In vitro effect of a novel protease inhibitor cocktail on Toxoplasma gondii tachyzoites

Toxoplasmosis is a zoonotic disease and a global food and water-borne infection. The disease is caused by the parasite Toxoplasma gondii, which is a highly successful and remarkable pathogen because of its ability to infect almost any nucleated cell in warm-blooded animals. The present study was done to demonstrate the effect of protease inhibitors cocktail (PIC), which inhibit both cysteine and serine proteases, on in vitro cultured T. gondii tachyzoites on HepG2 cell line. This was achieved by assessing its effect on the invasion of the host cells and the intracellular development of T.gondii tachyzoites through measuring their number and viability after their incubation with PIC. Based on the results of the study, it was evident that the inhibitory action of the PIC was effective when applied to tachyzoites before their cultivation on HepG2 cells. Pre-treatment of T.gondii tachyzoites with PIC resulted in failure of the invasion of most of the tachyzoites and decreased the intracellular multiplication and viability of the tachyzoites that succeeded in the initial invasion process. Ultrastructural studies showed morphological alteration in tachyzoites and disruption in their organelles. This effect was irreversible till the complete lysis of cell monolayer in cultures. It can be concluded that PIC, at in vitro levels, could prevent invasion and intracellular multiplication of Toxoplasma tachyzoites. In addition, it is cost effective compared to individual protease inhibitors. It also had the benefit of combined therapy as it lowered the concentration of each protease inhibitor used in the cocktail. Other in vivo experiments are required to validate the cocktail efficacy against toxoplasmosis. Further studies may be needed to establish the exact mechanism by which the PIC exerts its effect on Toxoplasma tachyzoites behavior and its secretory pathway.

Keywords: Cysteine protease; Protease inhibitor cocktail; Serine protease; Toxoplasma gondii.