Immune response against Giardia duodenalis infection in cattle

Giardia duodenalis is een belangrijke parasiet bij mensen en dieren. In runderen kan G. duodenalis diarree en gewichtsverlies veroorzaken en infecties zijn meestal chronisch. Dit suggereert dat Giardia in staat is om het immuunsysteem van de gastheer te onderdrukken of te ontwijken. Met uitzondering van G. duodenalis en G. muris in het muis-model, is de kennis over de immuniteit tegen Giardia beperkt. Daarom is de doelstelling van deze thesis om de immuunrespons tegen G. duodenalis te onderzoeken in runderen en om potentiele mechanismen te identificeren die zorgen voor het chronische verloop van giardiosis in de natuurlijke gastheer. In muizen onderdrukt G. duodenalis de respons van dendritische cellen (DC) op stimulatie van de Toll-like receptor liganden. Dit suggereert dat G. duodenalis een immuun-modulerend effect heeft op DC. Daarom werd in het eerste deel van deze thesis het effect van G. duodenalis onderzocht op DCs, afkomstig van monocyten van het rund (MoDC). Ook werd nagegaan of de parasiet in staat is om via MoDC verdere adaptieve immuunresponsen te induceren en te beïnvloeden. De maturatie en cytokineproductie van runder-MoDC na incubatie met G. duodenalis werd onderzocht, alsook de capaciteit van Giardia-gestimuleerde MoDC om lymfocytproliferatie te induceren. MoDC gestimuleerd met G. duodenalis waren functioneel actief en in staat om proliferatie van T-cellen te induceren, die zowel pro- als anti-inflammatoire cytokines produceren. G. duodenalis was niet in staat om de respons van de MoDC na stimulatie van TLR4 door LPS, te beïnvloeden. Er werd geen bewijs gevonden dat G. duodenalis een regulerend of inhiberend effect heeft op runder MoDC. In het tweede deel van deze thesis werd de kinetiek van de immuunrespons tegen G. duodenalis onderzocht in runderen, om het mechanisme te identificeren dat het chronische karakter van G. duodenalis infecties in runderen kan verklaren. Zes kalveren werden experimenteel geïnfecteerd met G. duodenalis, waarna de cysten-excretie, de kinetiek van de circulerende geheugencellen en van serum-antilichamen werd gevolgd. De cysten-excretie startte een week na de infectie en bleef hoog tot week 14, waarna een continue lage uitscheiding aangaf dat de dieren immuniteit ontwikkeld hadden. Vijf weken na de start van de infectie werd een significante proliferatie van CD4+ αβ T-cellen in het bloed gemeten. Tijdens de piek van de proliferatie werd door deze cellen vooral IL-17 tot expressie gebracht. Serum concentraties van anti-G. duodenalis IgG1 en IgA stegen significant vanaf week 11. Serum antilichamen waren vanaf de start van de infectie gericht tegen alle variant-specifieke oppervlakte eiwitten (vsp) en niet enkel tegen bepaalde subpopulaties. Dit duidt er op dat bescherming tegen G. duodenalis niet alleen gebaseerd is op de herkenning van alle vsp door antilichamen, maar ook op de antilichaamconcentratie. Deze data wijzen erop dat de immuun-respons in runderen tegen G. duodenalis traag verloopt en waarschijnlijk onderdrukt wordt door een nog onbekend mechanisme. Toekomstig onderzoek zal moeten gericht zijn op de inductie van een IL-17 respons, die mogelijk geassocieerd is met protectie

Comparative immune responses against Psoroptes ovis in two cattle breeds with different susceptibility to mange

The sheep scab mite, Psoroptes ovis, is a major problem in the beef cattle industry, especially in Belgian Blue (BB) cattle. This breed is naturally more predisposed to psoroptic mange but reasons for this high susceptibility remain unknown. Different immune responses could be a potential cause; thus in this study, the cutaneous immune response and in vitro cellular immune response after antigen re-stimulation were examined in naturally infested BB. Cytokine production in the skin and in circulating re-stimulated peripheral blood mononuclear cells (PBMC) demonstrated a mixed pro-inflammatory Th2/Th17 profile, with transcription of IL-4, IL-13, IL-6 and IL-17. Strong IL-17 up-regulation in the skin of BB was associated with an influx of eosinophils and other immune cells, potentially leading towards more severe symptoms. Virtually no changes in cutaneous IFN-gamma transcription were detected, while there was substantial IFN-gamma up-regulation in re-stimulated PBMC from infested and uninfested animals, potentially indicating a role of this pro-inflammatory cytokine in the innate immune response. In Holstein-Friesian (HF) cattle, generally more resistant to P.ovis infection, a largely similar immunologic response was observed. Differences between HF and BB were the lack of cutaneous IL-17 response in infested HF and low transcription levels of IFN-gamma and high IL-10 transcription in re-stimulated PBMC from both infested and uninfested animals. Further research is needed to identify potential cell sources and biological functions for these cytokines and to fully unravel the basis of this different breed susceptibility to P. ovis

Occurrence and potential health risk of Cryptosporidium and Giardia in different water catchments in Belgium

Human wastewater and livestock can contribute to contamination of surface water with Cryptosporidium and Giardia. In countries where a substantial proportion of drinking water is produced from surface water, e.g., Belgium, this poses a constant threat on drinking water safety. Our objective was to monitor the presence of Cryptosporidium and Giardia in different water catchment sites in Belgium and to discriminate between (oo) cysts from human or animal origin using genotyping. Monthly samples were collected from raw water and purified drinking water at four catchment sites. Cryptosporidium and Giardia were detected using USEPA method 1623 and positive samples were genotyped. No contamination was found in purified water at any site. In three catchments, only low numbers of (oo) cysts were recovered from raw water samples (<1/liter), but raw water samples from one catchment site were frequently contaminated with Giardia (92 %) and Cryptosporidium (96 %), especially in winter and spring. Genotyping of Giardia in 38 water samples identified the presence of Giardia duodenalis assemblage AI, AII, BIV, BIV-like, and E. Cryptosporidium andersoni, Cryptosporidium suis, Cryptosporidium horse genotype, Cryptosporidium parvum, and Cryptosporidium hominis were detected. The genotyping results suggest that agriculture may be a more important source of surface water contamination than human waste in this catchment. In catchment sites with contaminated surface water, such as the Blankaart, continuous monitoring of treated water for the presence of Cryptosporidium and Giardia would be justified and (point) sources of surface water contamination should be identified

Galectin-11 induction in the gastrointestinal tract of cattle following nematode and protozoan infections

Galectin-11 (LGALS11) has been suggested to play an important role in protective immunity against gastrointestinal nematodes in ruminants. However, in cattle, this molecule has not been characterized in detail. In the current study, it was shown that transcription of LGALS11 was highly inducible in the bovine abomasal mucosa after an Ostertagia ostertagi infection. LGALS11 protein expression was also increased in the abomasal mucosa following O. ostertagi infection and localized to the nucleus and cytoplasm of epithelial cells and the mucus. Using in vitro abomasal epithelial cell cultures, it was shown that LGALS11 induction was associated with the proliferative and dedifferentiated status of cells. However, LGALS11 was not induced following stimulation with O. ostertagi excretory-secretory products. These results suggest that LGALS11 induction in vivo may be an indirect rather than a direct effect of the parasite on the epithelium. In addition, LGALS11 transcript was also detected in the abomasal lymph nodes where it was shown to be transcribed in MHCII(+) cells; however, transcription levels in the lymph nodes were not altered after O. ostertagi infection. In addition, LGALS11 was also induced in the small intestine by different types of parasites, including the nematode Cooperia oncophora and the protozoan parasite Giardia duodenalis

Giardia duodenalis stimulates partial maturation of bovine dendritic cells associated with altered cytokine secretion and induction of T-cell proliferation

Giardia duodenalis is an important intestinal parasite in animals and humans. The role of dendritic cells (DC) in the initiation of the immune response against G.duodenalis is poorly documented. The aim of this study was to test the hypothesis that G.duodenalis interferes with bovine DC function. Therefore, the effect of trophozoites and excretion/secretion products on bovine monocyte-derived dendritic cells (MoDC) was investigated. We assessed MoDC maturation and cytokine production of G.duodenalis-stimulated MoDC and the ability of these MoDC to take up antigen and induce lymphocyte proliferation. Little or no upregulation of maturation markers CD40 and CD80 was measured, but MHCII expression was increased after stimulation with low parasite concentrations. A dose-dependent decrease in ovalbumin uptake was observed in G.duodenalis-stimulated MoDC. In addition, stimulated MoDC induced proliferation of CD3(-), -T-cells and TCR+ CD4(+) and CD8(+) T-cells. Increased transcription of TGF- was shown in CD4(+) T cells, and increased TNF-, TGF-, IL-10 and IL-4 were seen in -T-cells. We found no evidence that G.duodenalis has a regulatory or inhibitory effect on bovine MoDC. MoDC stimulated with G.duodenalis are functionally active and able to induce proliferation of T cells that produce both pro- and anti-inflammatory cytokines

Reduced expression of maturation markers in bovine dendritic cells after incubation with Giardia duodenalis trophozoites

Giardia duodenalis (syn. G. intestinalis, G. lamblia) is an important intestinal parasite in a wide range of mammals, including farm animals, companion animals and humans. The role of dendritic cells in the immune response against G. duodenalis is poorly documented and has only been studied in the mouse, which is not a natural host for this parasite. The objective of this study was to investigate whether G. duodenalis trophozoites or their excretion/secretion (ES) products activate and mature dendritic cells from cattle in vitro. Bovine monocyte-derived dendritic cells (MoDCs) were incubated with different concentrations of G. duodenalis trophozoites. An effect of ES products was investigated by preventing direct contact between the trophozoites and the MoDCs, using 0.4μm cell culture inserts. After 18h of stimulation, expression of the maturation markers CD80, CD40 and MHCII was measured and ovalbumin uptake was monitored. A dose-dependent decrease of ovalbumin uptake was observed in MoDCs incubated with trophozoites (but not ES), suggesting functional maturation. However, none of the maturation markers were upregulated after incubation of MoDCs with Giardia trophozoites or ES. In order to investigate whether or not MoDCs can induce lymphocyte proliferation after incubation with Giardia trophozoites, a mixed lymphocyte reaction (MLR) will be performed with allogenic CD2+ lymphocytes. A selection of cytokines will be determined by ELISA in the culture supernatant of the stimulated MoDCs before and after incubation with CD2+ lymphocytes, to determine the phenotype of the immune response that is induced