomal DNA. The genome shows low base polymorphism and a high gene density, with about 90% of the chromosome cores assigned to functional categories. The genome lacks many complex regulatory elements, such as retrogenes and transposable elements, and has a low number of protein-coding genes compared to other eukaryotes. The genome contains genes involved in DNA replication, RNA processing, and protein synthesis, but with significant simplification due to the absence of DNA-containing organelles. The parasite lacks genes for the tricarboxylic acid cycle, fatty acid β-oxidation, and the respiratory electron-transport chain, suggesting that ATP production occurs through substrate-level phosphorylation. The genome also contains genes for the synthesis of trehalose, a major sugar reserve in microsporidia, and a complete glycolytic pathway. The genome shows evidence of a simplified Golgi apparatus and potential endocytosis pathways. The parasite has a unique set of proteins involved in protein folding, glycosylation, and membrane transport. The genome also contains genes for a potential mitochondria-derived organelle, suggesting that microsporidia may have retained a remnant of a mitochondrion. The genome sequence provides insights into the evolution and biology of microsporidia, which are considered 'curious fungi' due to their lack of mitochondria. The genome may also be useful for comparative genomics of microbial eukaryotes.The genome of Encephalitozoon cuniculi, a eukaryotic parasite, was sequenced and analyzed, revealing a genome of approximately 2.9 Mb with 1,997 potential protein-coding genes. The genome is highly compact, with reduced intergenic regions and short protein sequences compared to eukaryotic orthologues. This compactness reflects the parasite's strong host dependence, as it lacks genes for certain biosynthetic pathways and the tricarboxylic acid cycle. Phylogenetic analysis supports a fungal affiliation for microsporidia, despite their lack of mitochondria. The genome contains genes related to mitochondrial functions, suggesting that microsporidia may have retained a mitochondria-derived organelle. The genome consists of 11 chromosomes, with the smallest chromosome sequenced. The genome is organized with a 'unique sequences' core region flanked by divergent regions containing ribosomal DNA. The genome shows low base polymorphism and a high gene density, with about 90% of the chromosome cores assigned to functional categories. The genome lacks many complex regulatory elements, such as retrogenes and transposable elements, and has a low number of protein-coding genes compared to other eukaryotes. The genome contains genes involved in DNA replication, RNA processing, and protein synthesis, but with significant simplification due to the absence of DNA-containing organelles. The parasite lacks genes for the tricarboxylic acid cycle, fatty acid β-oxidation, and the respiratory electron-transport chain, suggesting that ATP production occurs through substrate-level phosphorylation. The genome also contains genes for the synthesis of trehalose, a major sugar reserve in microsporidia, and a complete glycolytic pathway. The genome shows evidence of a simplified Golgi apparatus and potential endocytosis pathways. The parasite has a unique set of proteins involved in protein folding, glycosylation, and membrane transport. The genome also contains genes for a potential mitochondria-derived organelle, suggesting that microsporidia may have retained a remnant of a mitochondrion. The genome sequence provides insights into the evolution and biology of microsporidia, which are considered 'curious fungi' due to their lack of mitochondria. The genome may also be useful for comparative genomics of microbial eukaryotes.