Expanding the toolbox for Trypanosoma cruzi: A parasite line incorporating a bioluminescence-fluorescence dual reporter and streamlined CRISPR/Cas9 functionality for rapid in vivo localisation and phenotyping
A. Ex vivo bioluminescence image of organs from a chronically infected mouse (day 117), showing parasite presence in stomach, colon and lungs (organs placed as in Fig 2D). The red box identifies the tissue sample from which the section shown in B-F was taken. B. mNeonGreen fluorescence. C. Higher magnification of boxed area in B, showing a single amastigote in close conjunction with the nucleus of the host cell. The kinetoplast DNA (K) is indicated by an arrow. D. DNA staining of section shown in B. E. Phase image illustrating architecture of tissue section. F. Merged image of B, D and E, showing phase, DNA and mNeonGreen. The single amastigote is indicated by an arrow and appears yellow. For panel C, bar represents 5 μm; for all other panels, bar indicates 100 μm. Images B, D, E and F are taken with a 40x objective at 0.7 scan zoom, image C is taken with 100x objective, scan zoom 2.7. G. Schematic indicating individual layers of the section. MC, mucosa; MM, muscularis mucosae; SM, submucosa; MEC, muscularis externae (circular); MEL, muscularis externae (longitudinal). The approximate location of the infected cell is indicated within this schematic (red oval and green circle).