Giardia intestinalis

Five diagnostic tests were compared for the diagnosis of Giardia duodenalis in fecal samples of young dogs. Fecal samples were collected from 136 healthy dogs < 1 year old and examined using immunofluorescence antibody microscopy (IFA) after sucrose gradient centrifugation, zinc sulfate centrifugal flotation technique (ZSCT), SNAP ® Giardia test, and ProSpecT ® Giardia EZ Microplate assay. In addition, polymerase chain reaction (PCR) of the 16S rRNA gene was performed. Kappa (κ) statistic was calculated to assess diagnostic agreement between the IFA and each test. Using the IFA as the gold standard, the relative sensitivity and specificity of each test were determined. Subsequently, a Bayesian approach was used to estimate the sensitivity and specificity of each test in comparison to the IFA results. Giardia duodenalis was detected in 41% of the samples examined by IFA. The ZSCT resulted in 37% of positive samples, with a relative sensitivity and specificity of 86 and 98%, respectively. The SNAP ® Giardia test was positive in 40% of the samples, with a relative sensitivity and specificity of 91 and 96%, respectively. The ProSpecT ® test was positive in 51% of the samples, with a relative sensitivity and spe-cificity of 100 and 83%, respectively. The relative sensitivity and specificity for PCR were 58 and 56%, respectively , with 55% of samples being PCR-positive. While the sensitivity and specificity estimates of each test in comparison to the IFA changed when using a Bayesian approach, the conclusions remained the same. While the ProSpecT ® test was the most sensitive test in this study, it is not designed for dogs and more costly than the other tests. The SNAP ® Giardia test performed similar to the ZSCT but may be more favorable because it is fast and easy to perform. Performance of the PCR was poor and the benefit of PCR may be in determining genotypes for evaluating zoonotic transfer between dogs and humans.

Zebra mussels (Dreissena polymorpha) from throughout the Shannon River drainage area in Ireland were tested for the anthropozoonotic waterborne parasites Cryptosporidium parvum, Giardia lamblia, Encephalitozoon intestinalis, E. hellem, and Enterocytozoon bieneusi, by the multiplexed combined direct immunofluorescent antibody and fluorescent in situ hybridization method, and PCR. Parasite transmission stages were found at 75% of sites, with the highest mean concentration of 16, nine, and eight C. parvum oocysts, G. lamblia cysts, and Encephalitozoon intestinalis spores/mussel, respectively. On average eight Enterocytozoon bieneusi spores/mussel were recovered at any selected site. Approximately 80% of all parasites were viable and thus capable of initiating human infection. The Shannon River is polluted with serious emerging human waterborne pathogens including C. parvum, against which no therapy exists. Zebra mussels can recover and concentrate environmentally derived pathogens and can be used for the sanitary assessment of water quality.

Giardia duodenalis (syn. G. intestinalis, or G. lamblia) is a leading cause of waterborne diarrheal disease that infects hundreds of millions of people annually. Research on Giardia has greatly expanded within the last few years, and our understanding of the pathophysiology and immunology on this parasite is ever increasing. At peak infection, Giardia trophozoites induce pathophysiological responses that culminate in the development of diarrheal disease. However, human data has suggested that the intestinal mucosa of Giardia-infected individuals is devoid of signs of overt intestinal inflammation, an observation that is reproduced in animal models. Thus, our understanding of host inflammatory responses to the parasite remain incompletely understood and human studies and experimental data have produced conflicting results. It is now also apparent that certain Giardia infections contain mechanisms capable of modulating their host’s immune responses. As the oral route of Giardia infection is shared with many other gastrointestinal (GI) pathogens, co-infections may often occur, especially in places with poor sanitation and/or improper treatment of drinking water. Moreover, Giardia infections may modulate host immune responses and have been found to protect against the development of diarrheal disease in developing countries. The following review summarizes our current understanding of the immunomodulatory mechanisms of Giardia infections and their consequences for the host, and highlights areas for future research. Potential implications of these immunomodulatory effects during GI co-infection are also discussed.

Giardia lamblia is a major cause of parasite-induced diarrhoea, which resides in the upper part of the small intestine lumen. Giardia is transmitted by the faecal-oral route, either directly or indirectly. This waterborne pathogen is highly prevalent in industrialized countries recent studies using PCR suggest that prevalence in the United States (USA) maybe as high as 22% with a high incidence of asymptomatic carriage. In Europe, similar studies suggest that prevalence in Romania may be as high as 91%. In the United Kingdom (UK), case reporting suggests that the prevalence of giardiasis is approximately 1.89% representing some 3% of all reported cases of gastro-intestinal infectious diseases. Two genetically distinct lineages (or assemblages) cause human disease. Giardiasis is caused when Giardia trophozoites of either assemblage attach to the intestinal epithelium. This can result in malabsorption, disruption of epithelial barrier function and apoptosis of intestinal epithelial cells. Giardia is a common cause of outbreaks of waterborne disease worldwide. In the US from 1991 to 2006 there were 13 outbreaks of Giardia linked to drinking water.
By employing an Ussing chamber, we demonstrated that trans-epithelial electrical resistance (TEER) of human colonic adenocarcinoma derived epithelial cell line (CaCo-2) monolayers decreased after 24 hours of co-culture with the parasite. When these cells were co-cultured with Giardia supernatants, inhibition of the CaCo-2 cell short-circuit current (Isc) in response to forskolin and UTP was observed. This inhibition was governed by the availability of Cl- channels as evidenced with Cl- channel inhibitors 4,4'-disothiocyanatostibene-2,2'-disulfonic acid (DIDS) and Glybenclamide (Gly101). The loss of transpethelial flux in response to the different inhibitors as well as the decrease of the CaCo-2 TEER raised the possibility of the presence of a soluble mediator. SDS-PAGE electrophoresis analysis of the Giardia supernatants identified a number of soluble proteins present in the culture supernatants from assemblages A and B. Disruption of endothelial function was then further investigated by fluorescent microscopy.
Our results suggest there is a soluble factor secreted by Giardia whose activity compromises enterocyte function and in particular the transcytosis of electrolytes. This enterocytes’ disruption by a soluble mediator seems to be quite similar to other well known enterotoxins, such as the potent cholera toxin (CT) produced by Vibrio cholera. Interestingly, these two different toxins are both involved in the disruption of chloride secretion. V.cholerae binds to the enterocytes’ plasma membrane and, releases an enzymatically active subunit that causes an increase of the cyclic-adenosine 5’-monophosphate (cAMP) production. cAMP is involved in chloride secretion by the Cystic Fibrosis conductance Regulator (CFRT) channel; the high level of intracellular cAMP causes a massive secretion of electrolytes and water into the intestinal lumen. Little or no structural damage is observed during a V.cholerae infection. Unlike V.cholerae toxin, Giardia soluble factor seems to induce a lot of structural damage such as brush border absorptive surface loss and diffuse shortening of villi. Even though Giardia soluble factor seems to be at the origin of the enterocytes’ structural changes and the increase of chloride secretion, its pathogenic mechanism requires further study.

Giardia intestinalis is a microaerophilic protozoan that is an important etiologic agent of diarrhea worldwide. There is evidence that under diverse conditions, the parasite is capable of shedding extracellular vesicles (EVs) which modulate the physiopathology of giardiasis. Here we describe new features of G. intestinalis EV production, revealing its capacity to shed two different enriched EV populations: large (LEV) and small extracellular vesicles (SEV) and identified relevant adhesion functions associated with the larger population. Proteomic analysis revealed differences in proteins relevant for virulence and host-pathogen interactions between the two EV subsets. We assessed the effect of two recently identified inhibitors of EV release in mammalian cells, namely peptidylarginine deiminase (PAD) inhibitor and cannabidiol (CBD), on EV release from Giardia and their putative effects on host-pathogen interactions. PAD-inhibitor Cl-amidine and CBD were both able to effectively reduce EV shedding, the PAD-inhibitor specifically affecting the release of LEVs and reducing parasite attachment to host cells in vitro. The strong efficacy of the PAD-inhibitor on Giardia EV release indicates a phylogenetically conserved pathway of PAD-mediated EV release, most likely affecting the Giardia arginine deiminase (GiADI) homolog of mammalian PADs. Our results suggest that LEVs and SEVs are differently involved in protozoa communication, and that treatment with EV-inhibitors may be a novel strategy for recurrent giardiasis treatment.