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PubMed2000

Serological responses of cattle after treatment and during natural re-infection with Fasciola hepatica, as measured with a dot-ELISA system.

The dot-ELISA reaction was used to study the dynamics of IgG titers in cattle naturally infected with Fasciola hepatica after anthelmintic treatment and during reinfection. Excretion/secretion products (ES) of the parasite were used as antigens for the dot-ELISA. IgG antibodies were no longer detectable by dot-ELISA, 4-6 months after nine animals received the first of three weekly doses of triclabendazole (15 mg kg(-1)) and were then maintained on a pasture free of F. hepatica metacercariae. Six fluke-free cattle began shedding F. hepatica eggs 3-6 months after grazing a pasture contaminated with metacercariae of the parasite. A detectable increase in dot-ELISA IgG antibody levels was observed 2-4 weeks after natural reinfection by grazing a similar pasture contaminated with F. hepatica metacercariae. The usefulness of the dot-ELISA system to diagnose chronic infection by serology is complicated by previous treatment against the parasite. It is concluded that the ES antigens can be useful to detect early infection of cattle with F. hepatica in a dot-ELISA system

Animals↗
PubMed1984

Fasciola hepatica: the effects of neuropharmacological agents upon in vitro motility.

The effects of a wide range of neuropharmacological agents on the motility in vitro of Fasciola hepatica have been determined using an isometric transducer system. The neuromuscular blocking agents tubocurarine and decamethonium cause a long-term stimulation of the basal activity of the fluke. Acetylcholine causes an inhibition of activity. This effect is mimicked by the cholinergic agonists carbachol and nicotine, antagonised by the cholinergic blocking agents atropine and mecamylamine, and potentiated by eserine, a cholinesterase inhibitor. With nicotine and atropine the effects are accompanied by an increase in muscle tone at a concentration of 1 X 10(-2) M. Noradrenaline and adrenaline also cause some inhibition of activity, an effect antagonised by guanethidine, which blocks the release of noradrenaline. In contrast, dopamine stimulates fluke motility, whilst its antagonist dihydroergotamine causes an inhibition of activity. The monoamine oxidase inhibitors iproniazid and p-chloromercuribenzoic acid induce a stimulation of activity; with the latter there is an increase in muscle tone at a concentration of 1 X 10(-3) M. The amine depleting agents chloroamphetamine and reserpine, and the monoamine uptake inhibitors desipramine and nortriptyline produce an inhibition of fluke activity, as does the serotonin uptake inhibitor fluoxetine. High concentrations of chloroamphetamine (1 X 10(-2) M) and the uptake inhibitors (1 X 10(-3) M and above) also induce an increase in muscle tone. Serotonin causes a marked stimulation of motility. The pharmacological evidence is consistent with a neurotransmitter role of acetylcholine (inhibitory), dopamine (excitatory), and noradrenaline (inhibitory). The status of serotonin is discussed.

Animals↗
PubMedDate not supplied

[Reserve nutrients of the trematode Fasciola hepatica at various stages in its ontogeny].

A comparative histochemical study for the presence of reserve nutrients (glycogen, fat) in tissues, organs of parthenitae (sporocysts, rediae) and larvae (miracidia, cercariae, adolescariae) of Fasciola hepatica from liver of sheep and cattle was carried out. The glycogen amount prevails at all developmental stages of Fasciola from sheep as compared to those from cattle. Fat was found in tissues and organs of parthenitae and larvae of this trematode in equal amount. Its reserves are obviously smaller than those of glycogen.

Animal Nutritional Physiological Phenomena↗
PubMed2000

Fasciola hepatica infection downregulates Th1 responses in mice.

Immune responses induced with helminth parasites have been extensively studied, but there is limited information on those to Fasciola hepatica, especially on the subtype of T cell induced with this parasite. We investigated the local and systemic T cell responses of different strains of mice following oral infection with doses of metacercariae from F. hepatica. Spleen cells from BALB/c and 129Sv/Ev mice given a low-dose (5 metacercariae) infection exhibited a Th2 response, producing high levels of the cytokines IL-4 and IL-5, and low levels of IFN-gamma and IL-2. In contrast, C57BL/6 mice showed a mixed Th1/Th2 response. A more marked polarization to a Th2 response was observed in BALB/c, 129Sv/Ev exposed to a high-dose (15 metacercariae) infection and the C57BL/6 mice also exhibited a clear Th2 response. IL-4 defective (IL-4-/-) C57BL/6 mice infected with 5 metacercariae produced less IFN-gamma and more IL-5 compared to their wild-type C57BL/6 counterparts, suggesting that IL-4 is important in establishing the Th2 type response in murine fasciolosis. However, the secretion of IFN-gamma and IL-2 was completely suppressed in the high-dose infection and this was also observed in IL-4-/- mice. Thus, liver flukes may secrete molecules that downregulate Th1 responses. T cell responses in the mesenteric (MLN) and hepatic lymph nodes (HLN) were also examined since newly excysted juveniles infect through the intestinal wall of their host before migrating to the hepatic tissue. Cells from both MLN and HLN secreted higher levels of IL-4 and IL-5 compared to spleen cells. We also observed a difference in cytokine profiles secreted by the MLN and HLN, which may reflect responses to antigens liberated by newly excysted juveniles and hepatic stage parasites, respectively.

Animals↗
PubMed2003

Fasciola hepatica cathepsin L-like proteases: biology, function, and potential in the development of first generation liver fluke vaccines.

Fasciola hepatica secretes cathepsin L proteases that facilitate the penetration of the parasite through the tissues of its host, and also participate in functions such as feeding and immune evasion. The major proteases, cathepsin L1 (FheCL1) and cathepsin L2 (FheCL2) are members of a lineage that gave rise to the human cathepsin Ls, Ks and Ss, but while they exhibit similarities in their substrate specificities to these enzymes they differ in having a wider pH range for activity and an enhanced stability at neutral pH. There are presently 13 Fasciola cathepsin L cDNAs deposited in the public databases representing a gene family of at least seven distinct members, although the temporal and spatial expression of each of these members in the developmental stage of F. hepatica remains unclear. Immunolocalisation and in situ hybridisation studies, using antibody and DNA probes, respectively, show that the vast majority of cathepsin L gene expression is carried out in the epithelial cells lining the parasite gut. Within these cells the enzyme is packaged into secretory vesicles that release their contents into the gut lumen for the purpose of degrading ingested host tissue and blood. Liver flukes also express a novel multi-domain cystatin that may be involved in the regulation of cathepsin L activity. Vaccine trials in both sheep and cattle with purified native FheCL1 and FheCL2 have shown that these enzymes can induce protection, ranging from 33 to 79%, to experimental challenge with metacercariae of F. hepatica, and very potent anti-embryonation/hatch rate effects that would block parasite transmission. In this article we review the vaccine trials carried out over the past 8 years, the role of antibody and T cell responses in mediating protection and discuss the prospects of the cathepsin Ls in the development of first generation recombinant liver fluke vaccines.

Animals↗
PubMed1992

Isoelectric focusing of soluble proteins from Fasciola hepatica L, 1758 and Fascioloides magna B, 1875.

Isoelectric focusing was performed on the soluble proteins of whole-body and excretory-secretory products (ESP) of Fasciola hepatica and Fascioloides magna. Adult F hepatica flukes were recovered from experimentally infected sheep and ESP obtained from the flukes; portions of liver were cut and frozen at -70 C. Fascioloides magna adults were collected from naturally infected white-tailed deer and ESP obtained; portions of liver were collected from noninfected white-tailed deer. Adult flukes and their host tissues were homogenized and centrifuged; protein concentrations with their ESP were determined and adjusted to less than 2.50 mg/ml. Seven ESP samples from F hepatica and 1 from Fascioloides magna were subjected to isoelectric focusing with the 2 species of fluke and their respective host liver homogenates. After separation, gels were stained with silver and scanned on a laser densitometer. Protein banding patterns of the 2 species of flukes were dissimilar. In the pH range of 3.5 to 9.6, the body protein had approximately 30 peaks and ESP about 23 peaks in both species. Overall banding patterns of the body protein and ESP of both species were distinct from those of respective host tissues. Of the peaks reported as dominant, 3 of the body protein and 2 of ESP were shared between the 2 species. Fascioloides magna had more dominant peaks than F hepatica. This technique of soluble protein isoelectric focusing is simple and reproducible, and the 2 fluke species can easily be differentiated by this technique, as well as by morphologic characteristics.

Animals↗
PubMed2006

Exposure to Fasciola hepatica miracidia increases the sensitivity of Lymnaea (Fossaria) humilis to high and low pH.

Humidity and temperature have been considered important factors affecting the infectivity of Fasciola hepatica to its molluscan host. One hundred and thirty laboratory-reared Lymnaea humilis were exposed for 4 hr to the miracidia of F. hepatica over a pH range from 4.0 to 10.0, and their rates of survival were compared with 130 similarly treated but unexposed control snails. All control snails died within 24 hr at pH 4.0, but they showed better survival at pH 5.0-10.0. Their sensitivity to solutions with high and low pH, however, was increased if kept in the presence of F. hepatica miracidia. Snails exposed at pH 5.0 died within 24 hr, whereas most other pHs also affected survival such that by day 18 only those snails exposed at pH 7.2 remained alive. The increased sensitivity of the snails to pH could be explained by a damage-mediated release of parasite enzymes, because infectivity was highest at pHs associated with the lowest host mortality.

Animals↗
PubMed1987

The relationship between the density of Fasciola hepatica miracidia and the net rate of miracidial infections in Lymnaea truncatula.

Experimental studies on the dynamics of infection of Lymnaea truncatula by the miracidia of Fasciola hepatica show that there is a linear relationship between the net rate of attachment of miracidia to snail epithelium and the density of infective stages. The probability of a snail remaining uninfected after exposure to a single miracidium varies with the size class of the snail. Under the conditions of the experiment (temperature, 15 degrees C; exposure period, 30 min; water volume, 8 ml), the probability was 0.46-0.48, 0.27-0.26 and 0.13-0.19 for snail size classes, 0.5-2.9, 3.0-5.9 and 6.0-8.9 mm respectively.

Animals↗
PubMed1983

Protection against Fasciola hepatica in rats with adult fluke antigen in Freund's adjuvant: influence of antigen batch, antigen dose and number of sensitising injections.

Protection of rats against Fasciola hepatica infection induced by intraperitoneal injection of adult fluke antigen in Freund's adjuvant was dependent on the batch of antigen. Relatively modest protection of 41 per cent and 55 per cent provided by some batches at one dose could be improved to 65 to 84 per cent and 86 per cent respectively by increasing the amount of antigen used for sensitisation. Protection was found to require at least two sensitising injections. Subcutaneous sensitisation with adult fluke antigen in Freund's adjuvant did not protect, whereas when this was given in conjunction with intraperitoneal injections of saline in Freund's adjuvant protection was achieved.

Animals↗
PubMed1975

Prenatal infection with Fasciola Hepatica in calves.

The livers from 16,776 calves 1 to 3 weeks of age slaughtered at Shepparton abattoirs were examined for gross evidence of infection with Fasciola hepatica. Those livers with thickened bile ducts were examined for the presence of liver fluke by slicing. One hundred and eight F. hepatica were collected from 84 of these livers, giving a minimum prevalence rate of prenatal infection of calves of 0.5%. These results are discussed in context with reports from other countries.

Animals↗
PubMed1999

Short report: Immunodiagnosis of human fascioliasis using recombinant Fasciola hepatica cathepsin L1 cysteine proteinase.

Our laboratory recently developed a diagnostic test (ELISA) for human fascioliasis based on the detection of serum IgG4 antibodies reactive with Fasciola hepatica cathepsin L1 (CL1). In the present study, we have used recombinant CL1, generated by functional expression of the cDNA in Saccharomyces cerevisiae, in this immunodiagnostic test and compared its performance with native CL1. Sera obtained from 64 individuals living in Cutusuma village in the northern Altiplano of Bolivia, a region with a high prevalence of human fascioliasis, were analyzed by the IgG4-ELISA. A highly statistically significant correlation (r2 = 0.751, P < 0.001) was demonstrated between the absorbances obtained using the recombinant and native proteins. These assays showed that 38 (59%) of the individuals tested were seropositive for fascioliasis, whereas only 26 of them were coprologically positive for F. hepatica eggs. All seronegative patients were also coprologically negative. Serum from individuals infected with schistosomiasis mansoni, cysticercosis, hydatidosis, and Chagas disease did not contain antibodies reactive with the recombinant or native CL1. Therefore, recombinant CL1 shows excellent potential for the development of the first standardized assay for the sensitive and specific diagnosis of human fascioliasis. Finally, our data supports earlier reports on the high prevalence of human fascioliasis in the Bolivian Altiplano, which collectively suggest that the disease has been endemic there for more than a decade.

Adolescent↗
PubMed1983

Fasciola hepatica: effects of diamfenetide free amine on in vitro physiology, biochemistry, and morphology.

Short-term (1-3 hr) incubations in vitro of immature and adult Fasciola hepatica with 10(-4) to 10(-5) M free amine of diamfenetide (DPT-FA) demonstrated a time/dose-dependent, irreversible paralysis that involved an increase in muscular tension and decrease in contraction amplitude. The following events occurred preceding or concomitant with the paralysis: influx of Na+, decrease in surface membrane potential, increase in wet weight, swellings on the ventral surface, and inhibition of 3-O-methyl glucose transport. These events were all consistent with a disturbance in surface membrane functions. The effects of DPT-FA were more severe in immature flukes (3-5 weeks postinfection) than adults which agrees with observed in vivo efficacy.

3-O-Methylglucose↗
PubMed1986

Heterologous resistance to Fasciola hepatica conferred upon rats by passive transfer of serum from different breeds of sheep.

Two experiments compared the protection against oral challenge with 20 metacercariae of Fasciola hepatica conferred on rats by intraperitoneal injection of serum from three breeds of sheep infected with F. hepatica (Barbados Blackbelly, St. Croix, Florida Native). Experiment 1 used serum from sheep 5-6 months of age following two infections of 250 metacercariae each, while Experiment 2 utilized serum collected from the same sheep at 10-11 months of age following either a primary (first exposure) or challenge (after two previous exposures of 250 metacercariae each) infection with 500 metacercariae. Similar numbers of flukes were recovered from rats given either immune or nonimmune (control) serum from each breed of sheep in Experiment 1. In Experiment 2, rats given serum from infected St. Croix sheep had significantly fewer flukes than rats given either control or immune serum from Barbados Blackbelly or Florida Native sheep. There was no significant correlation of fluke counts between individual serum donors (sheep) and serum recipients (rats).

Analysis of Variance↗
PubMed1988

Histochemical localization of acetylcholinesterase in the cerebral ganglia of Fasciola hepatica, a parasitic flatworm.

Acetylcholinesterase activity was found in the cell bodies and extracellularly in the neuropile of the cerebral ganglia of the adult trematode parasite, Fasciola hepatica. Within neuronal cell bodies of the cerebral ganglion, acetylcholinesterase reaction product was found in the endoplasmic reticulum, in the cisternae of the Golgi apparatus, and in secretory vesicles near the inner (releasing face) cisternae. Acetylcholinesterase reaction product was not seen intracellularly within any nerve processes. The reaction product was found around the somatic cell membranes and in the extracellular space between closely apposed nerve processes in the neuropile. Acetylcholinesterase reaction product was associated with synaptic endings that contained clear spheroidal synaptic vesicles, and the reaction product was localized at the site of synaptic contact between the zone of apposition of the pre- and postsynaptic terminals. This intracellular and extracellular distribution of the enzyme is consistent with its function as the degrading enzyme in cholinergic transmission.

Acetylcholinesterase↗
PubMedDate not supplied

Triclabendazole biotransformation and comparative diffusion of the parent drug and its oxidized metabolites into Fasciola hepatica.

Triclabendazole (TCBZ) is an halogenated trematodicidal benzimidazole compound extensively used in veterinary medicine. It is active against immature and adult stages of the liver fluke Fasciola hepatica. Free and conjugated TCBZ metabolites have been identified in the bile of treated sheep. The experimental aims were to characterize the in vitro patterns of TCBZ biotransformation both in the animal host (sheep liver microsomes) and target parasite (F. hepatica microsomal preparation); and to compare the ex vivo diffusion of TCBZ parent drug and its oxidized metabolites (TCBZ sulphoxide [TCBZSO], TCBZ sulphone [TCBZSO2], and TCBZ-hydroxy derivatives) into F. hepatica. Additionally, the octanol-water partition coefficients for TCBZ and all its metabolites were estimated as an indicator of the relationship between drug lipophilicity and diffusion into the target parasite. Drug/metabolites concentrations were quantified by HPLC after sample clean up and a solvent-mediated chemical extraction. Sheep liver microsomes metabolized TCBZ into its sulphoxide and sulphone metabolites after 30 min of incubation. The rate of TCBZ sulphoxidation in the liver was significantly greater (p < 0.01) than that observed for the sulphonation of TCBZSO. The trematode parasite oxidized TCBZ into its sulphoxide metabolite after 60 min of incubation at a metabolic rate of 0.09 nmol min(-1) mg protein(-1). TCBZ and all its oxidized metabolic products were recovered from F. hepatica as early as 15 min after their ex vivo incubation in a Kreb's Ringer Tris buffer. However, the diffusion of the hydroxy-derivatives into the fluke was lower than that observed for TCBZ, TCBZSO and TCBZSO2. There was a high correlation (r=0.82) between drug lipophilicity (expressed as octanol-water partition coefficients) and drug availability measured within the parasite. Unlike the uptake pattern previously observed for albendazole, the parent TCBZ and its sulphoxide and sulphone metabolites showed a similar ability to penetrate into the trematode parasite. Understanding the relationship between TCBZ metabolism, the relative pharmacological potency of its metabolic products and their ability to reach the target parasite may be critical to optimize its flukicidal activity, particularly when TCBZ resistant flukes have been already isolated in the field.

Animals↗
PubMed2002

Patterns in size and shedding of Fasciola hepatica eggs by naturally and experimentally infected murid rodents.

Using samples collected on the island of Corsica, a comparative study was done of the morphometry of Fasciola hepatica eggs shed by cattle and by naturally and experimentally infected murid rodents (wild Mus musculus and Rattus rattus and Rattus norvegicus Wistar laboratory strain). Eggs shed by murids are smaller in size than those shed by naturally infected cattle. A second study analyzed the number of F. hepatica eggs shed in murid feces at different time intervals, i.e., months, days, and 6-hr periods, by the Kato-Katz technique. Both experimentally and naturally infected black rats (R. rattus) were used, and Wistar rats were experimentally infected and included for comparison. The present studies prove that black rats R. rattus are able to shed eggs independently from the liver fluke isolate and that egg shedding occurs throughout the life of this host species, uninterrupted during all the months analyzed in a 2-yr period. Moreover, the results suggest that this shedding is continuous, with eggs appearing in the feces daily. The results on egg shedding by wild black rats R. rattus reach their maximum shedding in spring and autumn and a maximum during twilight hr. These chronobiological patterns appear to favor parasite transmission, both seasonally and daily.

Animals↗
PubMed1984

Effects of diamfenetide on metabolic and excretory functions of Fasciola hepatica in vitro.

The effect of diamfenetide (DFT) on the time course of production of end-products of glucose metabolism, tissue ATP levels and NH3 production by adult Fasciola hepatica in vitro was determined. Products of glucose metabolism are increased between 6 and 24 hr incubation in 10(-4) M DFT. Tissue ATP levels and NH3 production are decreased during this time period. The observed metabolic effects of DFT are manifested at a much later time after drug exposure than previously described membrane-disruptive events indicating that metabolic effects of DFT on F. hepatica may be secondary to the initial effects on surface membranes.

Acetanilides↗