The first description of eggs in the male reproductive system of Physaloptera bispiculata (Nematoda: Spiruroidaea).

Physaloptera bispiculata (Nematoda: Spiruroidaea) is a parasite of Nectomys squamipes (Rodentia: Cricetidae), a water rat that only occurs in Brazil. Naturally infected rodents were captured in the municipality of Rio Bonito, Rio de Janeiro, Brazil. Adult P. bispiculata worms were collected, prepared and analysed by light and scanning electron microscopy. Under scanning electron microscopy, several eggs were seen glued by cement to the cloacal aperture. Light microscopy revealed that some male worms had an uncountable number of embryonated eggs in the ejaculatory duct, cloaca and also in the posterior portion of the intestine. The probable explanation is that the eggs developing in the female uterus are pumped by the female or sucked by the male to the cloacal opening and from this point to the intestine and ejaculatory duct. The male probably does not have the ability to expel the eggs and for this reason a large number were found in these organs. On the other hand, this could be an important adaptation for the parasite, i.e. male worms expelled by the host can carry a large number of eggs and spread them to intermediate hosts when ingested by these hosts. As far as we know this is the first record of a physalopterid nematode harbouring eggs in the cloacal region, ejaculatory duct or intestine.

Effects of a putrescine analog on Giardia lamblia.

The protozoan Giardia lamblia is the most frequent intestinal parasite of first-world countries and a major cause of waterborne disorder often referred to as traveler's diarrhea. We have previously noticed that the putrescine analog 1,4-diamino-2-butanone (DAB) remarkably inhibits the growth of anaerobic trichomonad and Trypanosoma cruzi parasites. Here, we examined the role of polyamines in Giardia cells using this putrescine analog. DAB impaired parasite proliferation dose-dependently. The analog induced increased flagella numbers and sometimes four ventral disks as well as asymmetrical division, indicating truncated or deregulated cytokinesis. Electron microscopy analysis revealed that DAB also triggered the encystment process. Oxidative stress was evaluated by measuring lipid peroxidation by thiobarbituric acid reactive substances (TBARS) detection. Trophozoites incubated either with 1 mM of DAB or putrescine for 18 h displayed increased lipoperoxide levels. Addition of 200 microM aminoguanidine, a polyamine/diamine oxidase inhibitor, partially reverted the DAB, but not the putrescine effects, indicating that the DAB effects are due, at least in part, to DAB oxidation end products. These data indicate that polyamines play a role in Giardia cell division, differentiation, and antioxidant defenses.

Fine structure of the biogenesis of Giardia lamblia encystation secretory vesicles.

Synthesis, transport, and assembly of the extracellular cyst wall is the hallmark of Giardia lamblia encystation. Much is known of the biochemical pathways and their regulation. However, from a cell biology point of view, the biogenesis of the encystation specific vesicles (ESVs) that transport cyst wall proteins to the periphery of the cell is poorly understood. Therefore, we exploited a number of complementary ultrastructural approaches to test the hypothesis that the formation of ESVs utilizes a novel regulated secretory pathway. We analyzed parasites at different stages of encystation in vitro by electron microscopy of thin sections, freeze fracture replicas, and three-dimensional reconstruction from serial sections of cells fixed for cytochemical localization of the endoplasmic reticulum (ER) marker, glucose 6-phosphatase. We also used a stereological approach to determine the area occupied by the ER, clefts, ESVs, and cyst wall. Taken together, our kinetic data suggest that some ER cisternae first dilate to form clefts, which enlarge into the ESVs. Living non-encysting and early-encysting trophozoites were labeled around the periphery of both nuclei with C(6)-NBD-ceramide. At 18-21 h, outward migration of some ESVs frequently caused protrusions at the periphery of encysting trophozoites. The presence of lysosome-like peripheral vesicles between the ESV and plasma membrane of the cell was confirmed using acridine orange, an acidic compartment marker. Our data suggest that G. lamblia has a novel secretory pathway in which certain functions of the ER and Golgi co-localize spatially and temporally. These studies will increase understanding of the evolutionary appearance of regulated secretory pathways for assembly of a primitive extracellular matrix in an early diverging eukaryote.

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.

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.

The peripheral vesicles of trophozoites of the primitive protozoan Giardia lamblia may correspond to early and late endosomes and to lysosomes.

Giardia lamblia, a primitive eukaryotic cell, lacks organelles such as mitochondria, peroxisomes, and a typical Golgi complex and presents a system of vesicles located below the plasma membrane. We used fluorescence and electron microscopy to better characterize the peripheral vesicles. Incubation of living cells with acridine orange showed that the peripheral vesicles correspond to an acidic compartment. Incubation with lucifer yellow, and with horseradish peroxidase, showed labeling of the peripheral vesicles even after several hours. Acid phosphatase was localized in the endoplasmic reticulum and in most of the peripheral vesicles. On the other hand, glucose 6-phosphatase, an endoplasmic reticulum marker, was observed in the endoplasmic reticulum cisternae and in some peripheral vesicles. A similar labeling pattern was observed using the zinc iodide technique, which reveals SH-containing proteins. Three-dimensional reconstruction and electron microscopy tomography of cells stained for acid phosphatase and glucose-6-phosphatase revealed the connection between some vesicles and profiles of the endoplasmic reticulum. Taken together, our observations suggest that trophozoites of G. lamblia present an endosomal-lysosomal system concentrated in a single system, the peripheral vesicles, which may represent an ancient organellar system that later on subdivided into compartments such as early and late endosomes and lysosomes.