Remarkable genetic diversity of Trypanosoma cruzi and Trypanosoma rangeli in two localities of southern Ecuador identified via deep sequencing of mini-exon gene amplicons

Abstract

Background: Trypanosoma cruzi, the causative agent of Chagas disease, and T. rangeli are kinetoplastid parasites endemic to Latin America. Although closely related to T. cruzi and capable of infecting humans, T. rangeli is non-pathogenic. Both parasite species are transmitted by triatomine bugs, and the presence of T. rangeli constitutes a confounding factor in the study of Chagas disease prevalence and transmission dynamics. Trypanosoma cruzi possesses high molecular heterogeneity: seven discrete typing units (DTUs) are currently recognized. In Ecuador, T. cruzi TcI and T. rangeli KP1(-) predominate, while other genetic lineages are seldom reported. Methods: Infection by T. cruzi and/or T. rangeli in different developmental stages of triatomine bugs from two communities of southern Ecuador was evaluated via polymerase chain reaction product size polymorphism of kinetoplast minicircle sequences and the non-transcribed spacer region of the mini-exon gene (n = 48). Forty-three mini-exon amplicons were also deep sequenced to analyze single-nucleotide polymorphisms within single and mixed infections. Mini-exon products from ten monoclonal reference strains were included as controls. Results: Trypanosoma cruzi genetic richness and diversity was not significantly greater in adult vectors than in nymphal stages III and V. In contrast, instar V individuals showed significantly higher T. rangeli richness when compared with other developmental stages. Among infected triatomines, deep sequencing revealed one T. rangeli infection (3%), 8 T. cruzi infections (23.5%) and 25 T. cruzi + T. rangeli co-infections (73.5%), suggesting that T. rangeli prevalence has been largely underestimated in the region. Furthermore, deep sequencing detected TcIV sequences in nine samples; this DTU had not previously been reported in Loja Province. Conclusions: Our data indicate that deep sequencing allows for better parasite identification/typing than amplicon size analysis alone for mixed infections containing both T. cruzi and T. rangeli, or when multiple T. cruzi DTUs are present. Additionally, our analysis showed extensive overlap among the parasite populations present in the two studied

Identification of Multiple Blastocystis Subtypes in Domestic Animals From Colombia Using Amplicon-Based Next Generation Sequencing

“…Because in samples from feces mixed infections are common ( 8 , 83 , 84 ), it is important to deploy NGS tools in studies of Blastocystis subtype diversity to have a better picture of STs present within a host. The usefulness of NGS in detecting mixed infections has also been demonstrated in other organisms such as Trypanosoma cruzi and Trypanosoma rangeli where NGS successfully detected co-infections of these two parasites in infected triatomines ( 85 ) and in triatomine food sources ( 86 ). NGS has aided in the determination of the diversity of low-density Plasmodium falciparum infections ( 87 ).…”

Development of an Amplicon-Based Next-Generation Sequencing Protocol to Identify Leishmania Species and Other Trypanosomatids in Leishmaniasis Endemic Areas

“…Various studies capitalizing on the advantages of this technology have been conducted for different purposes, for example, determining the prevalence and diversity of bacterial communities ( 18 , 19 ) and intestinal protozoa ( Blastocystis ) ( 20 ) or characterizing single-nucleotide polymorphisms in viruses ( 21 ). Moreover, NGS has allowed the identification of blood-feeding sources of triatomines ( 22 ), detection of Leishmania parasites, blood sources, and plant meals, and deciphering of the intestinal microbiome of phlebotomine sand flies ( 23 ), in addition to assessment of the genetic diversity of trypanosomatids in humans ( Trypanosoma cruzi and Trypanosoma rangeli ) ( 24 , 25 ) and mammalian reservoir hosts ( 17 , 26 , 27 ). However, relatively few studies have applied this technique to identify and detect coinfection events associated with Leishmania in different biological samples ( 28 ).…”

Whole-genome assembly of a hybrid Trypanosoma cruzi strain assembled with Nanopore sequencing alone

“…T. cruzi's phenotypic diversity is accompanied by a corresponding genomic diversity that is observed across both field isolates and lab-adapted strains ( Llewellyn et al . 2011 ; Maiguashca Sánchez et al . 2020 ; Talavera-López et al .…”