[Localization of acetylcholinesterase in the nervous system of Cotylophoron indicum].

The nervous system of Cotylophoron indicum was studied by using acetylcholine esterase histochemical staining techniques. Cranial ganglia and transverse commissure situate at dorso-lateral body between oral sucker and genital sucker. From the cranial ganglia four pairs of nerves proceed cephalad and connect with nerve network of the oral sucker. The posterior nerve cords from the cranial ganglia consist of 3 pairs and the ventral ones are the stoutest and longest nerves. A few branches from the 3 pairs of nerve cords connect to ventral sucker. There is a developed nerve network distributed in its genital sucker. The nerve fibers on body surface in pairs and parallel are diagonal and cross to form a nerve network on body surface. Three kinds of neurocytes distribute at the prosomal region. Results show that the nervous system structure of C. indicum is consistent with the essential features of Digenea, but more special and complicated around genital sucker.

In vitro effect of some anthelmintics on lactate dehydrogenase activity of Cotylophoron cotylophorum (Digenea: paramphistomidae).

Effects of praziquantel (PZQ), levamisole (LEV), mebendazole (MBZ), fenbendazole (FBZ) and albendazole (ABZ) on the lactate dehydrogenase (LDH) activity of Cotylophoron cotylophorum were studied in vitro. Maximum levels of inhibition of LDH catalysing both oxidation and reduction reactions were observed in PZQ- and LEV-treated worms. Similarly, benzimidazoles - MBZ, FBZ and ABZ - have also significantly inhibited the activity of LDH catalysing the oxidation of lactate; whereas the activity of LDH catalysing the reduction of pyruvate was accelerated. This affects the mitochondrial energy generating process which ultimately proves fatal to the parasite. Therefore, the mode of action of benzimidazoles is primarily on the activation of LDH catalysing the conversion of pyruvate to lactate.

[3H]-amino acid uptake and metabolic studies on Gigantocotyle explanatum and Gastrothylax crumenifer (Digenea: Paramphistomidae).

The amphistomes Gigantocotyle explanatum and Gastrothylax crumenifer utilize leucine, alanine, proline and methionine during in vitro incubations. Autoradiography on sections of these flukes reveal a time-dependent differential incorporation of tritium-labelled amino acids in various tissues. The tegument appears to be the primary surface through which amino acids are absorbed. Following absorption, the reappearance of [3H]-leucine and [3H]-alanine on the tegumental surface during late chase periods indicates their possible involvement in tegumental secretion. A combination of diffusion and carrier-mediated uptake, possibly involving gamma-glutamyl transpeptidase, is indicated. The transport loci show differences in carrier-affinity (Kt) and maximum uptake velocities (Vmax) for amino acids under study, which suggest multiple transport molecules. Metabolic studies reveal that aspartate, alanine, ornithine, proline, leucine and methionine undergo transamination through 2-oxoglutarate-linked transaminases, distributed in the cytosolic and mitochondrial fractions of G. explanatum and G. crumenifer. With the exception of alanine transaminase, the enzyme levels in the cytosolic fraction were higher than the mitochondrial fraction of the two amphistomes. Predominantly cytosolic glutamate dehydrogenase which was comparatively higher in G. explanatum, catalyse amination of alpha-ketoglutarate. A high level of cytosolic arginase alone does not indicate a functional urea cycle. A tentative pathway of amino acid metabolism in these amphistomes is proposed.

Histochemical characterization of esterases from three species of amphistomes.

Esterase activity of Gigantocotyle explanatum, Ceylonocotyle scoliocoelium and Cotylophoron cotylophorum were localized histochemically in various regions associated with nutrition, viz, integument, pharynx, sucker and gastrodermis. The intensity of the reactions varied in the different regions of the same parasite and also from species to species. The distribution and relative activity of the esterases were investigated and the results reveal that the influence of inhibitors/activators on enzyme activity varied in the different regions. Based on the inhibitory effects it is inferred that carboxylesterases are distributed in the region of the integument, pharynx and gastrodermis, acetylesterases in the suckers and cholinesterase in the subtegumental cells and muscle of these three trematodes.

The influence of Calicophoron microbothrium on the susceptibility of Bulinus tropicus to Schistosoma bovis.

A total of 480 snails were collected from 3 habitats on the Mau Escarpment, Kenya, and were identified as Bulinus tropicus. Of the 351 snails examined alive in London, 75 were infected with Calicophoron microbothrium, 39 with C. microbothrium and Schistosoma bovis, 1 with S. bovis, 24 with other species of trematodes and 212 were uninfected. Examination of digestive glands of B. tropicus either uninfected or infected with both C. microbothrium and S. bovis demonstrated that it is possible to differentiate between parasite and host enzyme activity using glucose phosphate isomerase. However, malate dehydrogenase enables a much clearer differentiation between the enzyme activity of the schistosome and that of the amphistome. Laboratory snail infection experiments demonstrated that it is possible successfully to infect B. tropicus with S. bovis if the snails have previously been exposed to miracidia of C. microbothrium.

Ultrastructure and cytochemistry of the tegument of Orthocoelium scoliocoelium and Paramphistomum cervi (Trematoda: Digenea).

The tegument of Orthocoelium scoliocoelium and Paramphistomum cervi was examined using histochemical techniques and electron microscopy. On the basis of the distribution of acid and alkaline phosphatase (E.C. 3.1.3.2, E.C. 3.1.3.1), non-specific esterase (E.C. 3.1.1.1), cholinesterase (E.C. 3.1.1.7) and succinate dehydrogenase (E.C. 1.3.99.1) at light microscope level two distinct regions were recognized, an outer and an inner zone. Electron microscopy revealed that the tegument comprises an outer surface syncytium underlain by a thick subsyncytial zone and musculature. Deeper still occur the nucleated "tegumental cells". The latter are in cytoplasmic continuity with the surface syncytium via vacuolated cytoplasmic trabeculae which traverse the muscle layers and the subsyncytial zone. Three types of tegumental cells each lacking mitochondria were observed. The T1 cells synthesize discoid and electron dense T1 bodies while T2 cells produce oval and electron lucent T2 bodies. The third type of tegumental cells apparently produce no secretory bodies and may represent an embryonic cell type. The surface syncytium contains T1 and T2 secretory bodies and is bounded apically by a plasma membrane invested externally by a fuzzy and filamentous glycocalyx. The surface syncytium lacks mitochondria and is traversed by infoldings of the basal plasma membrane. Beneath the surface syncytium the subsyncytial zone is largely comprised of fibrous interstitial material. This zone, which is particularly thick in the amphistomes, is traversed by trabeculae and extensions of underlying parenchymal cells which usually contain mitochondria and lysosomes. The subsyncytial zone overlies numerous circular and longitudinal muscle fibres. The absence of mitochondria and enzymes associated with active transport suggests that the amphistome tegument may be mainly specialized for protection of the worm against mechanical and chemical conditions prevailing in the rumen. Active uptake of nutrients is probably not a primary function.

Fermentation and the properties of some enzymes of carbohydrate metabolism in the trematode Calicophoron ijimai.

Glycogen content, glucose consumption and the production of metabolic end products by Calicophoron ijimai were determined under aerobic and anaerobic conditions. The major end products of fermentation were identified as lactic, acetic, propionic, isobutyric and alpha-methylbutyric acids, propionic acid predominating. The activities and properties of some of the enzymes of carbohydrate metabolism were determined. The worms showed high phosphoenolpyruvate carboxykinase, malate dehydrogenase and malate dehydrogenase (decarboxylating) but relatively low pyruvate kinase and very low lactate dehydrogenase activities. The pH optima, coenzyme, cofactor and ionic requirements of the enzymes were similar to those of other helminths. Malate dehydrogenase had an 8-fold greater affinity for oxaloacetate than malate, and was about 14 times more active for oxaloacetate reduction than malate oxidation. Phosphoenolpyruvate carboxykinase was 2.4 times more active and had a 2-fold greater affinity for phosphoenolpyruvate and dinucleotide than pyruvate kinase. The low activities of lactate dehydrogenase and pyruvate kinase but high activities of malate dehydrogenase and phosphoenolpyruvate carboxykinase suggest that anaerobic carbohydrate catabolism follows the fumarate reductase pathway.

Morphological and histochemical studies on the ovarian development and oogenesis in Paramphistomum cervi (Digenea: Paramphistomatidae).

The ovary in 4-week-old worms contains undifferentiated germ cells. The oogonial differentiation, together with the appearance of some PAS-positive material in the ooplasm occurs in the ovary of 8-week-old worms. The ovary of 16-week-old worms contains only oogonia and oocytes and no germ cells. In adult worms, the oocytes reach the pachytene stage and further development is arrested at this stage. Nucleolar fragmentation and subsequent transport of nuclei to the ooplasm have been observed. The ooplasm contains nutritive material rich in proteins, carbohydrates and lipids. Its amount varies with different stages of oogenesis. The localization of various phosphatases and dehydrogenases was studied during oogenesis and their functional significance was discussed.

[Cytosol malate dehydrogenase in Calicophoron ijimai trematodes and the effect of antiparasitic preparations on its activity]. / Tsitozol'naia malatdegidrogenaza v trematodakh Calicophoron ijimai i vliianie ne ee aktivnost' nekotorykh protivoparazitarnykh preparatov.

The activity and properties of malate dehydrogenase (MDH; EC 1.1.1.37) and of "malic" enzyme (EC 1.1.1.40) in cytosole of the trematode C. ijimai were determined. The activity of MDH directed to oxaloacetate formation was shown to be 14 times and maximum velocity 13 times lower than that of the reverse reaction. The apparent KM was one order higher in the direct reaction. This confirms the possibility of glycolytic pathway in C. ijimai via CO2 fixation into phosphoenolpyruvate to form oxaloacatate which is readily eliminated by active MDH. The presence of "malic" enzyme in C. ijimai testifies to the occurrence of different pathways of succinate formation in this species.

Isozyme pattern of lactate and malate dehydrogenases of Gastrothylax crumenifer (Trematoda: Amphistomatidae) from different hosts.

Isozyme pattern of lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) of Gastrothylax crumenifer from sheep, goat and buffalo was studied using polyacrylamide gel electrophoresis. LDH of G. crumenifer from buffalo, goat and sheep consists of four fractions, three fractions and two fractions, respectively. The parasite from buffalo shows two fractions of MDH, whereas those from goats or sheep show only a single fraction. The significance of these results is discussed.

Non-specific alkaline phosphomonoesterases of eight species of digenetic trematodes.

Alkaline phosphatases from different trematodes occupying the same habitat have identical pH otima but different levels of enzyme activities. Isoparorchis hypselobagri, from the fish Wallago attu, shows four to six times more enzyme activity than Fasciolopsis buski, Gastrodiscoides hominis and Echinostoma malayanum, from the pig Sus scrofa, and Fasciola gigantica, Gigantocotyle explanatum, Cotylophoron cotylophorum and Gastrothylax crumenifer, from the buffalo Bubalus bubalis. At least two peaks of activity at different levels of pH were obtained for each trematode examined. Both Gastrodiscoides hominis and Isoparorchis hypselobagri enzymes had three peaks of alkaline phosphatase activity. The optimum temperature for maximum enzyme activity was 40 degrees C, above which rapid inactivation occurred. At temperatures below 40 degrees C, the enzymes of fish and mammalian trematodes did not behave similarly; I. hypselobagri enzyme being active over a wider range of temperature (20 degrees-40 degrees C. Various concentrations of KCN and arsenate proportionately inhibited enzyme activity. NaF Did not significantly influence enzyme activity, while Mg++ and Co++ acted as activators. The extent of inhibition or activation of enzyme activity of different trematodes varied, probably due to species differences. Both inhibition and activation of I. hypselobagri enzyme was higher than in the case of other trematodes.