Chapters authored
Toxoplasma gondii Tissue Cyst: Cyst Wall Incorporation Activity and Matrix Cytoskeleton Proteins Paving the Way to Nutrient Acquisition
Toxoplasma gondii is an intracellular parasite that causes chronic infection by the development of bradyzoites housed in tissue cysts, preferably in the muscles and central nervous system. The composition and the function of the cyst wall are still not fully understood. Are T. gondii cysts able to incorporate nutrients through its wall? If so, how would these nutrients be traversed to cross the cyst matrix to reach the bradyzoite forms? Herein, we tested the uptake capacity of the Toxoplasma tissue cyst wall by employing some fluid-phase endocytosis tracers as peroxidase (HRP) and bovine serum albumin (BSA). Fluorescence images revealed these molecules on the cyst wall as well as in the cyst matrix. The subcellular localization of the tracer was confirmed by ultrastructural analysis showing numerous labeled vesicles and tubules distributed within the cyst matrix in close association with intracystic bradyzoite membrane, suggesting the cyst wall as a route of nutrient uptake. Furthermore, we confirmed the presence of cytoskeleton proteins, such as tubulin, actin, and myosin, in the tissue cyst matrix that may explain the nutrient input mechanism through the cyst wall. A better understanding of the nutrient acquisition process by the cyst might potentially contribute to the development of new therapeutic targets against chronic toxoplasmosis.
Part of the book: Toxoplasmosis
Congenital Toxoplasmosis: In Vivo Impact of Toxoplasma gondii Infection on Myogenesis and Neurogenesis
Congenital toxoplasmosis (TC) from Toxoplasma gondii positive mother to child transmission results in fetal death, abortion, or infantile neurologic and neurocognitive deficits as well as chorioretinitis. This study aims to analyze the morphological changes in brain and skeletal muscle cells of Swiss mouse embryos during experimental congenital toxoplasmosis. Swiss mice, before mating, were gavage inoculation infected with approximately 25 or 50 cysts of ME‐49 strain T. gondii. Eighteen day postcoitus maternal and embryonic muscle and brain samples were collected and processed for histopathological analysis. The muscle tissue from embryos of infected mothers, in comparison with healthy muscle myofibers, exhibited discontinuous and shorter myofibrils, more interfibrillar space and immature cells with fewer stained and poorly defined striated profiles. These in vivo findings might be related to an adhesion protein decrease, observed in vitro, where myogenesis was completely affected during Toxoplasma infection. The neurogenesis was severely affected with irregularly arranged cells, reduced cell density, and a significant intercellular space increase. The brain tissue presented ischemia, cell death, necrosis, and thrombi, increasing according to the degree of the acute infection, which compromised the neurogenesis, thereby justifying brain size decrease in these embryos.
Part of the book: Toxoplasmosis
Development of Schizont Stages of Toxoplasma gondii in Primary Cell Culture of Feline Enterocytes
Intestinal epithelial cell cultures are a potentially applicable model for investigating enteropathogens such as the protozoan Toxoplasma gondii, the etiological agent of toxoplasmosis. Felids such as domestic cats are the only known definitive hosts where the parasite undergoes sexual reproduction, which occurs in the enterocytes. Primary feline intestinal epithelial cell (FIEC) cultures were obtained from the fetal small gut of felines, and the epithelial nature of these cells was confirmed by the revelation of cytokeratin and intestinal alkaline phosphatase content by fluorescence microscopy, besides alignment, microvilli, and adherent intercellular junctions by ultrastructural analysis. FIECs infected with T. gondii bradyzoite forms showed that the parasite:cell ratio was determinant for establishing the lytic cycle and cystogenesis and the induction of schizont-like forms. Type C and D schizonts were identified by light and electron microscopies, which showed morphological characteristics like those previously described based on the analysis of cat intestines experimentally infected with T. gondii. These data indicate that FIECs simulate the microenvironment of the felid intestine, allowing the development of schizogony and classic endopolygeny. This cellular framework opens new perspectives for the in vitro investigation of biological and molecular aspects involved in the T. gondii enteric cycle.
Part of the book: Towards New Perspectives on Toxoplasma gondii
Nutrient Uptake Portals in Toxoplasma gondii Tachyzoites
The process of nutrient acquisition by Toxoplasma gondii tachyzoites is an attractive target for developing and designing drugs against toxoplasmosis, however, just recently it was revealed to be an important process to be understood. The present work helps address the lack of information about the exact sites where nutrient uptake in T. gondii. The endocytosis of proteins by tachyzoites of T. gondii was measured using both fluid-phase and receptor-mediated endocytic tracers. Quantitative analysis by flow cytometry revealed important differences in the percentage of labeled parasites, incubated with BSA, dextran, or transferrin. The analysis by confocal microscopy showed that the anterior portion of the conoid is one preferential site for binding BSA and transferrin to the tachyzoite, later localized within elongated structures present in the anterior region of the parasite. The ultrastructural analysis of multiple ultrathin sections displayed the endocytic markers at the following: (i) conoid, within rhoptries, (ii) in cup-shaped invagination of the parasite membrane (micropore) and, (iii) posterior pore. The present study brings data revealing three possible nutrient uptake portals in Toxoplasma tachyzoites that may contribute in the future to a therapeutic design with a view to treatment of toxoplasmosis.
Part of the book: Towards New Perspectives on Toxoplasma gondii