Morphology of Giardia agilis: observation by scanning electron microscopy and interference reflexion microscopy

The flagellated protozoan, Giardia agilis, was isolated from tadpole small intestine and examined by scanning electron microscopy and interference reflexion microscopy. The general morphology of the G. agilis trophozoite is similar to G. muris and G. duodenalis, but with modifications that reflect its elongated form. Interference reflexion microscopic analysis of attachment of G. agilis reveals a pattern of focal contacts by the lateral crest of the ventral disc, the ventrolateral flange, the lateral shield, and by numerous microvillus-like appendages found along the lateral border of the trophozoite. The pattern of focal contacts was observed to be dynamic; trophozoites were observed to make and break the focal contacts in a relatively short time and to glide along the surface of the substratum without breaking focal contacts.     

Morphology of Giardia agilis: Observation by Scanning Electron Microscopy and Interference Reflexion Microscopy1

The flagellated protozoan, Giardia agilis, was isolated from tadpole small intestine and examined by scanning electron microscopy and interference reflexion microscopy. The general morphology of the G. agilis trophozoite is similar to G. muris and G. duodenalis, but with modifications that reflect its elongated form. Interference reflexion microscopic analysis of attachment of G. agilis reveals a pattern of focal contacts by the lateral crest of the ventral disc, the ventrolateral flange, the lateral shield, and by numerous microvillus-like appendages found along the lateral border of the trophozoite. The pattern of focal contacts was observed to be dynamic; trophozoites were observed to make and break the focal contacts in a relatively short time and to glide along the surface of the substratum without breaking focal contacts.     

Giardia lamblia: evaluation of the in vitro effects of nocodazole and colchicine on trophozoites

Giardia lamblia is the most commonly detected parasite in the intestinal tract of humans and other mammals causing giardiasis. Giardia presents several cytoskeletal structures with microtubules as major components such as the ventral adhesive disk, eight flagella axonemes, the median body and funis. Many drugs have already been tested as antigiardial agents, such as albendazole and mebendazole, which act by specifically inhibiting tubulin polymerization and hence microtubule assembly. In the present work, we used the microtubule inhibitors nocodazole and colchicine in order to investigate their direct and indirect effects on Giardia ultrastructure and attachment to the glass surface, respectively. Axenically grown G. lamblia trophozoites were treated with nocodazole or colchicine for different time intervals and analyzed by light and electron microscopy. It was observed that trophozoites became completely misshapen, detached from the glass surface and failed to complete cell division. The main alterations observed included disc fragmentation, presence of large vacuoles, and appearance of electrondense deposits made of tubulin. The cytokinesis was blocked, but not the karyokinesis, and membrane blebs were observed. These findings show that Giardia behavior and cytoskeleton are clearly affected by the commonly used microtubule targetting agents colchicine and nozodazole.     

Recent insights into the mucosal reactions associated with Giardia lamblia infections

Giardia lamblia is an intestinal protozoan parasite infecting humans and various other mammalian hosts. The most important clinical signs of giardiasis are diarrhoea and malabsorption. Giardia lamblia is able to undergo continuous antigenic variation of its major surface antigen, named VSP (variant surface protein). While intestinal antibodies, and more specifically anti-VSP IgA antibodies, were proven to be involved in modulating antigenic variation of the parasite the participation of the local antibody response in control of the parasite infection is still controversial. Conversely, previous studies based on experimental infections in mice showed that cellular immune mechanisms are essential for elimination of the parasite from its intestinal habitat. Furthermore, recent data indicated that inflammatory mast cells have a potential to directly, or indirectly, interfere in duodenal growth of G. lamblia trophozoites. However, this finding was challenged by other reports, which did not find a correlation between intestinal inflammation and resistance to infection. Since intestinal infiltration of inflammatory cells and/or CD8+T-cells were demonstrated to coincide with villus-shortening and crypt hyperplasia immunological reactions were considered to be a potential factor of pathogenesis in giardiasis. The contribution of physiological factors to pathogenesis was essentially assessed in vitro by co-cultivation of G. lamblia trophozoites with epithelial cell lines. By using this in vitro model, molecular (through surface lectins) and mechanical (through ventral disk) adhesion of trophozoites to the epithelium was shown to be crucial for increased epithelial permeability. This phenomenon as well as other Giardia-induced intestinal abnormalities such as loss of intestinal brush border surface area, villus flattening, inhibition of disaccharidase activities, and eventually also overgrowth of the enteric bacterial flora seem to be involved in the pathophysiology of giardiasis. However, it remains to be elucidated whether at least part of these pathological effects are causatively linked to the clinical manifestation of the disease.     

Giardia flagellar motility is not directly required to maintain attachment to surfaces

Giardia trophozoites attach to the intestinal microvilli (or inert surfaces) using an undefined "suction-based" mechanism, and remain attached during cell division to avoid peristalsis. Flagellar motility is a key factor in Giardia's pathogenesis and colonization of the host small intestine. Specifically, the beating of the ventral flagella, one of four pairs of motile flagella, has been proposed to generate a hydrodynamic force that results in suction-based attachment via the adjacent ventral disc. We aimed to test this prevailing "hydrodynamic model" of attachment mediated by flagellar motility. We defined four distinct stages of attachment by assessing surface contacts of the trophozoite with the substrate during attachment using TIRF microscopy (TIRFM). The lateral crest of the ventral disc forms a continuous perimeter seal with the substrate, a cytological indication that trophozoites are fully attached. Using trophozoites with two types of molecularly engineered defects in flagellar beating, we determined that neither ventral flagellar beating, nor any flagellar beating, is necessary for the maintenance of attachment. Following a morpholino-based knockdown of PF16, a central pair protein, both the beating and morphology of flagella were defective, but trophozoites could still initiate proper surface contacts as seen using TIRFM and could maintain attachment in several biophysical assays. Trophozoites with impaired motility were able to attach as well as motile cells. We also generated a strain with defects in the ventral flagellar waveform by overexpressing a dominant negative form of alpha2-annexin::GFP (D122A, D275A). This dominant negative alpha2-annexin strain could initiate attachment and had only a slight decrease in the ability to withstand normal and shear forces. The time needed for attachment did increase in trophozoites with overall defective flagellar beating, however. Thus while not directly required for attachment, flagellar motility is important for positioning and orienting trophozoites prior to attachment. Drugs affecting flagellar motility may result in lower levels of attachment by indirectly limiting the number of parasites that can position the ventral disc properly against a surface and against peristaltic flow.     

Giardia lamblia: the effects of extracts and fractions from Mentha x piperita Lin. (Lamiaceae) on trophozoites

Giardia lamblia is a parasite that causes giardiasis in humans and other mammals. The common treatment includes different classes of drugs, which were described to produce unpleasant side effects. Mentha x piperita, popularly known as peppermint, is a plant that is frequently used in the popular medicine to treat gastrointestinal symptoms. We examined the effects of crude extracts and fractions from peppermint against G. lamblia (ATCC 30888) on the basis of trophozoite growth, morphology and adherence studies. The methanolic, dichloromethane and hexanic extracts presented IC(50) values of 0.8, 2.5 and 9.0microg/ml after 48h of incubation, respectively. The aqueous extract showed no effect against the trophozoites with an IC(50)>100microg/ml. The aqueous fraction presented a moderate activity with an IC(50) of 45.5microg/ml. The dichloromethane fraction showed the best antigiardial activity, with an IC(50) of 0.75microg/ml after 48h of incubation. The morphological and adhesion assays showed that this fraction caused several alterations on plasma membrane surface of the parasite and inhibited the adhesion of G. lamblia trophozoites. Cytotoxic assays showed that Mentha x piperita presented no toxic effects on the intestinal cell line IEC-6. Our results demonstrated antigiardial activity of Mentha x piperita, indicating its potential value as therapeutic agent against G. lamblia infections.     

Interaction between chondroitin-6-sulfate and Entamoeba histolytica as revealed by force spectroscopy

We have observed by atomic force microscopy (AFM) the amoeba surface and probed the interaction force between Entamoeba histolytica and chondroitin-6-sulphate (C6S). We have used several substrates to adhere trophozoites. The best reproducibility in sample preparation was obtained with fibronectin-coated coverslips and when the cells were fixed with paraformaldehyde. The images obtained with the AFM showed that the trophozoite exhibits an irregular surface. Pseudopods and waving adhesion plaques could be observed. Force spectroscopy analysis showed that the trophozoite surface strongly interacts with C6S-functionalized tips. During cantilever retraction, attractive force peaks were observed at distances up to 1.3 microm above the trophozoite surface. Statistical analysis of the force distributions collected for five samples shown a reproducible 2.2 nN mean adhesion force. We observed a reduction of the adhesion force and of the interaction distance after addition of galactose to the buffer solution suggesting that the observed interaction is also Gal/GalNAc-lectin-mediated.     

SEM evidence for a new species, Giardia psittaci

The genus Giardia has been subdivided by Filice (1952) into 3 species, G. agilis, G. muris, and G. duodenalis, based on the morphology of the median body and subtle variations in the dimensions of trophozoites. Giardia trophozoites were isolated from the small intestine of budgerigars (parakeets) and examined morphologically with light and scanning electron microscopy. These trophozoites, like other Giardia spp., possessed a flattened dorso-ventral shape, 8 flagella, and an adhesive disc on the ventral surface. The presence of a claw hammer-shaped median body suggested classification of these trophozoites as G. duodenalis. However, unlike any known members of G. duodenalis, the Giardia trophozoites from budgerigars were morphologically distinct in that they lacked the ventrolateral flange and therefore did not have a marginal groove bordering the anterior and lateral border of the adhesive disc. This distinct morphology clearly indicated that trophozoites from budgerigars should be considered as a separate species, G. psittaci. Our evidence has demonstrated that median body shape cannot serve as a sole criterion for speciation of Giardia. In addition, if other avian species of Giardia also resemble G. psittaci, then this would suggest that evolutionary divergence has occurred in the genus Giardia.     

Inhibition of in vitro attachment of Giardia trophozoites by mucin

An enzyme-linked immunoabsorbent assay was developed to quantify the number of Giardia sp. trophozoites attached to a substratum. Trophozoites were allowed to attach for 5 or 10 min to untreated or modified substratum, or allowed to attach in the presence of cytoskeletal inhibitors. Microtubule inhibitors, cochicine and nocodazole, were ineffective in blocking attachment, although the actin-disrupting agent cytochalasin B produced significant inhibition of attachment. Three different mucins were associated with significant inhibition of Giardia trophozoite attachment, which was reversed by treating the mucin-coated wells with 0.1% poly-L-arginine. Examination of mucin-treated substrata by high-resolution field emission scanning electron microscopy showed the presence of thin linear fibrils, whereas treatment of mucin-coated wells with poly-L-arginine revealed similar fibrils but of thicker dimensions. These data support the concept that mucin may inhibit Giardia sp. trophozoite attachment in vitro by electrostatic repulsion and that this can be eliminated by treatment of mucin-coated surfaces with polycationic agents, such as poly-L-arginine or poly-L-lysine.     

A Detailed, Hierarchical Study of Giardia lamblia's Ventral Disc Reveals Novel Microtubule-Associated Protein Complexes

Giardia lamblia is a flagellated, unicellular parasite of mammals infecting over one billion people worldwide. Giardia''s two-stage life cycle includes a motile trophozoite stage that colonizes the host small intestine and an infectious cyst form that can persist in the environment. Similar to many eukaryotic cells, Giardia contains several complex microtubule arrays that are involved in motility, chromosome segregation, organelle transport, maintenance of cell shape and transformation between the two life cycle stages. Giardia trophozoites also possess a unique spiral microtubule array, the ventral disc, made of approximately 50 parallel microtubules and associated microribbons, as well as a variety of associated proteins. The ventral disc maintains trophozoite attachment to the host intestinal epithelium. With the help of a combined SEM/microtome based slice and view method called 3View® (Gatan Inc., Pleasanton, CA), we present an entire trophozoite cell reconstruction and describe the arrangement of the major cytoskeletal elements. To aid in future analyses of disc-mediated attachment, we used electron-tomography of freeze-substituted, plastic-embedded trophozoites to explore the detailed architecture of ventral disc microtubules and their associated components. Lastly, we examined the disc microtubule array in three dimensions in unprecedented detail using cryo-electron tomography combined with internal sub-tomogram volume averaging of repetitive domains. We discovered details of protein complexes stabilizing microtubules by attachment to their inner and outer wall. A unique tri-laminar microribbon structure is attached vertically to the disc microtubules and is connected to neighboring microribbons via crossbridges. This work provides novel insight into the structure of the ventral disc microtubules, microribbons and associated proteins. Knowledge of the components comprising these structures and their three-dimensional organization is crucial toward understanding how attachment via the ventral disc occurs in vivo.     

Ultrastructure of Giardia duodenalis trophozoites group from hamster: some relevant aspects.

Trophozoites of Giardia duodenalis group obtained from fragments or scratched of hamster's mucosa were examined by transmission electron microscopy. The fine structure of the trophozoites are presented and compared with those described for other animals. Some of the trophozoites present the cytoplasm full of glycogen, rough endoplasmic reticulum-like structures and homogeneous inclusions not enclosed by membranes, recognized as lipid drops, which had not been observed in Giardia from other animals. The adhesive disk is composed of a layer of microtubules, from which fibrous ribbons extend into the cytoplasm; these ribbons are linked by layer of cross-bridge filaments that shows an intermediary dense band, described for the first time in this paper. The authors regard this band as the result of the cross-bridge filaments slinding in the medium region between adjacent fibrous ribbons, and suggest a contractile activity for them. The role of the adhesive disk on the trophozoite mechanism of attachment to host mucosa is also discussed.     

Giardia lamblia rearranges F-actin and alpha-actinin in human colonic and duodenal monolayers and reduces transepithelial electrical resistance

The mechanisms of epithelial injury in giardiasis remain unknown. The effects of live Giardia lamblia on cellular G-actin, F-actin, alpha-actinin, and electrical resistance of human intestinal epithelial monolayers were investigated using SCBN and Caco2 cell lines grown on chamber slides or Transwell filter membranes. In separate experiments, some monolayers were also exposed to sonicated trophozoites, some to supernatant from live G. lamblia cultures, and some with or without the Ca2+ channel blocker verapamil. After 2, 24, or 48 hr of coincubation with G. lamblia, monolayers were assessed for cytoskeletal arrangement under fluorescence and confocal laser microscopy, and transepithelial electrical resistance was measured. Exposure to live G. lamblia trophozoites induced localized condensation of F-actin and loss of perijunctional alpha-actinin while G-actin remained unchanged. Confocal laser microscopy indicated that F-actin rearrangement was not affected by verapamil and was localized within the terminal web area. Coincubation of monolayers with G. lamblia lysates or with spent medium alone similarly rearranged F-actin. Verapamil alone did not alter F-actin. Electrical resistance of SCBN and Caco2 monolayers exposed to G. lamblia was significantly decreased versus controls regardless of whether live or lysed trophozoite samples were used. The results indicate that G. lamblia-induced epithelial injury is associated with F-actin and alpha-actinin rearrangements in the terminal web area via mechanisms independent of extracellular Ca2+. These alterations are associated with reduced transepithelial electrical resistance and are due at least in part to trophozoite products.     

The Giardia ventrolateral flange is a lamellar membrane protrusion that supports attachment

Attachment to the intestinal epithelium is critical to the lifestyle of the ubiquitous parasite Giardia lamblia. The ventrolateral flange is a sheet-like membrane protrusion at the interface between parasites and attached surfaces. This structure has been implicated in attachment, but its role has been poorly defined. Here, we identified a novel actin associated protein with putative WH2-like actin binding domains we named Flangin. Flangin complexes with Giardia actin (GlActin) and is enriched in the ventrolateral flange making it a valuable marker for studying the flanges’ role in Giardia biology. Live imaging revealed that the flange grows to around 1 μm in width after cytokinesis, then remains uniform in size during interphase, grows in mitosis, and is resorbed during cytokinesis. A flangin truncation mutant stabilizes the flange and blocks cytokinesis, indicating that flange disassembly is necessary for rapid myosin-independent cytokinesis in Giardia. Rho family GTPases are important regulators of membrane protrusions and GlRac, the sole Rho family GTPase in Giardia, was localized to the flange. Knockdown of Flangin, GlActin, and GlRac result in flange formation defects. This indicates a conserved role for GlRac and GlActin in forming membrane protrusions, despite the absence of canonical actin binding proteins that link Rho GTPase signaling to lamellipodia formation. Flangin-depleted parasites had reduced surface contact and when challenged with fluid shear force in flow chambers they had a reduced ability to remain attached, confirming a role for the flange in attachment. This secondary attachment mechanism complements the microtubule based adhesive ventral disc, a feature that may be particularly important during mitosis when the parental ventral disc disassembles in preparation for cytokinesis. This work supports the emerging view that Giardia’s unconventional actin cytoskeleton has an important role in supporting parasite attachment.     

The biology of Giardia spp.

Gardia spp. are flagellated protozoans that parasitize the small intestines of mammals, birds, reptiles, and amphibians. The infectious cysts begin excysting in the acidic environment of the stomach and become trophozoites (the vegetative form). The trophozoites attach to the intestinal mucosa through the suction generated by a ventral disk and cause diarrhea and malabsorption by mechanisms that are not well understood. Giardia spp. have a number of unique features, including a predominantly anaerobic metabolism, complete dependence on salvage of exogenous nucleotides, a limited ability to synthesize and degrade carbohydrates and lipids, and two nuclei that are equal by all criteria that have been tested. The small size and unique sequence of G. lamblia rRNA molecules have led to the proposal that Giardia is the most primitive eukaryotic organism. Three Giardia spp. have been identified by light lamblia, G. muris, and G. agilis, but electron microscopy has allowed further species to be described within the G. lamblia group, some of which have been substantiated by differences in the rDNA. Animal models and human infections have led to the conclusion that intestinal infection is controlled primarily through the humoral immune system (T-cell dependent in the mouse model). A major immunogenic cysteine-rich surface antigen is able to vary in vitro and in vivo in the course of an infection and may provide a means of evading the host immune response or perhaps a means of adapting to different intestinal environments.     

The biology of Giardia spp.

Gardia spp. are flagellated protozoans that parasitize the small intestines of mammals, birds, reptiles, and amphibians. The infectious cysts begin excysting in the acidic environment of the stomach and become trophozoites (the vegetative form). The trophozoites attach to the intestinal mucosa through the suction generated by a ventral disk and cause diarrhea and malabsorption by mechanisms that are not well understood. Giardia spp. have a number of unique features, including a predominantly anaerobic metabolism, complete dependence on salvage of exogenous nucleotides, a limited ability to synthesize and degrade carbohydrates and lipids, and two nuclei that are equal by all criteria that have been tested. The small size and unique sequence of G. lamblia rRNA molecules have led to the proposal that Giardia is the most primitive eukaryotic organism. Three Giardia spp. have been identified by light lamblia, G. muris, and G. agilis, but electron microscopy has allowed further species to be described within the G. lamblia group, some of which have been substantiated by differences in the rDNA. Animal models and human infections have led to the conclusion that intestinal infection is controlled primarily through the humoral immune system (T-cell dependent in the mouse model). A major immunogenic cysteine-rich surface antigen is able to vary in vitro and in vivo in the course of an infection and may provide a means of evading the host immune response or perhaps a means of adapting to different intestinal environments.     

Electron and video-light microscopy analysis of the in vitro effects of pyrantel pamoate on Giardia lamblia

Electron and video-light microscopy analysis of the in vitro effects of pyrantel pamoate on Giardia lamblia. Experimental Parasitology 97, 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.     

In vitro excystation of Giardia from humans: a scanning electron microscopy study

The in vitro excystation of Giardia lamblia on cysts isolated from human feces was studied. After purification by sucrose gradient, cysts were incubated in a pepsin-acid solution, then placed in a modified HSP3 medium where excystation occurred within a few minutes. The excystation procedure was studied by continuous observations by light microscopy and sequential observations by scanning electron microscopy (SEM). The in vitro excystation was stopped at timed intervals during incubation by addition of a large amount of 1% glutaraldehyde. The excystation process began by the cyst wall opening at one pole. Flagella protruded rapidly, the parasite emerged progressively from the cyst envelope, posterior end first, the empty cyst collapsed and shrank. Although flagella emerging from the organism were distinguishable, the cell body had not yet shown all the morphological features of the G. lamblia trophozoite. A radical rearrangement of the organism occurred gradually: initially oval in shape, the parasite became round, then elongated, flattened, and underwent cytokinesis. The daughter trophozoites acquired their typical morphological features: the shape, the adhesive disc with the C-shaped structure distinctly visible on the ventral surface, and the definite placement of the flagella. These observations obtained on G. lamblia by SEM were comparable to those obtained with G. muris.     

Structure of the organs of attachment of brachiolaria larvae of Stichaster australis (Verrill) and Coscinasterias calamaria (Gray) (Echinodermata: Asteroidea)

The structure of the brachiolar arms and adhesive disk of the brachiolaria larvae of Stichaster australis (Verrill) and Coscinasterias calamaria (Gray) was determined from light microscopy and from scanning and transmission electron microscopy. The structure of these organs was very similar in both species.The brachiolar arms are comprised of a stem region terminating in a crown of adhesive papillae which are made up of a variety of secretory cell types. Principal among these are elongated cells producing very electron-dense secretory particles, which are released at the free cell surface attached to cilia. Secretory particles appear to be important in temporary attachment of the brachiolar arms to the substratum. Ciliary sense cells, possibly used in the recognition of specific substrata are located at the tip of adhesive papillae.The adhesive disk is comprised of large cells packed with secretory droplets and elongated intracellular fibres. In the attached adhesive disk, secretory droplets are lost, having formed the cement that attaches the disk to the substratum. It appears that adhesive papillae lateral to the adhesive disk hold the disk in position close to the substratum during secretion and hardening of the cement. The intracellular fibres are the principal anchoring structures running from microvilli (locked into the attachment cement) on the surface of the disk to the underlying connective tissue of the attachment stalk.     

Giardia muris: ultrastructural analysis of in vitro excystation

Giardia muris cysts were examined by transmission electron microscopy before treatment, after induction, and at timed intervals during the incubation phase of in vitro excystation. Untreated G. muris cysts had a thick cyst wall composed of a fibrous outer wall and a thin, electron-dense inner membrane which extended from the trophozoite plasma membrane. The cytoplasm was devoid of endoplasmic reticulum, Golgi bodies,and mitochondria. Numerous large vacuoles were present within the ectoplasm just beneath the plasma membrane in untreated cysts. Following induction these cysts lacked ectoplasmic vacuoles. Concurrently, numerous membrane bound vesicles were seen in the peritrophic space closely adhering to the surface of the trophozoite. These vesicles appear to be of cytoplasmic origin. The cytoplasm of fully excysted trophozoites lacked ectoplasmic vacuoles but displayed well-developed ribbons of microtubular bodies, probably precursors of ventral disk, lateral flange, and median bodies and also contained extensive granular endoplasmic reticulum. No more than two nuclei were observed within each organism. The earliest excysted organisms were observed 0-5 min after incubation had begun and most organisms had excysted within 10 min. Cytokinesis occurred only after excystation was complete.     

Protein SUMOylation is Involved in Cell‐cycle Progression and Cell Morphology in Giardia lamblia

The unicellular protozoa Giardia lamblia is a food‐ and waterborne parasite that causes giardiasis. This illness is manifested as acute and self‐limited diarrhea and can evolve to long‐term complications. Successful establishment of infection by Giardia trophozoites requires adhesion to host cells and colonization of the small intestine, where parasites multiply by mitotic division. The tight binding of trophozoites to host cells occurs by means of the ventral adhesive disc, a spiral array of microtubules and associated proteins such as giardins. In this work we show that knock down of the Small Ubiquitin‐like MOdifier (SUMO) results in less adhesive trophzoites, decreased cell proliferation and deep morphological alterations, including at the ventral disc. Consistent with the reduced proliferation, SUMO knocked‐down trophozoites were arrested in G1 and in S phases of the cell cycle. Mass spectrometry analysis of anti‐SUMO immunoprecipitates was performed to identify SUMO substrates possibly involved in these events. Among the identified SUMOylation targets, α‐tubulin was further validated by Western blot and confirmed to be a SUMO target in Giardia trophozoites.