Entamoeba histolytica: trophozoite, precyst and cyst studied by atomic force microscopy

Amoebiasis is a disease caused by the cosmopolite parasitic protozoa E. histolytica; it has been estimated that the 10% of the world population is infected by this protozoa, which represents approximately 50 million of invasive amoebiasis cases and as much as 100,000 deaths per year. E. histolytica have four stages: the trophozoite, precyst, cyst and metacyst; trophozoite is the invasive stage and the cyst is the infective stage. Trophozoite, or mobile form, is a uninucleated and pleomorphic structure, measures about 15-40 μm; the cyst measures 10-15 μm, and is a spherical tetranucleated structure covered by a chitin cell wall that confers its resistance to adverse conditions. Atomic force microscopy (AFM) has emerged in recent years as a powerful tool for the study of cellular structure at resolutions of a few nanometers.

Morphological studies revealing details of living cells, which were previously impossible due to the resolution limits of the light microscope, are now possible using AFM. The AFM´s major contribution to biology has been its ability to study the dynamics of live cells in physiological medium at ultrahigh resolution and in real time. Despite the encysting process has been studied in vitro by different authors, many of the aspects about this process still remain unknown. In this work, we studied in situ by atomic force microscopy (AFM) the E. histolytica trophozoite, pre-cyst and cyst, and we made clear some of the differences between each one of the stages, making evident the differences in roughness, composition, elasticity and size. In situ characterization of the E. histolytica trophozoite, pre-cyst and cyst stages was performed.

This is the first report of the pre-cyst stage observed on AFM. In addition, we performed a quantitative comparison of the elasticity of the outer surface of the cell, which shows that the trophozoite cell membrane is more elastic and less rigid than the cell wall of the cyst. Morphological differences between the different stages were also observed, both the morphology and measurements of all cell stages observed in this study are consistent with those reported in the literature. According to the data obtained, as cell goes through the encysting process it loses its elasticity, opposite occurs with the roughness and the adhesion work, which increase as the cell becomes encysted.

Keywords: amoebiasis; atomic force microscopy (AFM); cyst; Entamoeba histolytica; morphometric

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