Fig 3. Drug-cured infections confer significant protection against challenge with a homologous T . cruzi strain. (A) Outline of strategy. BALB/c mice were infected i.p. with 10 3 bioluminescent trypomastigotes (CL Brener strain) and subjected to curative benznidazole treatment initiated at various times post- infection. 21 days after the end of treatment, they were re-infected i.p. and monitored for a further 70 days. Bioluminescence-negative mice were then immunosuppressed using cyclophosphamide (Materials and Methods) and assessed by ex vivo imaging two weeks later. Results were derived from 2 independent experiments, each involving 6 mice per cohort. (B) Representative ventral images of benznidazole-cured mice following re-infection. The number of days at which drug treatment was initiated, following the primary infection, is indicated (left). Controls are naive mice that were infected at the same point as the re-infected experimental cohorts, and serve to illustrate the normal profile of infection in the absence of any form of conferred immunity. All images use the same log 10 -scale heat-map with minimum and maximum radiance values indicated. (C) Total body bioluminescence (sum of ventral and dorsal images) of drug-cured mice following re-infection (n = 12) (means ± SD) derived by in vivo imaging. The length of the primary infection is indicated (inset). (D) Representative ex vivo bioluminescence images. Upper inset, organs from control mouse 70 days post-infection. Central inset, organs from a mouse that was re-infected following curative treatment initiated on day 36 of the primary infection. On day 70 of the challenge infection, immunosuppressive treatment was initiated and the organs then harvested. In this instance, a residual infection was identified. Lower inset, organs from a mouse treated as above, which was protected against