3.4. In vivo BNZ resistance profile
First, parasitaemia profiles were monitored during the acute-phase (Figure 5A ). Significant differences were observed between untreated and BNZ-treated mice, showing a remarkable parasitaemia reduction even at subcurative BNZ doses. Second, the experimental cure was determined using a double checking widely used in in vivotrypanocidal assays: immunosuppression and PCR (Figure 5A-B ) (Santos et al. , 2010; Francisco et al. , 2016; Martín-Escolano et al. , 2020c). Mice whose parasitaemia and PCR remain negative after immunosuppression (enhancing the reactivation of any residual infection) are considered cured.
Finally, organomegaly of spleens and hearts was measured (Figure 6 ). They are manifested in both the acute- and chronic-phases of CD, allow us to characterise the pathogenesis of the T. cruziArequipa strain, and also allow us an indirect evaluation of the BNZ efficacy since they are directly associated with the parasitic load (Martín-Escolano et al. , 2018, 2019, 2020d). Splenomegaly and cardiomegaly occurred in experimentally infected mice in both phases of CD, during which the spleens of infected mice were approximately twice the mass of those from uninfected mice. The hearts also suffered a significant enlargement. Importantly, treatment with BNZ reduced infection-induced splenomegaly and cardiomegaly, even at subcurative doses and in the absence of a parasitological cure, since it is linked to a reduction of the parasite load (Francisco et al. , 2015).