S.M.A. Abidi

Biochemical characterization of Taenia hydatigena cysticerci from goats and pigs.

Analysis of the major biochemical components of Taenia hydatigena cysticerci collected from goats and pigs showed marked differences, particularly in glycogen, protein, lipid and DNA levels. Differences were also detected in the levels of cholesterol, triglycerides, free fatty acids and phospholipids. Furthermore, the profile of phospholipid fractions revealed quantitative differences between the two species. It is concluded that the cysticerci of goat and pig origin probably represent two different strains and possibly follow the same pattern of speciation as reported in the related taeniid, Echinococcus granulosus.

Incidence and histopathology of encysted progenetic metacercaria of Clinostomum complanatum (Digenea: Clinostomidae) in Channa punctatus and its development in experimental host

OBJECTIVE To study the incidence of encysted progenetic metacercariae of Clinostomum complanatum (C. complanatum) in Channa punctatus (C. punctatus), associated histopathology and the experimental infection to laboratory chicken to obtain ovigerous adult worms. METHODS Live C. punctatus were brought from local fish market of Aligarh, India, dissected and examined on a monthly basis for the presence of C. complanatum cysts. For histochemistry, infected tissue sections with attached cysts were processed for haematoxylene and eosin staining. Cysts were aseptically fed to 4 day old leghorn chicken to obtain adult worms. Mechanically excysted metacercaria and the ovigerous adult worms were stained in carmine to prepare permanent slides. RESULTS One year survey for the infection of encysted progenetic metacercaria of C. complanatum in C. punctatus revealed the prevalence, intensity and abundance of 24.7%, 2.27 and 0.608, respectively. Histopathology showed heavy infiltration of immune cells at the site of cyst attachment and some tissue damage was also evident. Following feeding to experimental chicken, about 41.07% of the encysted metacercariae were able to excyst and migrate back to bucco-pharyngeal region where they tenaciously attached and fed on blood, and transformed into ovigerous adult worms from 62 hours onwards of post infection. CONCLUSIONS The parasite is potentially pathogenic to the host, and the availability of a suitable intermediate host can be a contributing factor for the occurrence of C. complanatum metacercaria either in the excysted or encysted form, indicating loose host specificity and zoonotic potential.

Time-dependent tegumental surface changes in juvenile Fasciola gigantica in response to triclabendazole treatment in goat

Triclabendazole (TCBZ), the anthelmintic drug active against both mature and immature liver flukes, was used to investigate the effect of in vivo treatment on the tegumental surface of juvenile Fasciola gigantica. Five goats were infected with 150 F. gigantica metacercariae each by oral gavage. Four of them were treated with single dose of TCBZ at 10mg/kg at four weeks post-infection. They were euthanized at 0 (untreated), 24, 48, 72 and 96h post treatment. Juvenile flukes were manually retrieved from the goat livers and processed for scanning electron microscopy. In control flukes, the anterior region was adorned with sharply pointed spines projecting away from the surface, while in the posterior region, spines become shorter and narrower, loosing serration and with the appearance of distinct furrows and papillae. The dorsal surface retained the same pattern of surface architecture similar to that of ventral surface. Flukes obtained from 24h post-treatment did not show any apparent change and were still very active. However, there were limited movements and some blebbing, swelling, deposition of tegumental secretions and some flattening displayed by the flukes of 48h post-treatment. All the worms were found dead 72h post-treatment and showed advanced level of tegumental disruptions, consisting of severe distortion of spines, sloughing off the tegument to expose the basal lamina, formation of pores and isolated patches of lesions. By 96h post-treatment, the disruption was extremely severe and the tegument was completely sheared off causing deeper lesions that exposed the underlying musculature. The disruption was more severe at posterior than anterior region and on ventral than dorsal surface. The present study further establishes the time-course of TCBZ action in vivo with 100% efficacy against the juvenile tropical liver fluke.

Monoamine oxidase in amphistomes and its role in worm motility.

The quantitative assay of mitochondrial monoamine oxidase (MAO) activity revealed a higher enzyme level in Explanatum explanatum than Gastrothylax crumenifer. The specific MAO inhibitors, chlorgyline, pargyline, deprenyl and nialamide produced different degrees of interspecific inhibition. The differential effects on enzyme activity of chlorgyline and deprenyl suggests the possible existence of polymorphic forms of the enzyme, MAO-A and MAO-B, in amphistomes. These specific inhibitors also had a differential influence on the in vitro motility of amphistomes, further indicating the involvement of different forms of MAO in the oxidative deamination of biogenic monoamines which might be partly responsible for neuromuscular coordination in amphistomes. The experimental procedures used in this study could be conveniently used for quick screening and evaluation of some of the qualitative effects of anthelmintic drugs under in vitro conditions.

A rabbit eye model for in vivo transformation of progenetic metacercariae of Clinostomum complanatum into ovigerous adult worms

Clinostomum complanatum is a digenetic trematode that causes yellow grub disease in some fish species and also shows zoonotic potential by sporadically infecting humans. In this study, progenetic metacercariae of C. complanatum were obtained from the fish Trichogaster fasciatus, and were aseptically placed in conjunctival incisions made in the superior and inferior fornices of the eye of rabbits, which served as the experimental hosts. Worms were harvested without necropsy of the host on days 4 and 8 post infection, to observe in vivo transformation of the progenetic metacercariae into ovigerous adult worms. The worms appeared to cause minimal damage to the host although they were tenaciously attached. In vivo maturation was evident by the development of the vitellaria, enlargement of gonads, the presence of a large number of shelled eggs in a distended uterus and ramifications of the intestinal caeca. Obtaining mature ovigerous worms without sacrificing the host clearly gives the rabbit eye model an advantage over those described previously. Due to the relative advantage of the short time required for maturation and the prolific egg production by C. complanatum, it is suggested that this host-parasite system could be used as an excellent model for classroom teaching of trematode biology and to investigate the cues involved in in vivo transformation and host-parasite interactions.

Levels of some antioxidant molecules and lipid peroxidation during in vivo transformation of the progenetic metacercaria of Clinostomum complanatum to ovigerous adult worms.

The levels of oxidative stress markers are an important indicator of the physiological state of the parasite and its host. In the present study levels of lipid peroxidation, glutathione S transferase, glutathione, superoxide dismutase and catalase were determined in the Clinostomum complanatum progenetic metacercaria, obtained from the fish peritoneum (a hypoxic habitat). The in vivo transformed ovigerous adult worms were obtained from the aerobic environment of the buccopharyngeal region of experimentally infected chickens. Levels of antioxidant molecules were also determined in the blood of experimentally infected chickens. An increase in the levels of lipid peroxidation, and a significant decrease in the levels of glutathione S transferase, glutathione, superoxide dismutase and catalase was observed in the infected host as compared to the controls. In the ovigerous worms, the levels of lipid peroxidation, glutathione S transferase, glutathione, superoxide dismutase were found to be significantly less than the levels observed in the progenetic metacercaria. Since the establishment of worm in the buccal cavity of the avian host would lead to its exposure to oxygen and the haematophagous nature of the parasite also exposes it to the free radicals in the host blood, the progenetic metacercaria has evolved to produce excess free radical scavenging molecules reserved to combat the oxidative stress encountered within the microhabitat of the definitive host.

Biochemical variations in Gigantocotyle explanatum and Gastrothylax crumenifer with respect to their seasonal reproduction

Biochemical components, glycogen, protein, nucleic acids and lipid fractions were analysed every month from January to December 1986, in the liver and rumen amphistomes Gigantocotyle explanatum and Gastrothylax crumenifer, respectively. The results reveal a considerable seasonal variation. In the rumen amphistome the components reach their maximum level only once a year, whereas in the liver amphistome, more than one peak is observed in a year. In G. crumenifer the content of glycogen and nucleic acids increases before the onset of egg production while protein and lipids reach their maximum level during the egg production phase. The variations in biochemical components were associated with the reproductive cycles and gonad recrudescence of these parasites.

[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.

Biochemical changes during the development of the miracidium of Gigantocotyle explanatum

Analysis of various biochemical components during the development of the miracidium of G. explanatum showed marked changes, particularly in glycogen, protein and DNA levels. Though the total lipids remained more or less unchanged, alterations in the levels of cholesterol, triglycerides, free fatty acids, phospholipids and phospholipid fractions were also recorded. Such changes could be intrinsically programmed for the cellular differentiation and organogenesis in larval amphistomes.

A comparative study of the protein content of some helminths and the suitability of assay methods

The protein content of fresh homogenates and their corresponding TCA precipitated fractions of 10 different species of helminths was estimated by the methods of Lowry et al. and Spector using the Folin phenol reagent and Coomassie brilliant blue G-250 respectively. The former method gives exaggerated values as compared to the latter method. The parasite phenols, phenolic proteins and catecholamines could be responsible for interference in the Lowrys procedure. The TCA noln-precipitable moieties also give colour only with the Folin phenol assay. The pronounced intra-specific differences in the total protein content of helminths reflect their metabolic variations and adaptations. Habitat does not appear to influence the protein content of parasites, however, the effect of host variation was evident in the pouched amphistome G. crumenifer. It is concluded that the dye binding method gives more consistent results and it can be conveniently applied to crude tissue homogenates of helminths.