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Previous studies have shown that the protozoan parasite Blastocystis exhibits apoptotic features with caspase-like activity upon exposure to a cytotoxic monoclonal antibody or the anti-parasitic drug metronidazole. The present study reports that staurosporine (STS), a common apoptosis inducer in mammalian cells, also induces cytoplasmic and nuclear features of apoptosis in Blastocystis, including cell shrinkage, phosphatidylserine (PS) externalization, maintenance of plasma membrane integrity, extensive cytoplasmic vacuolation, nuclear condensation and DNA fragmentation. STS-induced PS exposure and DNA fragmentation were abolished by the mitochondrial transition pore blocker cyclosporine A and significantly inhibited by the broad-range cysteine protease inhibitor iodoacetamide. Interestingly, the apoptosis phenotype was insensitive to inhibitors of caspases and cathepsins B and L, while calpain-specific inhibitors augmented the STS-induced apoptosis response. While the identities of the proteases responsible for STS-induced apoptosis warrant further investigation, these findings demonstrate that programmed cell death in Blastocystis is complex and regulated by multiple mediators. DOI 10.1099/mic.0.034025-0

Previous studies have demonstrated that colony forms of Blastocystis undergo cell death with numerous membrane-bound vesicles containing organelles located within the central vacuole, resembling morphological features of autophagy. In this study, we investigated whether Blastocystis underwent autophagy upon amino acid starvation and rapamycin treatment. Concurrently, we provide new insight into a possible function of the central vacuole. The use of the autophagy marker monodansylcadaverine, and the autophagy inhibitors 3-methyladenine and wortmannin, showed the existence of autophagy in amino-acid-starved and rapamycin-treated Blastocystis. Confocal microscopy and transmission electron microscopy studies also showed morphological changes that were suggestive of autophagy. The unusually large size of the autophagic compartments within the parasite central vacuole was found to be unique in Blastocystis. In addition, autophagy was found to be triggered when cells were exposed to the cytotoxic antibody mAb 1D5, and autophagy was intensified in the presence of the caspase inhibitor zVAD.fmk. Taken together, our results suggest that the core machinery for autophagy is conserved in Blastocystis, and that it plays an important role in the starvation response and cell death of the parasite. DOI 10.1099/mic.0.033944-0