Third-generation sequencing revises the molecular karyotype for and identifies emerging copy number variants in sexual recombinants.

is a useful model for intracellular parasitism given its ease of culture in the laboratory and genomic resources. However, as for many other eukaryotes, the genome contains hundreds of sequence gaps owing to repetitive and/or unclonable sequences that disrupt the assembly process. Here, we use the Oxford Nanopore Minion platform to generate near-complete de novo genome assemblies for multiple strains of and its near relative, We significantly improved genome contiguity (average N50 of ∼6.6 Mb) and added ∼2 Mb of newly assembled sequence. For all of the strains that we sequenced (RH, ME49, CTG, II×III progeny clones CL13, S27, S21, S26, and D3X1), the largest contig ranged in size between 11.9 and 12.1 Mb in size, which is larger than any previously reported chromosome, and found to be due to a consistent fusion of Chromosomes VIIb and VIII. These data were validated by mapping existing ME49 Hi-C data to our assembly, providing parallel lines of evidence that the karyotype consists of 13, rather than 14, chromosomes. By using this technology, we also resolved hundreds of tandem repeats of varying lengths, including in well-known host-targeting effector loci like rhoptry protein 5 () and Finally, when we compared with , we found that although the 13-chromosome karyotype was conserved, extensive, previously unappreciated chromosome-scale rearrangements had occurred in and since their most recent common ancestry.