Deep sequencing reveals multiclonality and new discrete typing units of Trypanosoma cruzi in rodents from the southern United States.

BACKGROUND/
PURPOSE: The parasitic protozoa Trypanosoma cruzi, is widely distributed throughout the Americas. We explored the nature of T. cruzi infection in small rodents from New Orleans (LA, USA), an enzootic region of the parasite in North America.
METHODS: We characterized the full complement of discrete typing units (DTUs) in rodent hosts through next-generation metabarcoding, as conventional PCR and Sanger sequencing approaches only detect the dominant genotype in biological samples. We assayed DTU diversity in tissue samples from 6 T. cruzi PCR positive rodents. The intergenic region of the mini-exon gene was amplified and sequenced on a MiSeq platform. A total of 141 sequences were aligned using Muscle, and TCS networks were constructed to identify DTUs in the samples.
RESULTS: We detected distinct and varying assemblages of DTUs in the rodent hosts. Highly diverse DTU assemblages were detected, with 6-32 haplotypes recovered per individual, spanning multiple DTUs (TcI,TcII, TcIV, TcV and TcVI). Haplotypes varied in frequencies from 82% to less than 0.1%. DTU composition varied according to the tissue analyzed. Rural and urban rodents carried similarly diverse DTU assemblages, though urban rodent species tended to harbor more haplotypes than their sylvatic counterparts.
CONCLUSION: Our results affirm that mammalian hosts can concurrently harbor a diverse complement of parasites, and indicate that there is greater diversity of T. cruzi DTUs present in North America than previously thought. Further investigation is warranted to understand the role of commensal rodents as a reservoir for T. cruzi in sylvatic and peridomestic environments.