Autophagic cell death induction in a clinical isolate of Acanthamoeba sp. treated with methanolic extracts of Pereskia bleo Kuhn.

Aims: This study was aimed to investigate the effects of methanol extracts from various parts of the Pereskia bleo Kuhn plant on Acanthamoeba sp. The antioxidant levels of each extract from different plant parts were measured after the extraction process. These extracts were then exposed to Acanthamoeba sp. to assess dose-response, IC₅₀ values, changes in cell morphology, internal cell activity and apoptosis based on alterations in phospholipids. Methodology and results: The total phenolic content, carotenoid estimation and antioxidant activity of the leaves, flowers and fruits of P. bleo were measured based on 1,1-diphenyl-2-picrylhydrazyl radicals (DPPH) assay. Its antiamoebic properties were tested using 3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide (MTT) assay for the IC₅₀ determination. The morphological and biochemical changes in the Acanthamoeba sp. were observed under light and fluorescence microscopy using the acridine orange and propidium iodide double staining (AO/PI). The IC₅₀ values of P. bleo leaves, flowers and fruits methanolic extracts were 5.884%, 0.1646% and 20.69%, respectively. Morphological observation displayed shortened acanthapodia with darkened cytoplasms. AO/PI-stained Acanthamoeba sp. cells appear with orange-fluorescent organelles in their green cytoplasm, indicating autophagic cell deaths. Apoptotic and necrotic Acanthamoeba sp. cells were absent based on Annexin V labelling. Conclusion, significance and impact of study: This study confirmed that the methanolic crude extracts of P. bleo exhibit high cytotoxic potential towards Acanthamoeba sp. trophozoites by inducing an autophagic mode of cell death. [ABSTRACT FROM AUTHOR]