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.

Genetic Diversity and Distribution of Blastocystis Subtype 3 in Human Populations, with Special Reference to a Rural Population in Central Mexico.

Blastocystis subtype 3 (ST3) is a parasitic protist found in the digestive tract of symptomatic and asymptomatic humans around the world. While this parasite exhibits a high prevalence in the human population, its true geographic distribution and global genetic diversity are still unknown. This gap in knowledge limits the understanding of the spread mechanisms, epidemiology, and impact that this parasite has on human populations. Herein, we provided new data on the geographical distribution and genetic diversity of Blastocystis ST3 from a rural human population in Mexico. To do so, we collected and targeted the SSU-rDNA region in fecal samples from this population and further compared its genetic diversity and structure with that previously observed in populations of Blastocystis ST3 from other regions of the planet. Our analyses reveled that diversity of Blastocystis ST3 showed a high haplotype diversity and genetic structure to the world level; however, they were low in the Morelos population. The haplotype network revealed a common widespread haplotype from which the others were generated recently. Finally, our results suggested a recent expansion of the diversity of Blastocystis ST3 worldwide.

First genotyping of Blastocystis sp. in dairy, meat, and cashmere goats in northwestern China.

Blastocystis is one of the most common parasites inhabiting in small intestines of human and animals. Although its pathogenicity has been remaining controversial, the possibility of zoonotic transmission between human and animals was recognized. The goat was one of the most important economic animals supplying people with cashmere, meat, and dairy products. However, few studies were to investigate Blastocystis infection in goats. A total of 789 faecal specimens of goats (including 362 of dairy, 193 of meat and 234 of cashmere goats) were collected from multiple regions of Shaanxi province in northwestern China to investigate the colonization frequency and subtypes of Blastocystis, and to assess the zoonotic potential of these goats. The respective colonization frequencies of Blastocystis in dairy, meat and cashmere goats were 54.1% (196/362), 40.4% (78/193) and 78.6% (184/234). The prevalence of Blastocystis in pre-weaned (0-2-month) goats was significantly lower than that in goats of other age groups, and the highest colonization was observed in goats of 7-11-month age group. Sequence analysis of Blastocystis positive samples indicated the presence of seven subtypes in these goats, including six known subtypes (STs1, 3, 4, 5, 10, 14) and one possible novel subtype (isolate Sd26), with the subtype 10 as the predominant one. Additionally, zoonotic subtypes were found in dairy (ST1, ST3 and ST5) and cashmere (ST4 and ST5) goats, but not detected in meat goats. These results showed that Blastocystis is highly prevalent, widely distributed and genetically diverse in goats in Shaanxi province, northwestern China, and zoonotic potential of dairy and cashmere goats to transmit Blastocystis.

Proteogenomic Insights into the Intestinal Parasite Blastocystis sp. Subtype 4 Isolate WR1.

Blastocystis sp. is known for years as a highly prevalent anaerobic eukaryotic parasite of humans and animals. Several monophyletic clades have been delineated based on molecular data, and the occurrence of each subtype in humans and/or animal hosts has been documented. The genome of several representatives has been sequenced revealing specific traits such as an intriguing 3'-end processing of primary transcripts. Here, a first high-throughput proteomics dataset acquired on this difficult-to-cultivate parasite is presented for the zoonotic subtype T4 isolate WR1. Amongst the 2766 detected proteins, we highlighted the role of a small ADP ribosylation factor GTP-binding protein involved in intracellular traffic as major regulator of vesicle biogenesis and a voltage-dependent anion-selective channel protein because both were unexpectedly highly abundant. We show how these data may be used for gaining proteogenomics insights into Blastocystis sp. specific molecular mechanisms. We evidenced for the first time by proteogenomics a functional termination codon derived from transcript polyadenylation for seven different key cellular components.

Blastocystis spp.: avances, controversias y desafíos futuros / Blastocystis spp.: Advances, controversies and future challenges

Blastocystis spp. es el protista intracelular que se detecta con mayor frecuencia en muestras de materia fecal humana; las tasas de infección pueden superar el 20% en países en vías de desarrollo. El hallazgo de este parásito en heces de diversas especies animales sugiere su potencial zoonótico. La relevancia clínica y el papel patógeno de Blastocystis spp. en el tracto intestinal son inciertos. Varias son las publicaciones que lo reconocen como agente etiológico de desórdenes intestinales como diarrea, enfermedad inflamatoria intestinal y colitis ulcerosa, aunque la patogenicidad de este parásito no ha sido probada. Este amplio rango de respuestas a la infección podría estar relacionado con la diversidad genética de los aislamientos provenientes de hospedadores infectados.

Blastocystis spp. is the most common protozoan detected in human stool samples. In developing countries, infection rates are higher than 20%. The presence of this parasite in the feces of several host species suggests its zoonotic potential. The clinical relevance and the pathogenic role of Blastocystis spp. in the intestinal tract remain unclear. There are several clinical reports that recognize it as the etiologic agent of several intestinal disorders such as diarrhea, inflammatory bowel disease and ulcerative colitis, although the pathogenicity of this parasite has not been proved yet. This wide range of clinical manifestations could be related to the genetic diversity exhibited by this parasite.

[Blastocystis spp.: Advances, controversies and future challenges]. / Blastocystis spp.: avances, controversias y desafíos futuros.

Blastocystis spp. is the most common protozoan detected in human stool samples. In developing countries, infection rates are higher than 20%. The presence of this parasite in the feces of several host species suggests its zoonotic potential. The clinical relevance and the pathogenic role of Blastocystis spp. in the intestinal tract remain unclear. There are several clinical reports that recognize it as the etiologic agent of several intestinal disorders such as diarrhea, inflammatory bowel disease and ulcerative colitis, although the pathogenicity of this parasite has not been proved yet. This wide range of clinical manifestations could be related to the genetic diversity exhibited by this parasite.