Abstract
Infection with Trypanosoma cruzi causes Chagas disease. The methods provided here allow for the quantification of T. cruzi in the liver, heart, and blood of intraperitoneally-infected mice and analysis of the killing activity of the cells infected with T. cruzi in vitro.
Keywords: Trypanosoma cruzi, Quantitative PCR, Killing assay, Mouse, Heart, Liver, Blood, Bone marrow-derived macrophage
Background
Chagas disease, characterized by chronic cardiomyopathy, is caused by infection with the intracellular protozoan parasite Trypanosoma cruzi (Bonney et al., 2015). Approximately 20 million people in Latin America suffer from Chagas disease (Ribeiro et al., 2012; Flavia Nardy et al., 2015), and it has become a global health issue owing to the migration of infected individuals (Andrade et al., 2014; Garcia et al., 2015; Requena-Mendez et al., 2015). Several drugs, such as nifurtimox and benznidazole, have been developed for treating Chagas disease. However, these drugs need to be taken for several months and severe side effects have been reported (Viotti et al., 2009). A major aim of treatment is to inhibit T. cruzi transmission via blood as well as prevent the development of heart failure. Thus, the protocol for quantification of T. cruzi and the T. cruzi killing assay presented here might aid the development of novel diagnostic methods and therapeutic strategies for Chagas disease.
Part I: Quantification of T. cruzi in tissue
Materials and Reagents
Equipment
Procedure
Part II: T. cruzi killing assay
Recipes
Acknowledgments
This work was supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan, and the Japan Agency for Medical Research and Development (to K.T. and H. K.), and by the Ichiro Kanehara Foundation and the Kurata Memorial Hitachi Science and Technology Foundation (to H.K.). The protocols provided here were partially modified from previously described methods by S. Cencing et al. (2011) and S. Caldas et al. (2012) (Quantification of T. cruzi) and by J. Weischenfeldt and B. Porse (2008) (Preparation of bone marrow-derived macrophages). We thank Kate Fox, DPhil, from Edanz Group (www.edanzediting.com/ac) for editing a draft of this manuscript. The authors declare that they have no competing financial interests.
References
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