The Future of Protozoan Infection Research: 3D Cell Culture and beyond.

Protozoan infections remain a major challenge to human health, owing to the substantial impact these parasites exert on their hosts. In vitro culture systems are fundamental for elucidating the host-parasite interactions underlying these infections; however, traditional two-dimensional models are often limited and fail to recapitulate key aspects of human tissue architecture and function. In this context, advanced in vitro cell culture models have emerged as powerful tools, offering increased morphofunctional resemblance to human tissues. This review critically examines recent studies employing advanced platforms, including Transwell systems, spheroids, organoids, scaffolds and biomaterials, microfluidic devices, and organ-on-chip technologies. Collectively, these approaches enable the modeling of diverse human tissues, such as the intestine, brain, heart, and vascular compartments, and support the investigation of multiple pathogenic protozoa, including Trypanosoma cruzi, Plasmodium spp., and Giardia duodenalis. By integrating advanced in vitro models into protozoan infection research, it is possible to achieve a more faithful representation of the human host environment, simulating infection processes and to foster the development of more effective and targeted pharmacological interventions.