The present Project will apply a genome-wide approach in the investigation of two cases of host-parasite interactions, i.e. in a zoonotic subtype of the protist Blastocystis, and the nematode Anisakis pegreffii, and their accidental host (human). They are etiological agents of neglected "water/food-borne" zoonotic diseases whose occurrence in Italian patients is still underestimated. They both infect the gastrointestinal tract of humans. Further, A. pegreffii provokes allergic disease; it similarly happens in the infection with Blastocystis. It has been also supposed that infection with protozoans increases the intestinal permeability, leading to a major risk of allergic events even when in co-infection with helminths.
In spite of genetic heterogeneity and epidemiology so far investigated in the host-target parasite systems, knowledge on the association between host genetic susceptibility, parasite genome and gene expression analysis of molecules implicated in the pathogenesis correlated with the zoonotic infection of the target organisms of this Project, is still scarce.
The high-throughput "omics" technologies (genomics, transcriptomics, proteomics) are becoming very useful to explore the biology and pathogenicity role of parasites in their natural and accidental hosts. In this frame, the main goal of this project is the studying of the parasite-host interaction in the case of those zoonotic organism models. This will be reached using genomic and transcriptomic analyses, and by in vitro and in vivo studies.
The results achieved in the specific objectives (organized in WPs) will add knowledge on the qualitative and quantitative analyses of global genes possibly implicated in the interaction with the host, in the pathogenesis of diseases provoked by target parasites, as well as in human genetics of the host-susceptibility. This represents in some instances both an advance with respect to the state of the art and innovative aspects of the research on those parasites
The results achieved in the WPs will add knowledge on the qualitative and quantitative analysis of genes possibly implicated in pathogenesis of the diseases provoked by the target parasites, as well as in human genetics of the host-susceptibility. This represents in some instances both an advance with respect to the state of the art and innovative aspects of the research on those parasites.
WP1: The wide-genome sequencing of the zoonotic Blastocystis ST6, circulating in humans and animals in Italy, represents advance with respect to the state of the art. This approach will produce data sets to be compared with those gathered from other STs zoonotic and not-zoonotic (i.e. ST3); the last one is in progress by participants in the frame on another Project. The genome-wide data set will not only provide insights into the genome architecture of the target Blastocystis ST including the detection of genes responsible for the virulence factors, such as the proteases, protease inhibitors, but also the possible occurrence of HGT genes (horizontal gene transfers) which has been detected in other zoonotic Blastocystis STs. Further, the wide-genome library could allow studying simple sequence repeats to be used in genotyping of the ST target of this study, even correlated with its differential clinical manifestations observed in human patients.
Analogously, the genome-wide analysis of A. pegreffii will be an advanced result; it will help to identify gene families which could be implicated in the pathogenesis of human anisakiasis. Genome data set will also permit to detect genes possibly implicated in the biology of the species, including candidate genes in the parasite adaptation to their natural hosts (for instance, fish). Further, the obtained genome library of A. pegreffii will be also compared with that already partially known of A. simplex ss, providing further knowledge on the genotyping of the two Anisakis species, allowing to detect further diagnostic markers for their recognition, to be used in future research.
WP2: The comparative RNAseq analysis during in vitro studies of target parasites in presence of host cells will make possible to investigate the up-regulated genes coding for proteins implicated in the host-parasite interaction and in the pathogenesis provoked of both Blastocystis ST6 and A. pegreffii. This represents an innovative aspect in both the two species, which will also include the evaluation of the abilities of the studied parasites to trigger proinflammatory cytokine expression, even correlated to those up-regulated genes of the target parasites. Finally, the evaluation by qRT-PCR of those genes in vivo models, will help to find molecular effectors at the host-parasite interface which could be used in future proteomics as biomarkers of infection during the early pathogenesis.
WP3: The comparison between microbiota profile from Blastocystis ST6 infected/not-infected rats will represent and advance with respect to the state of the art in the relationships between microbiota and Blastocystis occurrence and patients' symptoms. Even more, the association with Anisakis represents an innovative aspect concerning the possible occurrence of allergic diseases as a consequence of the possible relationship among a common protist of the human intestinal tract (i.e. Blastocystis), the occurrence of the nematode Anisakis, and the human gut microbiota. This would provide information about the possible increase of the intestinal permeability provoked by the protozoan, the occurrence of a high IgE hypersensitivity to Anisakis, producing allergic diseases, including Chronic Urticaria (CU).
WP4 A genetic epidemiological investigation aimed to disentangle a potential host-genetic architecture of allergic disorders, even IgE-hypersensitivity associated with the target parasites was never been investigated in Italian patients.
The potential finding of the correlation between alleles/genotypes in the genes coding for major allergens of the target parasites will also contribute to enlarge the knowledge around the molecular mechanisms which are at the base of the host-parasite interaction.
The potential significant association between host genetic factors and IgE hypersensitization to Anisakis, will constitute a genetic data sets to be used in the comparative analysis performed on other Mediterranean populations from where also a high IgE-sensitization to the parasite Anisakis, has been reported.
The host-genetic perspective constitutes a novel approach to be considered in the research on these parasitic zoonoses. The potential association between HLA-DRB1/DBQ1 alleles, which will be detected in those patients showing IgE sensitization and CU occurrence, will allow identifying a "genetic risk factor" in the human manifestation of the zoonoses. This would expose humans having those specific alleles to encounter higher pathologic events during infection.