Trypanosoma cruzi genome plasticity and evolution

Abstract: Trypanosoma cruzi, a protozoan from the Kinetoplastidae family, is the etiologic agent of Chagas disease, a major public health problem affecting mostly the poorest areas of Latin America. Due to the complex nature of the parasite’s genome it has been impossible to produce a complete reference genome sequence, thus hampering the implementation of post- genomic approaches to unveil the mechanisms of generation of antigenic variation and the identification of new drug targets. My doctoral studies have focused on the application of combined genome sequencing and computational methods to produce a complete reference T. cruzi genome sequence and perform comparative analyses to better understand the mechanisms that allow T. cruzi to evade the mammalian host immune system and to briskly adapt to novel environments. In paper I and II, different genome assembly strategies and second generation sequencing technologies were implemented to perform comparative analyses to identify elements of virulence between T. cruzi and two trypanosomatids that are non-pathogenic to humans: Trypanosoma cruzi marinkellei, a bat-restricted sub-species of the T. cruzi clade and the human avirulent species Trypanosoma rangeli. The studies reveal the expansion of T. cruzi- specific genomic traits specialised in the invasion of mammalian cells. In paper III, using third-generation, PacBio sequencing data it was possible to assemble the complete reference genome sequence of a Trypanosoma cruzi isolate from the DTU-I clade. This breakthrough allowed us - for the first time - to explore in detail the genome architecture of the subtelomeric areas where many parasite virulence factors are encoded. One of the most interesting discoveries was the overrepresentation of interspersed retrotransposons and microsatellites in tandem gene arrays coding for surface molecules, hinting at a retrotransposon-driven mechanism of recombination for generating new sequence variants. Whole genome sequencing of 35 T. cruzi DTU-I isolates, collected from different locations in the American continent, made possible to identify and characterise the mechanisms of adaptability employed by the parasite. Finally, paper IV analyses the mechanisms of genomic hybridisation in T. cruzi and the evolution over time of the hybrid offspring. The analysis revealed that during hybrid formation, the parasite integrates genetic material from each parental strains with the aid of retrotransposons and microsatellites, and the genome of these hybrid isolates moves quickly from a tetraploid to a diploid state. As a result, the hybrid strain has more genetic material, mostly in the subtelomeres, providing the parasite with a pool of new surface molecule genes with the potential to possibly increase its fitness in a new environment. In conclusion, the work presented here has advanced the understanding of parasite biology and provided a genomic resource to be exploited for the identification of drug targets and vaccine candidates.

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