Blastocystis is a single-celled intestinal monoxenus parasite with fecal-oral transmission. It represents the most prevalent intestinal parasites in humans from all over the world, including from Italy. To date, based on the SSU rDNA and MLST loci analysis, it was discovered to comprise 17 subtypes (STs), likely representing distinct species, colonizing humans, other mammals, birds, reptiles and amphibians. Some STs from animals are zoonotic to humans. Currently, there are no data on the molecular epidemiology of blastocystosis in domestic and farmed animals, which could represent the source of infection to humans in Italy.
The pathogenesis of STs still remains uncertain. Recent findings indicate that some STs of Blastocystis could be considered as "markers" of gastrointestinal health, rather than of disease. Further, so far, little is known about whether and how STs alter the faecal bacterial microbiota.
The Project aims to continue to perform the research on the argument, with following objectives: 1) to carry out molecular epidemiological study of Blastocystis STs on: a) human groups (patients) from Italy, both immunocompetent and immunodepressed ones; b) domestic, farmed and sylvatic animals; 2) compare the faecal microbiota in human groups infected with STs; 3) develop a molecular method, for the rapid and specific molecular detection of STs in humans; 4) detect protease released by the parasite and their gene expression level, implicated in the pathogenicity of distinct STs.
Results that will be obtained will allow to: a) add knowledge on the occurrence of STs in animals as possible sources of blastocystosis to humans, in Italy; b) study the association between STs, faecal microbiota and intestinal disorders in different patients groups; c) use a rapid, sensitive and specific tool in the diagnosis of STs in humans; d) detect target proteins (proteases) which could be also used as a possible "biomarker" of pathogenicity of different STs, in humans.
The occurrence of Blastocystis infected animals will allow to explain the result of an infection in humans in Italy. Thus, the results achieved on domestic and farmed animals will contribute to the knowledge about the risk of transmission to humans. Preliminary studies in this contest already achieved by the PI, has indeed evidenced the occurrence of zoonotic STs from animals. The present Project aim to cover as much as possible, the source of human infections in Italy. This aspect has never been investigated in Italy, and thus the results which will be achieved will represent an advance with respect to the state of the art.
It has been has suggested that other human variables, are determining the ability of Blastocystis STs to colonize the human gut. For example, the human gut flora could have an impact. For instance, the present Project, is also aiming to investigate the carriage of Blastocystis is associated with differences in the faecal microbiota, as a proxy of gut microbiota, and what might be the impact of this differences on gut homeostasis. Studies on the gut microbiota in humans with and without Blastocystis, are likely to provide valuable information to help in determine the role of Blastocystsis in human health and disease. Populations among immunocompromised patients from as well as the possible association of different STs with faecal microbiota in subjects with HIV infection and their possible implications to gut homeostasis, have been preliminary studied by the PI and collaborators. Other human immunocompromised populations will be studied, including patients with Lyme disease. The faecal microbiota and STs in people from two different epidemiological scenarios will be compared, to our knowledge, for the first time in Italy.
The Project aimed to develop a rapid, sensitive and specific method based on a qRT-PCR using STs-specific primers and probes, for the detection of various STs in humans stools, will represent a diagnostic advance with respect to the conventional PCR-DNA, also in terms of less time and money consuming. This aspect will represent also a strong advantage to the screening tools used routinely in human diagnostic at the UOS of Parasitology at Policlinico Umberto I.
The genetic characterization of the subtypes from stool samples from both humans and animals will allow to implement the epidemiological data concerning the occurrence of subtypes in animals (domestic and sylvatic), having also a zoonotic role to humans. In addition, the estimates of genetic polymorphism at the intrasubtype level, as preliminarly observed in some STs (Mattiucci et al., 2016), will be able to detect a possible association, in a multivariate analysis, between distinct haplotypes and degree of intestinal diseases in infected subjects (humans).
Finally, because the pathogenesis of STs, remains as a controversial issue, the gene expression analysis of some target proteins (i.e. Proteases) having a major role in the pathogenicity, which will be detected in the present study, will allow to evidence a possible differential pathogenic role of STs, to humans. This aspect will allow in the future to add further diagnostic markers, out of PCR-DNA in the infection by STs, such as the RT-RNA analysis of some target proteins could be also used as a "biomarkers" of distinct ST affecting humans.