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Mechanical integration of muscle, tegument, and subtegumental tissues by anchoring fibrils and microfibrils in the cestode Hymenolepis diminuta.

Electron microscopic examination of the epidermal basement membrane region of the rat tapeworm, Hymenolepis diminuta, has revealed specialized connective tissue structures that appear to anchor the epidermis, or tegument, to the parenchymal tissues of the helminth, as well as interconnect subtegumental muscle fibers, tegument, and parenchyma. Anchoring fibrils-cross-banded bundles of ca. 3 nm diameter filaments--were observed to directly interlink tegument and muscle, muscle and muscle, and tegument, muscle, and parenchymal connective tissue. Anchoring fibrils therefore appear to mechanically integrate epidermal tissue movements in response to subtegumental muscle contraction. A well-developed stratum of microfibrils, forming the lamina reticularis of the tegumental basement membrane, may also help anchor the tegument as well as to serve as a flexible, reinforcing sheath that protects parenchymal tissues from excessive radial displacement due to muscle contraction.

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Demonstration of specific high-affinity binding sites for [3H]5-hydroxytryptamine in the cestode Hymenolepis diminuta.

[3H]5-HT (0.16-8.32 nM) exhibited saturable and specific binding in membrane preparations of Hymenolepis diminuta. The saturation data produced a non-linear Scatchard plot which could be resolved into sites having apparent dissociation constants (Kd) of 0.17 and 8.30 nM for the high-affinity and low-affinity components, respectively. Drug displacement studies, using radioligand concentrations of 0.6 and 6 nM, revealed that the two [3H]5-HT binding components are pharmacologically distinct and do not conform to any known class of 5-HT recognition site. The physiological significance of these putative 5-HT receptors and their potential usefulness for the selection of new antiparasitic agents are discussed.

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Serotonin stimulates protein phosphorylation in the cestode Hymenolepis diminuta.

1. Serotonin stimulated the incorporation of 32P from [gamma-32P] ATP into crude membrane preparations (P2) of Hymenolepis diminuta in a dose-dependent manner (EC50 of approximately 0.79 microM). 2. This response was seen with several serotonin agonists, and was inhibited by several serotonin antagonists, which were identical to the previously described activation and inhibition of serotonin-sensitive adenylate cyclase. 3. Cyclic AMP produced a dose-dependent stimulation of 32P incorporation into the P2 fraction, with an EC50 of approximately 2.51 microM. 4. The targets for the serotonin stimulated incorporation of 32P were found to be in trypsin-labile proteins with Mr's of 134,000, 110,000, 82,000, 80,000 and 31,000.

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Effects of phytochemicals of Flemingia vestita (Fabaceae) on glucose 6-phosphate dehydrogenase and enzymes of gluconeogenesis in a cestode (Raillietina echinobothrida).

The crude root-peel extract of Flemingia vestita, containing genistein as the major isoflavone, has a vermifugal/vermicidal effect. It acts by causing flaccid paralysis accompanied by alterations in the activities of several tegumental enzymes and other metabolic activities in the fowl tapeworm, Raillietina echinobothrida. To elucidate the mode of action of the putative phytochemicals on energy metabolism, crude root-peel extract, pure genistein and praziquantel were tested on glucose 6-phosphate dehydrogenase (G6PDH) and enzymes of gluconeogenesis--pyruvate carboxylase (PC), phosphoenolpyruvate carboxykinase (PEPCK) and fructose 1,6-bisphosphatase (FBPase)--in R. echinobothrida. The activities of G6PDH, PEPCK and FBPase were largely restricted to the cytosolic fraction, while PC was confined to the mitochondrial fraction. Following treatments, the G6PDH activity was decreased by 23-31%, whereas the activities of PC and PEPCK were increased by 32-44% and 44-49%, respectively. There was no significant effect by any of the treatments on FBPase activity. We hypothesize that the phytochemicals from F. vestita, genistein in particular, influence the key enzymes of these pathways, which is perhaps a function of high energy demand of the parasite under anthelmintic stress.

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Regulation of the immune response to cestode infection by progesterone is due to its metabolism to estradiol.

The aim of this work was to investigate the role of progesterone during Taenia crassiceps cysticercosis, and the immunological mechanisms involved in its effects, by relating progesterone treatment to whole parasite counts, to host humoral and cellular immune response, to the presence or absence of nuclear receptors to sex steroids in splenocytes, and to serum sex steroid levels in infected mice of both genders. Progesterone treatment increased parasite loads two-fold in females and three-fold in males compared with control mice. The expression of the Th2 cytokine profile (IL-4, IL-6 and IL-10) was markedly increased in infected mice of both genders, while progesterone treatment returned this expression to basal levels. However, the Th1 cytokine profile (IFN-gamma and TNF-alpha) was not affected by infection, whilst progesterone treatment increased the expression of both cytokines two-fold compared to uninfected, infected and placebo-treated mice. Testosterone serum levels decreased in infected male mice by 95%, and treatment with progesterone did not affect them. In females, no change in testosterone levels was observed. Progesterone levels increased three-fold only in progesterone-treated infected mice of both sexes, while estradiol levels in female and male progesterone-treated infected mice increased two-fold compared to infected control mice. The infection markedly induced the expression of progesterone receptor (PR) isoforms A and B in splenocytes of infected mice of both genders (five-fold). Metabolism of progesterone to estradiol was demonstrated by the use of the anti-estrogen tamoxifen, which reduced parasite loads 100% in infected mice of both sexes treated with progesterone. These results suggest that progesterone, possibly through its metabolism to estradiol, affects establishment, growth and reproduction of the helminth parasite T. crassiceps.

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Molecular discrimination of taeniid cestodes.

DNA approaches are now being used routinely for accurate identification of Echinococcus and Taenia species, subspecies and strains, and in molecular epidemiological surveys of echinococcosis/taeniasis in different geographical settings and host assemblages. The publication of the complete sequences of the mitochondrial (mt) genomes of E. granulosus, E. multilocularis, T. solium and Asian Taenia, and the availability of mtDNA sequences for a number of other taeniid genotypes, has provided additional genetic information that can be used for more in depth phylogenetic and taxonomic studies of these parasites. This very rich sequence information has provided a solid molecular basis, along with a range of different biological, epidemiological, biochemical and other molecular-genetic criteria, for revising the taxonomy of the genus Echinococcus and for estimating the evolutionary time of divergence of the various taxa. Furthermore, the accumulating genetic data has allowed the development of PCR-based tests for unambiguous identification of Echinococcus eggs in the faeces of definitive hosts and in the environment. Molecular phylogenies derived from mtDNA sequence comparisons of geographically distributed samples of T. solium provide molecular evidence for two genotypes, one being restricted to Asia, with the other occurring in Africa and America. Whether the two genetic forms of T. solium differ in important phenotypic characteristics remains to be determined. As well, minor DNA sequence differences have been reported between isolates of T. saginata and Asian Taenia. There has been considerable discussion over a number of years regarding the taxonomic position of Asian Taenia and whether it should be regarded as a genotype, strain, subspecies or sister species of T. saginata. The available molecular genetic data do not support independent species status for Asian Taenia and T. saginata. What is in agreement is that both taxa are closely related to each other but distantly related to T. solium. This is important in public health terms as it predicts that cysticercosis in humans attributable to Asian Taenia does not occur, because cysticercosis is unknown in T. saginata.

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Spermatological characters of the pseudophyllidean cestode Bothriocephalus scorpii (Müller, 1776).

Spermiogenesis of Bothriocephalus scorpii (Cestoda, Pseudophyllidea) includes an orthogonal development of two flagella, followed by a flagellar rotation and a proximo-distal fusion with the median cytoplasmic process. The fusion occurs at the level of four attachment zones. The presence of dense material in the apical region of the differentiation zone in the early stage of spermiogenesis appears to be a characteristic feature for the Pseudophyllidea. The mature spermatozoon possesses two axonemes of 9+"1" pattern of the Trepaxonemata, nucleus, cortical microtubules, electron-dense granules and crested body. The anterior part of the gamete exhibits a centriole surrounded by electron-dense tubular structures arranged as incomplete spiral. When the crested body disappears, the electron-dense tubular structures are arranged into a ring encircling the axoneme. The electron-dense tubular structures and their arrangement appear to be a specific feature for the clade "Bothriocephalidea". The organization of the posterior extremity of the gamete with the nucleus is described for the first time in the Pseudophyllidea.

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Molecular and genetic characterisation of the host-protective oncosphere antigens of taeniid cestode parasites.

Highly effective recombinant vaccines have been developed against Taenia ovis infection in sheep, Taenia saginata infection in cattle, Taenia solium infection in pigs, Echinococcus granulosus and Echinococcus multilocularis infections in a variety of intermediate host species. These vaccines have been based on the identification and expression in Escherichia coli of antigens derived from the oncosphere life cycle stage, contained within the parasites' eggs. Investigation of the molecular aspects of these proteins and the genes encoding them have revealed a number of common features, including the presence of a predicted secretory signal sequence, and one or two copies of a fibronectin type III domain, each encoded by separate exons within the associated gene. Evidence has been obtained to confirm glycosylation of some of these antigens. Ongoing investigations will shed light on the biological roles played by the proteins within the parasites and the mechanism by which they make the parasites vulnerable to vaccine-induced immune responses.

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