Successful Genetic Transfection of the Colonic Protistan Parasite Blastocystis for Reliable Expression of Ectopic Genes.

The microbial parasite Blastocystis colonizes the large intestines of numerous animal species and increasing evidence has linked Blastocystis infection to enteric diseases with signs and symptoms including abdominal pain, constipation, diarrhea, nausea, vomiting, and flatulence. It has also recently been reported to be an important member of the host intestinal microbiota. Despite significant advances in our understanding of Blastocystis cell biology and host-parasite interactions, a genetic modification tool is absent. In this study, we successfully established a robust gene delivery protocol for Blastocystis subtype 7 (ST7) and ectopic protein expression was further tested using a high sensitivity nano-luciferase (Nluc) reporter system, with promoter regions from several genes. Among them, a strong promoter encompassing a region upstream of the legumain 5' UTR was identified. Using this promoter combined with the legumain 3' UTR, which contains a conserved, precise polyadenylation signal, a robust transient transfection technique was established for the first time in Blastocystis. This system was validated by ectopic expression of proteins harbouring specific localization signals. The establishment of a robust, reproducible gene modification system for Blastocystis is a significant advance for Blastocystis research both in vitro and in vivo. This technique will spearhead further research to understand the parasite's biology, its role in health and disease, along with novel ways to combat the parasite.

Blastocystis is associated with decrease of fecal microbiota protective bacteria: comparative analysis between patients with irritable bowel syndrome and control subjects.

Blastocystis is a protistan parasite living in the digestive tract of many animals, including humans. This highly prevalent intestinal parasite is suspected to be linked to Irritable Bowel Syndrome (IBS), a chronic functional bowel disorder. Here, we first compared the prevalence of Blastocystis among 56 IBS patients (40 IBS with constipation (IBS-C), 9 IBS with diarrhea (IBS-D), 4 mixed IBS (IBS-M) and 3 unsubtyped IBS (IBS-U) according to the Rome III criteria) and 56 control (i.e. without any diagnosed chronic or acute gastrointestinal disorder) subjects. The highest prevalence of Blastocystis spp. was observed in the IBS group, but was only statistically significant in men (36.8% in the IBS group versus 4.8% in the control group). We then conducted a meta-analysis including epidemiological studies attempting to determine whether Blastocystis carriage could be linked to IBS, and highlighted that IBS patients had a relative risk of 2.34 to be infected by Blastocystis when compared to non-IBS subjects. We also looked for Dientamoeba fragilis, which is often associated with IBS, and identified this parasite only in some IBS patients (n = 6/56). Several studies provided evidence for a major role of the gut microbiota in the pathophysiology of IBS. Thus, we investigated the possible impact of Blastocystis carriage on the enteric bacterial community through quantification of 8 major bacterial groups from the enteric flora. Our data indicated that men with IBS-C had a significant decrease in Bifidobacterium sp. when infected by Blastocystis. Interestingly, in control subjects (i.e. without any gastrointestinal disorder) positive for Blastocystis, Faecalibacterium prausnitzii, which is known for its anti-inflammatory properties, was significantly decreased in men. Our results support the hypothesis that Blastocystis might be linked to the pathophysiology of IBS-C and intestinal flora imbalance.

Investigación de blastocystis spp, giardia spp y cryptosporidium spp en aguas de consumo en una comunidad de Caracas- Venezuela: Reporte preliminar / Surveillance of blastocystis spp, giardia spp and cryptosporidium spp in human consumption water in a community of Caracas- Venezuela

Uno de los mecanismos de transmisión de protozoarios intestinales, es el consumo de agua contaminada con quistes y ooquistes, cuya eliminación por cloración o filtración no resulta eficaz. En una comunidad de Caracas, se evaluó la posible contaminación del agua de consumo, con Blastocystis spp, Giardia spp y Cryptosporidium spp. El sedimento obtenido mediante filtración y separación inmunomagnética de 15 muestras de agua, se examinó microscópicamente al fresco, con coloraciones, inmunofluorescencia y cultivo en medio de Boeck- Drbohlav modificado (BDM). Se recopiló información sobre las condiciones de suministro, almacenamiento y consumo del agua, además del procedimiento utilizado para el lavado de frutas y vegetales. El único parásito observado fue Blastocystis spp (60%), mediante examen directo/cultivo (33%). Se observó un mayor consumo de agua filtrada que hervida (p= 0,001). Predominó el uso del agua de chorro para el lavado de vegetales y frutas, más que con agua y vinagre (p= 0,011). Se observó una mayor proporción de averías en los sistemas de recolección de aguas servidas (78,6%), más que en los sistemas de aguas blancas (28,6%, p= 0,011). El hallazgo de Blastocystis spp en el agua, se correlaciona con la prevalencia del parásito en habitantes de este sector. Destaca el papel del agua en la transmisión de Blastocystis spp, por lo cual se recomienda filtrarla y hervirla para prevenir la infección con este parásito.

Many intestinal protozoa are transmitted by contaminated water with cysts and oocysts and methods for their elimination as filtration or chlorination are not completely effective. We evaluated a possible consumption water contamination with Blastocystis spp, Giardia spp and Cryptosporidium spp in a community located in Caracas, Venezuela. The pellet obtained by immunomagnetic separation filtration of 15 water samples were examined by microscopic observation (direct examination), stain techniques, immunofluorescence and culture in Drbohlav Boeck-modified medium (BDM). We also collected information about consuming habits, water supply, storage and washing procedures of vegetables and fruits at assessed homes. The only parasite detected was Blastocystis spp (60%), by direct examination/culture (33%). A higher consumption of boiled filtered water (p = 0.001) was observed. The use of tub water for washing vegetables and fruits was predominant, instead of using water and vinegar (p = 0.011). We observed a higher proportion of nonfunctioning sewage collection (78.6%), rather than white water systems (28.6%, p = 0.011). Finding Blastocystis spp in water samples correlates with prevalence of this parasite in residents of this sector. The role of water in Blastocystis spp transmission is significant, so we recommend filtering and boiling it to prevent infection with this parasite.