Extracellular Vesicles Mediate Epimastigogenesis in : Strain-Specific Dynamics and Temperature-Dependent Differentiation.

, the causative agent of Chagas disease, undergoes epimastigogenesis-a critical differentiation step in which trypomastigotes transform into epimastigotes. While this process is essential for the parasite's survival in its insect vector, the molecular mechanisms regulating it remain poorly understood. Here, we present the first evidence implicating extracellular vesicles (EVs) as enhancing mediators of epimastigogenesis. Using in vitro models with strains CL Brener and Dm28c, we demonstrate that EVs, membrane-bound vesicles, were shown to enhance differentiation in a strain-specific and temperature-dependent manner. We observed strain-specific EV release patterns: Dm28c produced more EVs at 24 h, whereas CL Brener peaked at 72 h. Additionally, we confirm that epimastigogenesis occurs exclusively at 28 °C after 72 h. These findings establish EVs as novel regulators of differentiation and suggest new insight into parasite development, highlighting potential targets for therapeutic intervention. The observed enhancement of differentiation in the presence of EVs indicates a potential role for these vesicles in this developmental process, although the underlying mechanisms remain undefined.