Deep kinetoplast genome analyses result in a novel molecular assay for detecting Trypanosoma brucei gambiense-specific minicircles

Abstract

The World Health Organization targeted Trypanosoma brucei gambiense (Tbg) human African trypanosomiasis for elimination of transmission by 2030. Sensitive molecular markers that specifically detect Tbg type 1 (Tbg1) parasites will be important tools to assist in reaching this goal. We aim at improving molecular diagnosis of Tbg1 infections by targeting the abundant mitochondrial minicircles within the kinetoplast of these parasites. Using Next-Generation Sequencing of total cellular DNA extracts, we assembled and annotated the kinetoplast genome and investigated minicircle sequence diversity in 38 animal- and human-infective trypanosome strains. Computational analyses recognized a total of 241 Minicircle Sequence Classes as Tbg1-specific, of which three were shared by the 18 studied Tbg1 strains. We developed a minicircle-based assay that is applicable on animals and as specific as the TgsGP-based assay, the current golden standard for molecular detection of Tbg1. The median copy number of the targeted minicircle was equal to eight, suggesting our minicircle-based assay may be used for the sensitive detection of Tbg1 parasites. Annotation of the targeted minicircle sequence indicated that it encodes genes essential for the survival of the parasite and will thus likely be preserved in natural Tbg1 populations, the latter ensuring the reliability of our novel diagnostic assay.

Genome diversity of Leishmania aethiopica

“…Extensive LOH regions have been described previously for obligate asexual eukaryotes, such as the water flea Daphnia pulex ( Tucker et al., 2013 ) and the protozoan parasite Trypanosoma brucei gambiense ( Weir et al., 2016 ; Geerts et al., 2022 ). In T. b. gambiense , most LOH regions are ancestral and thus present in all strains, or at least within sets of strains that share a common ancestry ( Weir et al., 2016 ).…”

Genome diversity ofLeishmania aethiopica

“…Extensive LOH regions have been described previously for obligate asexual eukaryotes, such as the water flea Daphnia pulex [42] and the protozoan parasite Trypanosoma brucei gambiense [25,43]. In T. b. gambiense , most LOH regions are ancestral and thus present in all strains, or at least within sets of strains that share a common ancestry [25].…”

High-throughput analysis of the Trypanosoma cruzi minicirculome (mcDNA) unveils structural variation and functional diversity

“…Minicircle analysis was conducted using the Kinetoplast Genomics (KOMICS) automated Python package, a proven tool used for mitochondrial genome characterization in Leishmania 3 and Trypanosoma brucei 6 . The KOMICS pipeline involves three main steps: assembly, circularization, and polishing.…”

“…These include Leishmania species 2 , 3 , T. brucei sp. 4 – 6 , T. vivax 7 , T. lewisi 8 and T. cruzi 9 , 10 , among others. While most studies concur that maxicircles are valuable for identifying species or strains and for conducting phylogenetic studies, there is emerging evidence of heteroplasmy and genome-introgression in T. cruzi .…”