We have examined by gelatin-SDS-PAGE the protease activity in cell lysates of Giardia duodenalis trophozoites of two axenic strains isolated in Brazil from a symptomatic patient and an asymptomatic carrier , and the reference strain Portland 1 (P1). The proteolysis band patterns showed differences among strains isolated from asymptomatic and symptomatic individuals. The lysate of the strain BTU-10, showed only five hydrolysis bands, while a greater number of bands (10-11 bands) Giardia duodenalis (syn. G. lamblia, G. intestinalis) is a flagellated protozoan found in the small intestine of man and other mammals. Giardiasis is currently considered to be an important public health problem in developing countries, where it is one of the most common causes of diarrhea among children.Recent advances in the understanding of the biochemistry and molecular biology of protozoa have permitted the study of molecules that directly participate in the parasites life cycle or in the pathogenicity (North et al. 1990). Many important parasite species such as Entamoeba histolytica, Trypanosoma cruzi, Leshmania spp., Trichomonas vaginalis and Plasmodium spp. are actively proteolytic and contain multiple proteases that play a role in the life cycle, morphogenesis and infectivity (McKerrow et al. 1993). With respect to Giardia, despite evidence that proteolysis is involved in its nutrition, development and pathogenicity, relatively little is known. Initially, the intracelular activity of proteases in Giardia trophozoites was attributed to two major proteases corresponding to 105 and 40 kDa bands detected by SDS-PAGE using gelatin as a substrate (Hare et al. 1989, Parenti 1991 on the protease activity in trophozoites of other axenic strains. Moreover, there is increasing interest to conduct studies in attempting to learn more about strains isolated and axenized in endemic areas, in order to know the biological, clinical and epidemiological inter-strain differences in these areas where the search for control measures is still frequent.In the present investigation, we analyzed the protease activities in lysates of G. duodenalis trophozoites of two strains from a symptomatic (BTU-11) and an asymptomatic patient (BTU-10) isolated and axenized in Brazil, and the reference strain P1. The objectives were to analyse the protease activity in trophozoites and to determine if there are differences in such activity among the strains. MATERIALS AND METHODSParasites and culture -Trophozoites of three axenic strains of G. duodenalis were studied. Two of them were isolated in Brazil, at the Giardiasis Laboratory (IB/Unesp) in Botucatu, São Paulo: BTU-10 from a 21 year-old asymptomatic male and BTU-11 from a 40 year-old symptomatic male presenting diarrhea, flatulence and abdominal cramps. Both strains were obtained from cysts in the patient's feces. The third strain P1, originally isolated by Meyer (1976) from a 36 year-old symptomatic female from Portland, Oregon (USA), was obtained from HW Pillai (Division of Geographic Medicine Tufts University...

RSPU-B/218OLIVEIRA, M. R. et al.

SUMMARYIn the present study, we have analyzed by sodium docecyl sulphate -polyacrilamide gel electrophoresis (SDS-PAGE), immunoblotting and Concanavalin A blotting (Con A blotting) proteins of membrane fractions and soluble fractions obtained from Giardia duodenalis trophozoites of two axenic strains isolated in Brazil from a symptomatic (BTU-11) and an asymptomatic patient (BTU-10), as compared to the reference strain Portland 1. Both Brazilian strains showed a complex and homogeneous electrophoretic pattern of proteins, but some differences could be observed. Several glycoproteins were detected, particularly the proteins of 81, 72, 59 kDa and the protein of 62 kDa in the membrane proteins and cytosol, respectively. Many antigenic components were revealed by anti-Giardia rabbit IgG antibodies in the immunoblotting analysis. Among these components, the membrane protein of 32 kDa and the cytosol protein of 30 kDa could be related to giardin, as previously demonstrated.

Summary Trophozoites of Giardia canis obtained from scrapings of the intestinal mucosa of naturally infected dogs were prepared by Karapetyan's technique. A high concentration of trophozoites with good preservation was observed in the electron microscope. The striated disc is formed of concentrically arranged microtubules, surrounded by a small ventro‐lateral flange, supported by a marginal plate. Vacuoles are mainly present in the dorsal region. Invagination of the plasma membrane in association with the vacuoles suggests the formation of the vacuoles by pinocytosis. The cytoplasm contains a discrete rough endoplasmic reticulum and many ribosomes, but no glycogen particles. Cytoplasmatic cisterns, sometimes containing electron‐dense material, are seen. These cisterns are connected to the plasma membrane. The nuclei sometimes contain crystalloid inclusions. Close to the flagella, there is a microtubular structure and osmiophilic material. The median body is composed of rows of curved microtubules without any apparent relationship with other cytoplasmic structures. Zusammenfassung Ultrastruktur von Giardia canis‐Trophozoiten Trophozoiten von Giardia canis aus Geschabseln der Darmschleimhaut natürlich infizierter Hunde wurden nach der Technik von Karapetyan aufbereitet. Im Elektronenmikroskop wurde eine große Anzahl gut erhaltener Trophozoiten nachgewiesen. Die gestreifte Scheibe besteht aus konzentrisch angeordneten Mikrotubuli, umgeben von einem ventro‐lateralem Randstück und unterstützt durch eine Randplatte. Vakuolen finden sich hauptsächlich in der dorsalen Region. Invaginationen der Plasmamembran in Verbindung mit den Vakuolen lassen die Bildung der Vakuolen durch Pinozytosis vermuten. Das Zytoplasma enthält ein diskretes rauhes Endoplasmatisches Retikulum und viele Ribosomen aber keine Glykogenpartikel. Zytoplasmatische Zisternen, die zum Teil elektronendichtes Material enthalten, sind mit der Plasmamembran verbunden. Die Kerne enthalten zum Teil kristalloide Einschlüsse. In der Näher der Flagellen finden sich mikrotubuläre Strukturen und osmiophiles Material. Der Mediankörper besteht aus Reihen gekrümmter Mikrotubuli ohne eine auffallende Beziehung zu anderen zytoplasmatischen Strukturen. Résumé Ultrastructure des trophozoïtes de Giardia canis Des trophozoïtes de Giardia canis ont été préparés selon la technique de Karapetyan à partir de râclages de la muqueuse intestinale de chiens infectés. Un grand nombre de trophozoïtes bien conservés a été mis en évidence au microscope électronique. Le disque rayé se compose de microtubules rangés de façon concentrique, entouré d'une bordure ventro‐latérale et soutenu par une plaque de bordure. Des vacuoles se situaient principalement dans la région dorsale. Des invaginations de la membrane du plasma en liaison avec les vacuoles ont laissé supposer la formation des vacuoles par pinocytose. Le cytoplasme contient un réseau grossier discret endoplasmique et beaucoup de robosomes mais pas de particule de glycogène. Les cisternes cytoplasmiques contenant un matériel en par...