Bioluminescent imaging of Trypanosoma cruzi infection

Abstract

Chagas disease, caused by infection with the protozoan parasite Trypanosoma cruzi, is a major public health problem in Central and South America. The pathogenesis of Chagas disease is complex and the natural course of infection is not completely understood. The recent development of bioluminescence imaging technology has facilitated studies of a number of infectious and noninfectious diseases. We developed luminescent T. cruzi to facilitate similar studies of Chagas disease pathogenesis. Luminescent T. cruzi trypomastigotes and amastigotes were imaged in infections of rat myoblast cultures, which demonstrated a clear correlation of photon emission signal strength to the number of parasites used. This was also observed in mice infected with different numbers of luminescent parasites, where a stringent correlation of photon emission to parasite number was observed early at the site of inoculation, followed by dissemination of parasites to different sites over the course of a 25 day infection. Whole animal imaging from ventral, dorsal and lateral perspectives provided clear evidence of parasite dissemination. The tissue distribution of T. cruzi was further determined by imaging heart, spleen, skeletal muscle, lungs, kidneys, liver and intestines ex vivo. These results illustrate the natural dissemination of T. cruzi during infection and unveil a new tool for studying a number of aspects of Chagas disease, including rapid in vitro screening of potential therapeutic agents, roles of parasite and host factors in the outcome of infection, and analysis of differential tissue tropism in various parasite-host strain combinations.

Putting Infection Dynamics at the Heart of Chagas Disease

“…Broad tissue-type sampling can be easily conducted for groups of identically-treated animals, although in practice most animal studies concentrate on a limited set of tissues. Real-time imaging methods have also been developed that allow serial analysis of T. cruzi infections in individual mice [26-29]. These methods employ transgenic parasites expressing luciferases or fluorescent proteins, such that light signals emitted by parasites can be quantified and pinpointed to anatomical locations.…”

“…The broad range of infected organs was noted in the earliest autopsy studies of fatal acute T. cruzi infection [40]. Analyses of acutely infected mice have shown that diverse parasite strains indeed infect a huge array of cell types in virtually any tissue [16, 20-22, 26, 33, 41-47]. However, the relative abundance of parasites in different cell or tissue types varies greatly.…”

“…It is worth making a distinction between cell and tissue types, which is possible with microscopic detection of parasites, but not with methods that analyse homogenized tissue samples or macro-scale imaging. For example, bioluminescence imaging readily identifies the GI tract and lung as sites of infection [16, 26], but not the infected cell types. Using histological analysis, Guarner et al [21] also described lung infections and found that amastigotes were only localised to the muscular stratum of pulmonary blood vessels.…”

High-content imaging for automated determination of host-cell infection rate by the intracellular parasite Trypanosoma cruzi

“…Alternatively, an assay based on the selective incorporation of radioactive uracil by parasite-infected cells has been described [22]. More recently, Hyland et al (2008) engineered T. cruzi expressing firefly luciferase for bioluminescent determination of infection rates [23], These approaches have been applied for screening of compounds against T. cruzi [24, 25]. However, in addition to other disadvantages, these methods require separate assays for assessment of mammalian cell toxicity and the results are an average of the biological responses of thousands of cells.…”

Drug Susceptibility of Genetically Engineered Trypanosoma cruzi Strains and Sterile Cure in Animal Models as a Criterion for Potential Clinical Efficacy of Anti-T. cruzi Drugs

“…While the use of bioluminescence in Trypanosoma cruzi research has been reported before (3)(4)(5)(6), the improved sensitivity of the red-shifted reporter facilitates following tissue distribution more accurately (2). However, there are issues concerning the nature and interpretation of the experimental results to consider.…”