Giardia Lamblia Trophozoites Research Articles

Electron and Video-Light Microscopy Analysis of the in Vitro Effects of Pyrantel Pamoate on Giardia lamblia

Campanati, L., Gadelha, A. P. R., and Monteiro-Leal, L. H. 2001. Electron and video-light microscopy analysis of the in vitro effects of pyrantel pamoate on Giardia lamblia. Experimental Parasitology97, 9–14. Giardia infection is predominant in the small intestine of vertebrates, where the trophozoites attach to epithelial cells and adversely affect the microvilli and other epithelial cell structures. Giardiasis, the disease caused by this protozoan, is very common in developing countries and mainly affects children. Drugs currently used to treat Giardia infection, such as some benzimidazole derivatives, were originally designed to treat helminthic infections. Many of the drugs are known to cause severe side effects and disturbances to the patient. Using transmission electron microscopy and video-light microscopy, we studied the effects of pyrantel pamoate, a drug commonly used in the treatment of helminthic infections in horses and ruminants, on Giardia lamblia trophozoites. Pyrantel pamoate was administered to Giardia cells in four different concentrations. Using video-light microscopy, we observed the decrease in flagella beating frequency and severe changes in the lateral flange and in the general aspect of the cell. Using transmission electron microscopy, we observed changes in the cytoplasm and peripheral vesicles. The flagella and adhesive disk structure were not affected. Apparently, the effects of pyrantel pamoate are irreversible.

Human T-lymphotropic virus 1 adult T-cell lymphoma with Giardia and Strongyloides parasitism.

A man originally from Togo, West Africa, developed a 2-month history of abdominal cramps, diarrhea, and weight loss, 4 years after immigrating to Canada. The patient was seropositive for human T-lymphotropic virus 1 (HTLV-1). No human immunodeficiency virus 1 or human immunodeficiency virus 2 antibodies were detected. The patient’s complete blood count values were within laboratory reference intervals, and there specifically was no evidence of adult T-cell leukemia. Upper endoscopy revealed only esophageal candidiasis, which was confirmed histologically. Duodenal biopsies contained clusters of lymphocytes with large pleomorphic nuclei that infiltrated mucosa, including glandular epithelium, and submucosa (Figure, large arrow). These lymphocytes stained with antibodies directed against CD45RO and showed no immunoreactivity for CD20. Adult T-cell lymphoma was diagnosed and was also found involving the stomach. In addition to the diffuse neoplastic process, Giardia lamblia (small arrow) and Strongyloides stercoralis parasites were identified in duodenal biopsies. Occasional cytomegalovirus inclusions within gastric glandular epithelium were also present. Stool examination for ova and parasites revealed S stercoralis rhabditiform larvae, eggs, and parasitic females. Giardia lamblia cysts and trophozoites were also identified. No filariform Strongyloides larvae were present. Human T-lymphotropic virus 1 is an oncogenic retrovirus with a global distribution that is endemic to Southeast Asia, southwestern Japan, the Caribbean, the southeastern United States, and West Africa.1,2 Environmental cofactors, such as Strongyloides parasitism, have been suggested as possible contributing mechanisms to HTLV-1– associated leukemogenesis.3 It is controversial whether HTLV-1 carriers are at increased risk for Strongyloides infection.1 Human T-lymphotropic virus 1–infected patients with adult T-cell leukemia/lymphoma, however, are immunocompromised and specifically have defects of cellmediated immunity.2 These patients are at increased risk for multiple opportunistic infections, including cytomegalovirus, herpes zoster, disseminated herpes simplex, Pneumocystis carinii, Cryptococcus, Candida, and enteric parasites, such as Strongyloides.2,4 Immunocompromised hosts with Strongyloides infections may develop autoinfection and disseminated strongyloidiasis as a result of direct transformation of rhabditiform larvae to filariform larvae

Role of exogenous inositol and phosphatidylinositol in glycosylphosphatidylinositol anchor synthesis of GP49 by Giardia lamblia

Although Giardia lamblia trophozoites are unable to carry out de novo phospholipid synthesis, they can assemble complex glycophospholipids from simple lipids and fatty acids acquired from the host. Previously, we have reported that G. lamblia synthesizes GP49, an invariant surface antigen with a glycosylphosphatidylinositol (GPI) anchor. It is therefore possible that myo-inositol (Ins), phosphatidylinositol (PI) and other GPI precursors are obtained from the dietary products of the human small intestine, where the trophozoites colonize. In this report, we have investigated the role of exogenous Ins and PI on GPI anchor synthesis by G. lamblia. The results demonstrate that [ 3H]Ins and PI internalized by trophozoites, metabolically transformed into GlcN(acyl)-PI and downstream GPI molecules. Further investigations suggest that G. lamblia expresses cytidine monophosphate (CMP)-dependent (Mg 2+-stimulated) and independent (Mn 2+-stimulated) inositol headgroup exchange enzymes, which are responsible for exchanging free Ins with cellular PI. We observed that 3-deoxy-3-fluoro- D- myo-inositol (3-F-Ins) and 1-deoxy-1-F- scyllo-Ins (1-F- scyllo-Ins), which are considered potent inhibitors of Mn 2+-stimulated headgroup exchange enzyme, inhibited the incorporation of [ 3H]Ins into PI and GPI molecules significantly, suggesting that CMP-independent (Mn 2+-stimulated) exchange enzyme may be important for these reactions. However, 3-F-Ins and 1-F- scyllo-Ins were not effective in blocking the incorporation of exogenously supplied [ 3H]PI into GPI glycolipids. Thus, it can be concluded that G. lamblia can use exogenously supplied [ 3H]PI and [ 3H]Ins to synthesize GPI glycolipids of GP49; while PI is directly incorporated into GPI molecules, free Ins is first converted into PI by headgroup exchange enzymes, and this newly formed PI participates in GPI anchor synthesis.

Trophozoites of Giardia lamblia may have a Golgi-like structure.

Trophozoites of the primitive protozoan Giardia lamblia have been considered as cells which do not present the Golgi complex. Using C(6)-NBD ceramide, which has been shown to label the Golgi complex of mammalian cells, labelling of the perinuclear region of G. lamblia was observed by confocal laser scanning microscopy. Transmission electron microscopy of thin sections and of replicas of freeze-fractured cells revealed the presence of concentric perinuclear membranes resembling the Golgi complex.

A new method for assessing metronidazole susceptibility of Giardia lamblia trophozoites.

A quantitative, simple, and rapid assay has been developed to assess Giardia lamblia trophozoite sensitivity to metronidazole [1-(2-hydroxyetyl)-2-methyl-5-nitroimidazole] (MTZ). This new assay utilizes the ability of live (surviving) trophozoites to take up oxygen after have been exposed to MTZ. The effect of MTZ on oxygen uptake was compared with its effect on viability as evaluated by a culture method and morphological assays. Oxygen uptake rates decreased in trophozoites treated with MTZ, and this effect was drug concentration dependent: O(2) uptake rates went from 3.04 microM O(2) min(-1) per 10(6) cells to 0.72 microM O(2) min(-1) per 10(6) cells with increasing drug concentration (0.15 to 0.6 mM) in the preincubation. Concentrations of the drug which inhibited oxygen uptake by 28 to 76% in trophozoites killed from 39 to 82% of trophozoites, as evaluated by the culture method, and altered the morphology of 21 to 86% of the trophozoites. Thus, the trophozoites killed by MTZ are nonmotile cells and do not take up oxygen. A good correlation was found between the inhibitory effects of MTZ, as evaluated by oxygen uptake, and cellular viability. Similar 50% inhibitory concentrations were obtained: 0.33 mM by oxygen uptake, 0. 26 mM by the culture method, and 0.35 mM by morphological criteria. Oxygen uptake appears to be a good indicator of parasite viability. Therefore, this new method can provide a convenient means to assess MTZ susceptibility in G. lamblia and can be applied for screening potential antigiardial agents.

Trophozoites of Giardia lamblia may have a Golgi-like structure

Trophozoites of the primitive protozoan Giardia lamblia have been considered as cells which do not present the Golgi complex. Using C 6-NBD ceramide, which has been shown to label the Golgi complex of mammalian cells, labelling of the perinuclear region of G. lamblia was observed by confocal laser scanning microscopy. Transmission electron microscopy of thin sections and of replicas of freeze-fractured cells revealed the presence of concentric perinuclear membranes resembling the Golgi complex.

Presence of a protrusion on the ventral disk of adhered trophozoites of Giardia lamblia.

About 30% of the trophozoites of Giardia lamblia, fixed while adherent to the substrate using fixative solutions designed to better preserve cytoskeletal elements, showed the presence of a ventral disk. This structure varied in shape and size and could be seen by scanning and transmission electron microscopy, as well as by confocal laser scanning microscopy of cells incubated with 3-3'-dihexyloxacarbocyanine iodide, which labels cisternae of the endoplasmic reticulum. It could also be observed in living adherent cells. A row of microtubules, a large number of glycogen particles, peripheral vesicles and concentric membranes were seen within the protrusion.

Cytotoxicity Induced by Bismuth Subcitrate in Giardia lamblia Trophozoites

The cytotoxicity of colloidal bismuth subcitrate (CBS) was investigated in cultured Giardia lamblia trophozoites on the basis of cell attachment, morphology and viability studies. The effects on cell membrane integrity were evaluated by the permeability to trypan blue, and the morphological alterations were studied by phase-contrast microscopy and transmission electron microscopy. Our data show that although CBS induced loss of cellular viability (morphological and regrowth studies), the cell membrane permeability was not altered. The attachment of G. lamblia trophozoites to culture vials was rapidly disrupted by CBS. Ultrastructural observations revealed that this drug promoted modifications on the cell shape, displacement of nucleus and of cytoskeletal structures, pronounced cytoplasmic vacuolization, dorsal and ventral protuberance of cytoplasmic membrane and heavy deposition of electron-dense precipitates in adhesive disc, nuclear membrane and cytoplasmic components. In contrast, membranes and microtubules were apparently undamaged. Some of these results suggest that the cytotoxicity of CBS to G. lamblia cultures is the result of its binding to cytoskeletal components. As far as we know, this work is the first demonstration that bismuth subcitrate could kill a human protozoan parasite suggesting its potential in the therapy of giardiasis.

Giardia lamblia:Incorporation of Free and Conjugated Fatty Acids into Glycerol-Based Phospholipids

Gibson, G. R., Ramirez, D., Maier, J., Castillo, C., and Das, S. 1999.Giardia lamblia:Incorporation of free and conjugated fatty acids into glycerol-based phospholipids.Experimental Parasitology92, 1–11.Giardia lambliatrophozoites are flagellated protozoa that inhabit the human small intestine, where they are exposed to various dietary lipids and fatty acids. It is believed thatG. lamblia, which colonizes a lipid-rich environment of the human small intestine, is unable to synthesize phospholipids, long-chain fatty acids, and sterolsde novo. Therefore, it is possible that this protozoan has developed a special process for acquiring lipids from its host. We have previously shown thatG. lambliacan take up saturated fatty acids and incorporate them into phosphatidylglycerol (PG) and other glycerol-based phospholipids (Stevenset al., Experimental Parasitology,86, 133–143, 1997). In the present study, an attempt has been made to investigate the underlying mechanisms of transesterification and interesterification reactions of giardial phospholipids by free and conjugated fatty acids. Results show that exogenously supplied, unsaturated, fatty acids were taken up byGiardiaand incorporated into various phosphoglycerides, including PG. To test whether this intestinal pathogen can utilize conjugated fatty acids, live trophozoites were exposed to either [3]H;cbphosphatidylcholine (PC), where the fatty acid was3H-labeled at its sn2position, or to [14C]lyso-PC (fatty acid was14C-labeled at the sn1position) for 90 min, followed by phospholipid analysis using thin-layer chromatography. The results suggest that conjugated fatty acids, like free fatty acids, were incorporated into PG. It was also observed that aristolochic acid, an inhibitor of Ca2+-ionophore-stimulated phospholipase A2, decreased the transfer of fatty acids from [3H]PC to PG, indicating that giardial phospholipases were involved in these esterification reactions. Additional experiments, which include culturing trophozoites in serum-supplemented and serum-deprived medium, along with numerous biochemical analyses suggest that (i) PG is a major transesterified and interesterified product, (ii) it is likely that giardial phospholipases are involved in esterification reactions, (iii) inG. lamblia, PG is localized in perinuclear membranes, as well as intracellularly, but not in the plasma membrane, and (iv) various synthetic analogs of PG inhibit the growth of the parasitein vitro. These studies suggest that PG is an important phospholipid ofGiardiaand a potential target for lipid-based chemotherapy against giardiasis.

Detection of a giardia lamblia coproantigen by using a commercially available immunoenzymatic assay, in belo horizonte, brazil

It is known that fecal examination to detect Giardia lamblia cysts or trophozoites produces a high percentage of false-negative results. A commercially available immunoenzymatic assay (ProSpecT Giardia Microplate Assay, Alexon, Inc., BIOBRAS) to detect G. lamblia specific coproantigen was evaluated for the first time in Brazil. A total of 90 specimens were tested. Each specimen was first tested as unpreserved stool, and then it was preserved in 10% Formalin to be tested 2 months later. The assay was able to identify all the 30 positive patients (sensitivity = 100.0%) by visual or spectrophotometric examination in the unpreserved specimens and was negative in 57 of the 60 patients without G. lamblia (specificity = 95.0%). The assay identified 27 of the 30 positive patients (sensitivity = 90.0%) and was negative in 59 of the 60 negatives (specificity = 98.3%) in the preserved stools according to both readings. A marked difference was observed in the optical densities in both groups, preserved and unpreserved stools, when the G. lamblia-positive specimens were compared to the negative or positive for other intestinal parasites than G. lamblia. The assay seems a good alternative for giardiasis diagnosis, especially when the fecal examination was repeatedly negative and the patient presents giardiasislike symptoms.

Geranins A and B, new antiprotozoal A-type proanthocyanidins from Geranium niveum.

Bioassay-guided fractionation of the antiprotozoal extract of Geranium niveum led to the isolation of two new A-type proanthocyanidins, epi-afzelechin-(4beta-->8, 2beta-->O-->7)-afzelechin (1) and epi-catechin-(4beta-->8, 2beta-->O-->7)-afzelechin (2). Compounds 1 and 2 were given the trivial names of geranins A and B, respectively. In addition, five known compounds, mahuannin B (3), reynoutrin (4), hyperin (5), methyl gallate (6), and 3-beta-caffeoyl-12-oleanen-28-oic acid (7), were obtained. The structures of the new proanthocyanidins were elucidated by spectroscopic and chemical methods as well as CD measurements. Compounds 1, 2, and 4-7 were tested against axenically grown trophozoites of Giardia lamblia and Entamoeba histolytica.

Role of protozoa as risk factors for persistent diarrhea.

A case control study including 175 children aged 0-36 months suffering from diarrhea of > or = 14 days duration was undertaken to determine whether there is an association between Giardia lamblia, Entamoeba histolytica or Cryptosporidium infection and persistent diarrhea (PD). Subjects were identified by ongoing household surveillance and enrolled as cases. For each case two controls were selected by survey of neighbouring households--a child with acute diarrhea and one without diarrhea. Both the controls were matched with the case for age and nutritional status. Two fresh stool samples were collected from all cases and controls at enrollment and examined for trophozoites of Giardia lamblia, Entamoeba histolytica and Cryptosporidium. Giardia lamblia trophozoites were detected in a significantly higher proportion of PD cases (20.0%) than acute diarrheal and non diarrheal controls (4.6% each, p < 0.0001). There were no significant differences in the proportion of cases and controls who passed E. histolytica trophozoites or cryptosporidium in their stools. There was a consistent trend towards poorer weight gain in PD cases who passed Giardia trophozoites in stool; the differences were statistically significant at days 14 and 21, after enrollment. Giardia lamblia infection is more prevalent in PD cases than in acute diarrhea or non-diarrheal controls. This prevalence is not high enough to warrant routine anti-giardia therapy in patients with PD. However, as giardiasis was observed to have adverse growth impact in PD cases, stool microscopy for detection and subsequent treatment of Giardia lamblia seems to be justified.

Ethanol Effects on Intestinal Functions in Giardia lamblia-Infected Rats.

The effect of Giardia lamblia infection on D-glucose uptake and brush-border enzymes was studied in chronically ethanol-fed rats. Giardia lamblia trophozoites counts were significantly lower in the intestine of ethanol-fed rats than in the controls. Also, the Na+-dependent uptake of D-glucose and brush-border enzymes was significantly reduced (p<0.0001, p<0.01, respectively) in the G. lamblia-infected rat intestine. There was no change in Na+-independent glucose uptake under these conditions. Levels of mRNA encoding Na+-dependent D-glucose transporter (SGLT1) and brush-border sucrase and lactase were reduced in G. lamblia-infected animals compared with those of the controls. However, the mRNA levels encoding these proteins in ethanol-fed and control animals were similar in rat intestine. These findings suggest that down-regulation of SGLT1 and disaccharidases could be responsible for the observed D-glucose malabsorption and decrease in the activity of disaccharidases in G. lamblia infection.

Symptoms in patients with giardiasis undergoing upper gastrointestinal endoscopy.

The aim of the study was to compare symptoms of Giardia-positive and -negative patients undergoing upper gastrointestinal endoscopy. Multi-center questionnaire-based case-control study of 120 Giardia-positive and -negative patients who underwent upper endoscopy. Cases were identified through histological detection of trophozoites of Giardia lamblia in duodenal biopsy specimens. Giardia-negative patients with inconspicuous duodenal histology served as controls. There was a clear male predominance in the Giardia-positive groups, with 75 cases (62.5 %) being male and 45 (37.5 %) female (P < 0.0001). Giardia-positive cases tended to be older than controls (median 60 years vs. 45.2 years, P < 0.009). The main symptom of positive cases were tenderness to palpation (52.1%), abdominal discomfort (43.3 %), sensation of fullness (40.8%), distension (37.5%), epigastralgia (37.5%), nausea and/or vomiting (35.8%), heartburn (26.7%), and diarrhea (25.8%). Only sensation of fullness (P < 0.02), nausea/vomiting (P < 0.02) and heartburn (P < 0.03) were observed significantly more often in positive cases. Foul-smelling stools, considered typical for giardiasis, were observed in 5.8% of cases. Symptoms lasted for less than 1 month in 28.2 % of cases, for 1-6 months in 35.9%, and for more than 12 months in 35.9% (ns to controls). Treatment resulted in symptomatic cure in 60.5 % and in improvement of symptoms in 21.1 % of cases (P < 0.0001 vs controls). There are no symptoms that reliably allow the recognition of giardiasis in patients undergoing upper endoscopy.

Giardia lamblia:Evidence for Carrier-Mediated Uptake and Release of Conjugated Bile Acids

Giardia lambliatrophozoites colonize the human small intestine, where they are exposed to high concentrations of conjugated bile acids. Previous work has shown that bile acids enhance trophozoite survival, multiplication, and differentiation into the cyst stage. Therefore, experiments were performed to test whether carrier-mediated uptake of conjugated bile acids is present in this primitive parasite. Uptake of both cholyltaurine (C-tau) and cholylglycine (C-gly) was increased manyfold after culturing trophozoites in medium lacking bile acids. Absence of uptake at 4°C and inhibition by other conjugated bile acids provided additional evidence for carrier-mediated uptake. Uptake of C-tau was greater than that of C-gly under all experimental conditions and appeared to be mediated by a different carrier. The major evidence for different carriers is that C-tau uptake was Na+-dependent, while C-gly uptake was not. In addition, C-tau uptake was more strongly inhibited by DTNB and several organic anions than C-gly uptake. Radiolabeled C-tau and C-gly were each released rapidly from trophozoites at 37°C but not at 4°C, suggesting that release of conjugated bile acids was also carrier-mediated. These findings are consistent with the notion that multiple transporters for conjugated bile acids are present in a lower eukaryote. We speculate that intracellular bile acids may facilitate lipid trafficking and membrane biosynthesis.

Mechanisms of Giardia lamblia differentiation into cysts.

Microbiologists have long been intrigued by the ability of parasitic organisms to adapt to changes in the environment. Since most parasites occupy several niches during their journey between vectors and hosts, they have developed adaptive responses which allow them to survive under adverse conditions. Therefore, the life cycles of protozoan and helminthic parasites are excellent models with which to study numerous mechanisms involved in cell differentiation, such as the regulation of gene expression, signal transduction pathways, and organelle biogenesis. Unfortunately, many of these studies are very difficult because the conditions needed to elicit developmental changes in parasites remain undetermined in most cases. Recently, several interesting findings were reported on the process of differentiation of Giardia lamblia trophozoites into cysts. G. lamblia is a flagellated protozoan that inhabits the upper small intestine of its vertebrate host and is a major cause of enteric disease worldwide. It belongs to the earliest identified lineage among eukaryotes and therefore offers a unique insight into the progression from primitive to more complex eukaryotic cells. The discovery of a specific stimulus that induces trophozoites to differentiate into cysts, the identification and characterization of encystation-specific molecules, the elucidation of novel biochemical pathways, and the development of useful reagents and techniques have made this parasite an excellent model with which to study differentiation in eukaryotic cells. In this review, we summarize the most recent fundings on several aspects of Giardia differentiation and discuss the significance of these findings within the context of current knowledge in the field.

Biochemical and immunological characterization of soluble antigens of Giardia lamblia.

The crude soluble antigens (CSA) of Giardia lamblia trophozoites and their analytically purified fractions were characterized biochemically and immunologically to determine the most immunogenic fraction and its localization on the parasite. Both Sephacryl S-300 column chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed the highly complex and heterogeneous nature of CSA. Gel filtration of CSA showed four fractions (FI-FIV) with molecular masses of 250, 150, 110, and 10 kDa for fractions I-IV, respectively. Protein profiles of CSA demonstrated 28 Coomassie-blue bands in the range of 200-14 kDa. Similar banding patterns with fewer polypeptides were observed in the FI fraction. However, fractions II and III showed polypeptide bands in the region of 97-14 kDa. The glycoprotein nature of CSA and its-fractions were demonstrated in physicochemical analysis. In antigenic activity analysis the high-molecular-weight FI antigen was found to be 8 times more immunogenic than CSA as well as the other fractions. Major differences in the immunoreactivity of CSA and FI antigens were noted at 220, 30, and 22 kDa for the FI antigen and at 205, 84, 55, 43, and 20 kDa for CSA. Some of these FI polypeptides were found to be surface-associated as revealed by immunofluorescence and immunoblot assay. These results suggest the future use of the FI antigen in the serodiagnosis of and immunoprophylaxis against giardiasis.

Uptake and Cellular Localization of Exogenous Lipids byGiardia lamblia,a Primitive Eukaryote

Giardia lambliatrophozoites are unable to carry outde novolipid synthesis. It is therefore likely that lipids are acquired from the small intestine of the host, in which the trophozoites are exposed to free and conjugated fatty acids, various sterols, phospholipids, bile acids, and bile–lipid mixed micelles. Here we show thatG. lambliais capable of taking up exogenous phosphatidylcholine (PC), phosphatidylinositol (PI), sphingomyelin (SM), cholesterol, ceramide (Cer), and fatty acids. Results from epifluorescence and high-resolution confocal microscopy suggest that fluorescent analogs of SM and PC were accumulated in the plasma membranes, whereas palmitic acid and Cer were localized intracellularly. Interestingly, many of these analogs were also concentrated in perinuclear regions. Similar labeling patterns were observed when the fluorescent analogs were delivered to the parasite via liposomes. To test whetherG. lambliawas capable of esterifying exogenous fatty acids into membrane or cellular phospholipids, trophozoites were pulse-labeled with3H-labeled palmitic or myristic acids and the phospholipids analyzed by thin-layer chromatography. Results document thatG. lambliawas able to incorporate exogenous fatty acids into various phospholipids, i.e., PI, PC, PE, and PG. Interestingly, a major portion of radiolabeled fatty acids was incorporated into PG, a phospholipid characteristic of prokaryotic membranes.

Role of nitric oxide and superoxide in Giardia lamblia killing.

Giardia lamblia trophozoites were incubated for 2 h with activated murine macrophages, nitric oxide (NO) donors or a superoxide anion generator (20 mU/ml xanthine oxidase plus 1 mM xanthine). Activated macrophages were cytotoxic to Giardia trophozoites (approximately 60% dead trophozoites). The effect was inhibited (> 90%) by an NO synthase inhibitor (200 microM) and unaffected by superoxide dismutase (SOD, 300 U/ml). Giardia trophozoites were killed by the NO donors, S-nitroso-acetyl-penicillamine (SNAP) and sodium nitroprusside (SNP) in a dose-dependent manner (LD50 300 and 50 microM, respectively). A dual NO-superoxide anion donor, 3-morpholino-sydnonimine hydrochloride (SIN-1), did not have a killing effect in concentrations up to 1 mM. However, when SOD (300 U/ml) was added simultaneously with SIN-1 to Giardia, a significant trophozoite-killing effect was observed (approximately 35% dead trophozoites at 1 mM). The mixtures of SNAP or SNP with superoxide anion, which yields peroxynitrite, abolished the trophozoite killing induced by NO donors. Authentic peroxynitrite only killed trophozoites at very high concentrations (3 mM). These results indicate that NO accounts for Giardia trophozoites killing and this effect is not mediated by peroxynitrite.

Mechanisms of Giardia lamblia differentiation into cysts.

Microbiologists have long been intrigued by the ability of parasitic organisms to adapt to changes in the environment. Since most parasites occupy several niches during their journey between vectors and hosts, they have developed adaptive responses which allow them to survive under adverse conditions. Therefore, the life cycles of protozoan and helminthic parasites are excellent models with which to study numerous mechanisms involved in cell differentiation, such as the regulation of gene expression, signal transduction pathways, and organelle biogenesis. Unfortunately, many of these studies are very difficult because the conditions needed to elicit developmental changes in parasites remain undetermined in most cases. Recently, several interesting findings were reported on the process of differentiation of Giardia lamblia trophozoites into cysts. G. lamblia is a flagellated protozoan that inhabits the upper small intestine of its vertebrate host and is a major cause of enteric disease worldwide. It belongs to the earliest identified lineage among eukaryotes and therefore offers a unique insight into the progression from primitive to more complex eukaryotic cells. The discovery of a specific stimulus that induces trophozoites to differentiate into cysts, the identification and characterization of encystation-specific molecules, the elucidation of novel biochemical pathways, and the development of useful reagents and techniques have made this parasite an excellent model with which to study differentiation in eukaryotic cells. In this review, we summarize the most recent fundings on several aspects of Giardia differentiation and discuss the significance of these findings within the context of current knowledge in the field.