Abstract
BackgroundTrypanosoma 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.MethodsInfection 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.ResultsTrypanosoma 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.ConclusionsOur 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 localities (c.28 km apart), suggesting active parasite dispersal over the study area. Our results highlight the value of amplicon sequencing methodologies to clarify the population dynamics of kinetoplastid parasites in endemic regions and inform control campaigns in southern Ecuador.
Highlights
Trypanosoma cruzi, the causative agent of Chagas disease, and T. rangeli are kinetoplastid parasites endemic to Latin America
Our results highlight the value of amplicon sequencing methodologies to clarify the population dynamics of kinetoplastid parasites in endemic regions and inform control campaigns in southern Ecuador
Genotyping by multiplex polymerase chain reaction (PCR) and Next-generation sequencing (NGS) Among the 48 samples analyzed by multiplex PCR, agarose electrophoresis patterns indicated that 37.5% (n = 18) of the samples were co-infected with T. cruzi and T. rangeli, while 52.1% (n = 25) presented infection exclusively with T. cruzi and 6.25% (n = 3) were infected only with T. rangeli
Summary
Trypanosoma cruzi, the causative agent of Chagas disease, and T. rangeli are kinetoplastid parasites endemic to Latin America. 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. Chagas disease (CD) is caused by the kinetoplastid hemoflagellate parasite Trypanosoma cruzi and affects 6–7 million people worldwide [1] This neglected disease is endemic to Latin America, where it poses risk of infection for > 65 million people and kills an estimated 50,000 every year [2]. Two previous reports suggest the occurrence of parasites belonging to DTUs other than TcI, in triatomines and in patients of CD [16, 17]
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