Multiple Mitochondrial Introgression Events and Heteroplasmy in Trypanosoma cruzi Revealed by Maxicircle MLST and Next Generation Sequencing

Abstract

Background Mitochondrial DNA is a valuable taxonomic marker due to its relatively fast rate of evolution. In Trypanosoma cruzi , the causative agent of Chagas disease, the mitochondrial genome has a unique structural organization consisting of 20–50 maxicircles (∼20 kb) and thousands of minicircles (0.5–10 kb). T. cruzi is an early diverging protist displaying remarkable genetic heterogeneity and is recognized as a complex of six discrete typing units (DTUs). The majority of infected humans are asymptomatic for life while 30–35% develop potentially fatal cardiac and/or digestive syndromes. However, the relationship between specific clinical outcomes and T. cruzi genotype remains elusive. The availability of whole genome sequences has driven advances in high resolution genotyping techniques and re-invigorated interest in exploring the diversity present within the various DTUs. Methodology/Principal Findings To describe intra-DTU diversity, we developed a highly resolutive maxicircle multilocus sequence typing (mtMLST) scheme based on ten gene fragments. A panel of 32 TcI isolates was genotyped using the mtMLST scheme, GPI , mini-exon and 25 microsatellite loci. Comparison of nuclear and mitochondrial data revealed clearly incongruent phylogenetic histories among different geographical populations as well as major DTUs. In parallel, we exploited read depth data, generated by Illumina sequencing of the maxicircle genome from the TcI reference strain Sylvio X10/1, to provide the first evidence of mitochondrial heteroplasmy (heterogeneous mitochondrial genomes in an individual cell) in T. cruzi . Conclusions/Significance mtMLST provides a powerful approach to genotyping at the sub-DTU level. This strategy will facilitate attempts to resolve phenotypic variation in T. cruzi and to address epidemiologically important hypotheses in conjunction with intensive spatio-temporal sampling. The observations of both general and specific incidences of nuclear-mitochondrial phylogenetic incongruence indicate that genetic recombination is geographically widespread and continues to influence the natural population structure of TcI, a conclusion which challenges the traditional paradigm of clonality in T. cruzi .

Ecological host fitting of Trypanosoma cruzi TcI in Bolivia: mosaic population structure, hybridization and a role for humans in Andean parasite dispersal

“…Consistent with previous studies, no evidence of reciprocal nuclear hybridization was detected among recombinant strains (Messenger et al . 2012; Ramírez et al . 2012).…”

“…For NADH dehydrogenase subunit 4 ( ND4 ), an alternate set of primers was designed to improve amplification efficiency: ND4 forward (5′‐TTYTTCCCAATATGTATBGTMAG‐3′) and ND4 reverse (5′‐TGTATTAYCGAYCAATTYGC‐3′), and reactions were performed using the same conditions as previously (Messenger et al . 2012). …”

“…(2012) and are available from Dryad (doi: 10.5061/dryad.b8465). Additional mitochondrial multilocus sequence typing (mtMLST) data from 24 previously published TcI strains were included in selected analyses, as indicated (Messenger et al .…”

“…Additional mitochondrial multilocus sequence typing (mtMLST) data from 24 previously published TcI strains were included in selected analyses, as indicated (Messenger et al . 2012). The most appropriate nucleotide substitution model was selected from 1624 candidates, based on the Akaike information criterion (AIC), in jmodeltest 2.1.4 (Darriba et al .…”

Is Predominant Clonal Evolution a Common Evolutionary Adaptation to Parasitism in Pathogenic Parasitic Protozoa, Fungi, Bacteria, and Viruses?

“…'far from being a clonal species' (Coscoll a et al, 2011), 'extensive genetic exchange' (Caugant and Maiden, 2009), 'gross' incongruences (between phylogenetic trees) (Messenger et al, 2012), 'widespread genetic exchange' Ramírez and Lllewellyn, 2014), 'intense lateral exchange of genetic information' (Ramírez and Llewellyn, 2015), 'genetic exchange is a fairly common phenomenon' (Lima et al, 2014), 'genetic exchange playing a major role in T. cruzi population structure' (Minning et al, 2011), and others. It is impossible to totally discard such expressions in the scientific literature.…”