Blastocystis ratti contains cysteine proteases that mediate interleukin-8 response from human intestinal epithelial cells in an NF-kappaB-dependent manner.

Blastocystis is a ubiquitous enteric protozoan found in the intestinal tracts of humans and a wide range of animals. Evidence accumulated over the last decade suggests association of Blastocystis with gastrointestinal disorders involving diarrhea, abdominal pain, constipation, nausea, and fatigue. Clinical and experimental studies have associated Blastocystis with intestinal inflammation, and it has been shown that Blastocystis has potential to modulate the host immune response. Blastocystis is also reported to be an opportunistic pathogen in immunosuppressed patients, especially those suffering from AIDS. However, nothing is known about the parasitic virulence factors and early events following host-parasite interactions. In the present study, we investigated the molecular mechanism by which Blastocystis activates interleukin-8 (IL-8) gene expression in human colonic epithelial T84 cells. We demonstrate for the first time that cysteine proteases of Blastocystis ratti WR1, a zoonotic isolate, can activate IL-8 gene expression in human colonic epithelial cells. Furthermore, we show that NF-kappaB activation is involved in the production of IL-8. In addition, our findings show that treatment with the antiprotozoal drug metronidazole can avert IL-8 production induced by B. ratti WR1. We also show for the first time that the central vacuole of Blastocystis may function as a reservoir for cysteine proteases. Our findings will contribute to an understanding of the pathobiology of a poorly studied parasite whose public health importance is increasingly recognized.

Blastocystis ratti induces contact-independent apoptosis, F-actin rearrangement, and barrier function disruption in IEC-6 cells.

Blastocystis is an enteric protozoan purportedly associated with numerous clinical cases of diarrhea, flatulence, vomiting, and other gastrointestinal symptoms. Despite new knowledge of Blastocystis cell biology, genetic diversity, and epidemiology, its pathogenic potential remains controversial. Numerous clinical and epidemiological studies either implicate or exonerate the parasite as a cause of intestinal disease. Therefore, the aim of this study was to investigate the pathogenic potential of Blastocystis by studying the interactions of Blastocystis ratti WR1, an isolate of zoonotic potential, with a nontransformed rat intestinal epithelial cell line, IEC-6. Here, we report that B. ratti WR1 induces apoptosis in IEC-6 cells in a contact-independent manner. Furthermore, we found that B. ratti WR1 rearranges F-actin distribution, decreases transepithelial resistance, and increases epithelial permeability in IEC-6 cell monolayers. In addition, we found that the effects of B. ratti on transepithelial electrical resistance and epithelial permeability were significantly abrogated by treatment with metronidazole, an antiprotozoal drug. Our results suggest for the first time that Blastocystis-induced apoptosis in host cells and altered epithelial barrier function might play an important role in the pathogenesis of Blastocystis infections and that metronidazole has therapeutic potential in alleviating symptoms associated with Blastocystis.

Protease activity of Blastocystis hominis.

Parasite-derived proteases are important for the parasite life cycle and the pathogenesis of the disease they produce. Proteases of intestinal protozoan parasite Blastocystis hominis were studied for the first time with azocasein assays and gelatin SDS-PAGE analysis. Parasitic lysates were found to have high protease activity and nine protease bands of low (20-33 kDa) and high (44-75 kDa) molecular weights were reported. Proteases were found to be pH-dependent and highest proteolytic activity was observed at neutral pH. Inhibition studies showed that B. hominis isolate B, like many other protozoan parasites, contains mainly cysteine proteases.

Degradation of human secretory immunoglobulin A by Blastocystis.

Microbial immunoglobulin A (IgA) proteases cleave human secretory IgA, promoting the mucosal adhesion of pathogens. To investigate if the enteric protozoan Blastocystis degrades human secretory IgA, cell lysate and conditioned medium from two species were exposed to immunoglobulin A. Secretory IgA was cleaved by both cell lysate and conditioned medium with mainly cysteine proteinase activity in B. hominis B isolate and aspartic proteinase activity in B. ratii WR1 isolate. These findings suggest that Blastocystis proteases may play a role in parasite survival in vivo.

Plasmodium falciparum: a simplified technique for obtaining singly infected erythrocytes.

We report the development of a simple technique involving 15 ml polypropylene tubes and a rotatory incubator for obtaining erythrocytes singly infected with Plasmodium falciparum. This technique will be useful for cloning of the parasite. Our finding that P. falciparum merozoite invasion is inhibited during rotation suggests that this method may also be useful for the study of parasite-erythrocyte interactions under dynamic circulatory conditions.