Kinetoplastid flagellates are known for several unusual features, one of which is their complex mitochondrial genome, known as kinetoplast (k) DNA, composed of mutually catenated maxi- and minicircles. Trypanosoma lewisi is a member of the Stercorarian group of trypanosomes which is, based on human infections and experimental data, now considered a zoonotic pathogen. By assembling a total of 58 minicircle classes, which fall into two distinct categories, we describe a novel type of kDNA organization in T. lewisi. RNA-seq approaches allowed us to map the details of uridine insertion and deletion editing events upon the kDNA transcriptome. Moreover, sequencing of small RNA molecules enabled the identification of 169 unique guide (g) RNA genes, with two differently organized minicircle categories both encoding essential gRNAs. The unprecedented organization of minicircles and gRNAs in T. lewisi broadens our knowledge of the structure and expression of the mitochondrial genomes of these human and animal pathogens. Finally, a scenario describing the evolution of minicircles is presented.

Center for Parasitic Organisms State Key Laboratory of Biocontrol School of Life Sciences and Key Laboratory of Tropical Diseases Control of the Ministry of Education Zhongshan School of Medicine Sun Yat Sen University Guangzhou 510275 The People's Republic of China

Ecosystems and Environment Research Centre and Biomedical Research Centre School of Science Engineering and Environment University of Salford Salford UK

Institute of Parasitology Biology Centre Czech Academy of Sciences and Faculty of Science University of South Bohemia České Budějovice 37005 Czech Republic

Key Laboratory of Gene Engineering of the Ministry of Education State Key Laboratory of Biocontrol School of Life Sciences Sun Yat Sen University Guangzhou 510275 The People's Republic of China

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