Project description:Giardia duodenalis is a prevalent intestinal pathogens known to cause giardiasis, a condition characterized by diarrhea and frequently linked to malnutrition and growth impairments in children. The presence of Giardiavirus (GLV) in Giardia strains has been associated with heightened immune cytokine responses in the host compared to the GLV-free strains. However, the transmission mode and biological significance of GLV remain unclear. In this study, by using G. duodenalis DH (GLV-containing) and WBC6 (GLV-free) strains, we demonstrated that the DH strain produced extracellular vesicles (EVs), which originated from unique peripheral vesicle and bead-like structures in the ventrolateral flange. Nanoparticle tracking analysis revealed that GLV infected-G. duodenalis DH strain secreted fewer EVs than the GLV-free WBC6 strain. Biochemical and electron microscopy demonstrated that GLV virions can exploit the Giardia EVs pathway to facilitate their spread among parasites. Giardia uptake of GLV-containing EVs occured through clathrin-mediated endocytosis, leading to rapid infection of trophozoites by GLV. Furthermore, GLV infection enhanced messenger RNA translation efficiency, influencing protein abundance in Giardia trophozoites. The presence of GLV also upregulated the glycolytic pathway, with Giardia enolase closely associated with GLV replication. Importantly, Giardia infected with GLV alleviated the pathogenicity of parasite compared to the GLV-freeGiardia strain. These findings highlight the pivotal role of GLV in regulating Giardia biology, suggesting its potential for informing the development of novel intervention strategies against Giardia infections.
Project description:To elucidate the impact of GLV infection, we conducted a comparative analysis of the total protein profiles of tachyzoite samples. By analysing the mass spectrometry (MS) data from G. duodenalis DHGLV, using the G. duodenalis WBC6 isolate database as a reference, proteomics analyses revealed the actual protein species identified from the proteomics data were 1274 in the Giardia WBC6-GLV strain and 1494 in the Giardia DH+GLV strain.
Project description:Despite of Giardia duodenalis being one of the most commonly found intestinal pathogens in humans and animals, little is known of the host-parasite interactions in natural hosts. Therefore, the objective of this study was to investigate the intestinal response in calves following a G. duodenalis infection, using a bovine high-density oligo microarray to analyze global gene expression in the small intestine. The resulting microarray data suggested a decrease in inflammation, immune response and immune cell migration in infected animals, which was examined in more detail by quantitative real-time PCR on a panel of cytokines combined with histological analyses. The cytokine transcription levels showed a trend of down regulated expression in infected animals compared to the negative controls, best seen in jejunum for IL-6 and IL-8 and statistically significant for IL-17, IL-13 and IFN-?. No increased immune cell recruitment could be seen after infection, as well as no intestinal pathologies, such as villus shortening or increased levels of apoptosis. Key regulators in this intestinal response seem to be the nuclear peroxisome proliferator-activated receptors alpha (PPARA) and gamma (PPARG), for which an up-regulated expression was seen on microarray and qRT-PCR data. The activation of PPARs can exert an anti-inflammatory effect with inhibition of pro-inflammatory cytokines and a decrease in cell recruitment. . How the PPARs are activated during a Giardia infection still needs to be further elucidated. Eight male Holstein calves aged two to four weeks old were used for the trial. Prior to arrival, all animals were screened for the presence of Giardia cysts in their faecal samples. After confirming their negative status for all these pathogens, four of the animals were randomly chosen and placed in a G. duodenalis contaminated environment, whereas the four remaining animals were kept as negative controls in separate G. duodenalis-free stables. All calves in the study received the same commercial milk replacer. After three weeks, the presence or absence of a G. duodenalis infection was confirmed by IFA on faecal samples after which the animals were euthanized. Changes in gene expression profiles induced by Giardia duodenalis infection were compared using a high-density 60mer bovine oligo microarray.
Project description:Giardia duodenalis is a protozoan parasite responsible for gastroenteritis in vertebrates, including humans. The prevalence of G. duodenalis is partly owed to its direct and simple life cycle, as well as the formation of the environmentally resistant and infective cysts. Several proteomic and transcriptomic studies have previously analysed global changes during the encystation process using the well-characterised laboratory isolate and genome strain, WBC6. To expand current comparative analyses, this study presents the first quantitative global study of encystation using pathogenically relevant and alternative assemblage A strains: the human-derived BRIS/82/HEPU/106 and avian-derived BRIS/95/HEPU/2041. We have utilised tandem MS/MS with a label-free quantitative approach to compare cysts and trophozoite life stages between strains for variation, as well as confirm universal encystation markers of Assemblage A.
2015-06-10 | PXD002002 | Pride
Project description:Giardia duodenalis assemblage B whole genome comparative study
Project description:Giardiasis, caused by the protozoan parasite Giardia duodenalis, is an intestinal diarrheal disease affecting almost one billion people worldwide. A small endosymbiotic dsRNA viruses, G. lamblia virus (GLV), genus Giardiavirus, family Totiviridae, might inhabit human and animal isolates of G. duodenalis. Three GLV genomes have been sequenced so far, and only one was intensively studied, moreover a positive correlation between GLV and parasite virulence is yet to be proved. To understand the biological significance of GLV infection in Giardia, the characterization of several GLV strains from naturally infected G. duodenalis isolates is necessary. In the contest of high-throughput sequencing of four GLVs strains, from Giardia isolates of human and animal origin, we report on a new, unclassified viral sequence (designed GdRV-2), unrelated to Giardiavirus, encoding and expressing for a single large protein with a RdRp domain homologous to Totiviridae and Botybirnaviridae. We have analyzed the sequence of the GLV capsid protein (CP) and RNA-dependent RNA polymerase (RdRp) by LC-MS/MS analysis using different enzymatic strategies.
Project description:Giardia duodenalis a species-complex of common gastrointestinal protists of major medical and veterinary importance. This complex is currently subclassifed as ‘Assemblages’, with Assemblage A and B infective to humans. To date, post-genomic proteomics are derived exclusively from Assemblage A, biasing understanding of these parasites’ biology. This bias is particularly notable, as Assemble B is the more prevalent cause of human infections. To address this gap, we quantitatively analysed proteomes of the intestinal ‘trophozoite’ stage of three Assemblage B isolates, including the genome reference (GS/M) and two clinical isolates (BRIS/91/HEPU/1279 and BRIS/92/HEPU/1487), during in vitro axenic culture. We used spectrum-to-peptide matching metrics to infer currently unknown intra-assemblage variation. We identified and quantified over 3000 proteins in the GS isolate, but demonstrated significant isolate-dependent losses in peptide and protein identifications in non-reference isolates, suggesting significant intra-assemblage variation. We also explore differential protein expression between in vitro cultured subpopulations enriched for dividing relative to feeding cells. This data is an important proteomic baseline for Assemblage B, and highlights unique differences heretofore avoided in post-genomic Giardia proteomics.