RESUMO
Entamoeba histolytica is a parasitic protozoan that causes diarrheal disease in approximately 100 million people worldwide every year. E. histolytica has two forms, the growing trophozoite and the infectious cyst. Trophozoites colonizing the large intestine form cysts that are released into the environment. The ingestion of the cysts in contaminated food and water continues the disease cycle. Here, we investigated the role of glycogen in trophozoite growth and encystation. Glycogen is thought to provide precursors for the synthesis of chitin, a major component of the protective cyst wall. We propose that glycogen also serves as an energy source during metabolic adaptation to different nutrient environments. We examined the role of glycogen in E. histolytica by analyzing the growth and encystation of RNAi strains with reduced expression of the single gene-encoding glycogen synthase (GYS) or two of three genes encoding glycogen phosphorylase (PYG). The GYS RNAi strain had a greatly reduced glycogen accumulation, and both the GYS and PYG RNAi strains exhibited reduced growth in the glucose-poor medium. Both RNAi strains also showed reduced cyst production. Our results suggest glycogen synthesis and degradation are vital to the growth and adaptation of E. histolytica to a low-glucose environment such as that encountered in the large intestine.
RESUMO
Entamoeba histolytica is a parasitic protozoan that causes amoebic dysentery, which affects approximately 90 million people each year worldwide. E. histolytica is transmitted through ingestion of food and water contaminated with the cyst form, which undergoes excystation in the small intestine to the trophozoite form that colonizes the large intestine. The reptile pathogen Entamoeba invadens has served as a model for studying stage conversion between the trophozoite and cyst form due to lack of reproducible encystation of E. histolytica in the laboratory. Although much has been learned about encystation and excystation using E. invadens, the findings do not fully translate to E. histolytica due to the extensive genetic and host differences between these species. Here, we present the first reproducible encystation of E. histolytica in vitro. The cysts produced were viable and displayed the four characteristic hallmarks: round shape, chitinous cell wall, tetranucleation, and detergent resistance. Using flow cytometry analysis, glucose limitation and high cell density were key for encystation, as for E. invadens. Entry into encystation was enhanced by the short-chain fatty acids acetate and propionate, unlike for E. invadens. This new model will now allow the further study of E. histolytica stage conversion, transmission, and treatment.