Chromosomal copy number variation analysis by next generation sequencing confirms ploidy stability in Trypanosoma brucei subspecies

Although aneuploidy usually results in severe abnormalities in multicellular eukaryotes, recent data suggest that it could be beneficial for unicellular eukaryotes, such as yeast and trypanosomatid parasites, providing increased survival under stressful conditions. Among characterized trypanosomatids, Trypanosoma cruzi, Trypanosoma brucei and species from the genus Leishmania stand out due to their importance in public health, infecting around 20 million people worldwide. The presence of aneuploidies in T. cruzi and Leishmania was recently confirmed by analysis based on next generation sequencing (NGS) and fluorescence in situ hybridization, where they have been associated with adaptation during transmission between their insect vectors and mammalian hosts and in promoting drug resistance. Although chromosomal copy number variations (CCNVs) are present in the aforementioned species, PFGE and fluorescence cytophotometry analyses suggest that aneuploidies are absent from T. brucei. A re-evaluation of CCNV in T. b gambiense based on NGS reads confirmed the absence of aneuploidies in this subspecies. However, the presence of aneuploidies in the other two T. brucei subspecies, T. b. brucei and T. b. rhodesiense, has not been evaluated using NGS approaches. In the present work, we tested for aneuploidies in 26 T. brucei isolates, including samples from the three T. brucei subspecies, by both allele frequency and read depth coverage analyses. These analyses showed that none of the T. brucei subspecies presents aneuploidies, which could be related to differences in the mechanisms of DNA replication and recombination in these parasites when compared with Leishmania

Replacement of Leishmania (Leishmania) infantum populations in an endemic focus of visceral Leishmaniasis in Brazil

Visceral leishmaniasis is an important global health problem with an estimated of 50,000 to 90,000 new cases per year. VL is the most serious form of leishmaniasis as it can be fatal in 95% of the cases if it remains untreated. VL is a particularly acute problem in Brazil which contributed with 97% of all cases reported in 2020 in the Americas. In this country, VL affects mainly the poorest people in both urban and rural areas and continues to have a high mortality rate estimated around 8.15%. Here, we performed a temporal parasite population study using whole genome sequence data from a set of 34 canine isolates sampled in 2008, 2012 and 2015 from a re-emergent focus in Southeastern Brazil. Our study found the presence of two distinct sexual subpopulations that corresponded to two isolation periods. These subpopulations diverged hundreds of years ago with no apparent gene flow between them suggesting a process of rapid replacement during a two-year period. Sequence comparisons and analysis of nucleotide diversity also showed evidence of balancing selection acting on transport-related genes and antigenic families. To our knowledge this is the first population genomic study showing a turn-over of parasite populations in an endemic region for leishmaniasis. The complexity and rapid adaptability of these parasites pose new challenges to control activities and demand more integrated approaches to understand this disease in New World foci

Ancestral aneuploidy and stable chromosomal duplication resulting in differential genome structure and gene expression control in trypanosomatid parasites

Aneuploidy is widely observed in both unicellular and multicellular eukaryotes, usually associated with adaptation to stress conditions. Chromosomal duplication stability is a tradeoff between the fitness cost of having unbalanced gene copies and the potential fitness gained from increased dosage of specific advantageous genes. Trypanosomatids, a family of protozoans that include species that cause neglected tropical diseases, are a relevant group to study aneuploidies. Their life cycle has several stressors that could select for different patterns of chromosomal duplications and/or losses, and their nearly universal use of polycistronic transcription increases their reliance on gene expansion/contraction, as well as post-transcriptional control as mechanisms for gene expression regulation. By evaluating the data from 866 isolates covering seven trypanosomatid genera, we have revealed that aneuploidy tolerance is an ancestral characteristic of trypanosomatids but has a reduced occurrence in a specific monophyletic clade that has undergone large genomic reorganization and chromosomal fusions. We have also identified an ancient chromosomal duplication that was maintained across these parasite's speciation, named collectively as the trypanosomatid ancestral supernumerary chromosome (TASC). TASC has most genes in the same coding strand, is expressed as a disomic chromosome (even having four copies), and has increased potential for functional variation, but it purges highly deleterious mutations more efficiently than other chromosomes. The evidence of stringent control over gene expression in this chromosome suggests that these parasites have adapted to mitigate the fitness cost associated with this ancient chromosomal duplication

Whole genome sequencing of Trypanosoma cruzi field isolates reveals extensive genomic variability and complex aneuploidy patterns within TcII DTU

Abstract Background Trypanosoma cruzi, the etiologic agent of Chagas disease, is currently divided into six discrete typing units (DTUs), named TcI-TcVI. TcII is among the major DTUs enrolled in human infections in South America southern cone, where it is associated with severe cardiac and digestive symptoms. Despite the importance of TcII in Chagas disease epidemiology and pathology, so far, no genome-wide comparisons of the mitochondrial and nuclear genomes of TcII field isolates have been performed to track the variability and evolution of this DTU in endemic regions. Results In the present work, we have sequenced and compared the whole nuclear and mitochondrial genomes of seven TcII strains isolated from chagasic patients from the central and northeastern regions of Minas Gerais, Brazil, revealing an extensive genetic variability within this DTU. A comparison of the phylogeny based on the nuclear or mitochondrial genomes revealed that the majority of branches were shared by both sequences. The subtle divergences in the branches are probably consequence of mitochondrial introgression events between TcII strains. Two T. cruzi strains isolated from patients living in the central region of Minas Gerais, S15 and S162a, were clustered in the nuclear and mitochondrial phylogeny analysis. These two strains were isolated from the other five by the Espinhaço Mountains, a geographic barrier that could have restricted the traffic of insect vectors during T. cruzi evolution in the Minas Gerais state. Finally, the presence of aneuploidies was evaluated, revealing that all seven TcII strains have a different pattern of chromosomal duplication/loss. Conclusions Analysis of genomic variability and aneuploidies suggests that there is significant genomic variability within Minas Gerais TcII strains, which could be exploited by the parasite to allow rapid selection of favorable phenotypes. Also, the aneuploidy patterns vary among T. cruzi strains and does not correlate with the nuclear phylogeny, suggesting that chromosomal duplication/loss are recent and frequent events in the parasite evolution