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PubMed1995

Fasciola hepatica: rapid identification of newly excysted juvenile proteins.

The sensitivity of N-terminal sequencing has been used to identify proteins expressed by the newly excysted juvenile stage of the parasite Fasciola hepatica. Of the seven proteins identified, a number have significant sequence homology to the cysteine proteases: cathepsin B, cathepsin L and asparaginyl endoproteinase. Proteolytic activity was demonstrated using gelatin substrate sodium dodecyl sulphate polyacrylamide gel electrophoresis. In addition, a number of novel proteins were identified which shared no significant sequence homology to proteins in the databases. The availability of such N-terminal sequence information allows rapid identification of major proteins from scarce developmental stages and provides the basis for further molecular studies.

Amino Acid Sequence↗

PubMed1998

Developmental expression of a Fasciola hepatica sequence homologous to ABC transporters.

Polymerase chain reaction and cDNA library screening approaches were employed to identify a putative member of the highly conserved family of ATP-binding cassette transport proteins from Fasciola hepatica. At the predicted protein level, the F. hepatica sequence identified in the present study shares 43% and 36% identity with the Schistosoma mansoni SMDR2 and human MDR1 ATP-binding cassette transport sequences, respectively. Northern blot and reverse transcriptase-PCR analyses have demonstrated that expression of the F. hepatica ABC-transporter homologue is confined to immature parasites. The biochemical basis for the stage-specific expression of the ATP-binding cassette transporter homologue within F. hepatica remains to be determined.

ATP-Binding Cassette Transporters↗

PubMed2005

Thioredoxin peroxidase secreted by Fasciola hepatica induces the alternative activation of macrophages.

Alternatively activated macrophages (AAMphi) are primarily associated with the chronic stages of parasitic infections and the development of a polarized Th2 response. We have shown that Fasciola hepatica infection of BALB/c mice induces a polarized Th2 response during both the latent and chronic stage of disease. The activation status of macrophages was analyzed in this model of helminth infection by evaluating the expression of genetic markers of alternative activation, namely, Fizz1, Ym1, and Arg1. AAMphi were recruited to the peritoneum of mice within 24 h of F. hepatica infection and after intraperitoneal injection of parasite excretory-secretory (ES) products. Administration of a recombinant antioxidant thioredoxin peroxidase (TPx), which is contained within the ES products, also induced the recruitment of AAMphi to the peritoneum. In vitro studies showed that this recombinant TPx directly converts RAW 264.7 macrophages to an alternatively activated phenotype characterized by the production of high levels of interleukin-10 (IL-10), prostaglandin E(2), corresponding with low levels of IL-12. Our data suggest that the Th2 responses induced by the helminth F. hepatica are mediated through the secretion of molecules, one of which is TPx, that induce the recruitment and alternative activation of macrophages.

Animals↗

PubMed1999

[Exensivity of invasion Fasciola hepatica L. in cattle in the period 1957-1998 and valuation of losses caused by staggers disease among cattle in Poland, 1995-1997].

On the background of informations about frequency of appearance Fasciola hepatica L. among cattle in Poland in the time from 1995-1997, as well es in the former period, there is presented in the article an estimative apperiation of economical losses caused by staggers disease among cattle. The highest extensiveness of invasion of F. hepatica L. among cattle was noticed in the fifties and sixties manty in the north-eastern Poland. Losses caused by confiscation of a liver to amounted to 1 197 031 zł in 1995, 1 277 154 zł in 1996 and 1 312 078 zł in 1997.

Animals↗

PubMed2004

An ultrasensitive capture ELISA for detection of Fasciola hepatica coproantigens in sheep and cattle using a new monoclonal antibody (MM3).

A capture enzyme-linked immunosorbent assay (ELISA) using a new monoclonal antibody (mAb MM3) is reported for the detection of Fasciola hepatica excretory-secretory antigens (ESAs) in feces of infected hosts. The mAb MM3 was produced by immunization of mice with a 7- to 40-kDa purified and O-deglycosylated fraction of F. hepatica ESAs, which has previously been shown to be specific for the parasite. The specificity and sensitivity of the MM3 capture ELISA were assessed using feces from sheep and cattle. Sheep feces were obtained from a fluke-free herd (with most animals harboring other nematodes and cestodes), from lambs experimentally infected with 5-40 F. hepatica metacercariae and in some cases treated with triclabendazole at 14 wk postinfection (PI), and from uninfected control lambs. Cattle feces were collected at the slaughterhouse from adult cows naturally infected with known numbers of flukes (from 1 to 154) or free of F. hepatica infection (though in most cases harboring other helminths). The MM3 capture ELISA assay had detection limits of 0.3 (sheep) and 0.6 (cattle) ng of F. hepatica ESA per milliliter of fecal supernatant. The assay detected 100% of sheep with 1 fluke, 100% of cattle with 2 flukes, and 2 of 7 cattle with 1 fluke. The false-negative animals (5/7) were probably not detected because the F. hepatica individuals in these animals were immature (5-11 mm in length). As expected, coproantigen concentration correlated positively (r = 0.889; P < 0.001) with parasite burden and negatively (r = 0.712; P < 0.01) with the time after infection at which coproantigen was first detected. Nevertheless, even in animals with low fluke burdens (1-36 parasites), the first detection of F. hepatica-specific coproantigens by the MM3 capture ELISA preceded the first detection in egg count by 1-5 wk. In all sheep that were experimentally infected and then untreated, coproantigen remained detectable until at least 18 wk PI, whereas in sheep that were experimentally infected and then flukicide treated, coproantigen became undetectable from 1 to 3 wk after treatment. None of the fecal samples from sheep or cattle negative for fascioliasis but naturally infected with other parasites including Dicroelium dendriticum showed reactivity in the MM3 capture ELISA. These results indicate that this assay is a reliable and ultrasensitive method for detecting subnanogram amounts of F. hepatica antigens in feces from sheep and cattle, facilitating early diagnosis.

Animals↗

PubMed2004

Comparative assessment of the access of albendazole, fenbendazole and triclabendazole to Fasciola hepatica: effect of bile in the incubation medium.

The work reported here describes the comparative ability of albendazole (ABZ), fenbendazole (FBZ) and triclabendazole (TCBZ) to penetrate through the tegument of mature Fasciola hepatica, and the influence of the physicochemical composition of the incubation medium on the drug diffusion process. The data obtained from the trans-tegumental diffusion kinetic studies were complemented with the determination of lipid-to-water partition coefficients (octanol-water) for the benzimidazole (BZD) anthelmintic drugs assayed. Sixteen-week-old F. hepatica were obtained from untreated artificially infected sheep. The flukes were incubated (37 degrees C) over 60 and 90 min in incubation media (pH 7.4) prepared with different proportions of ovine bile and Krebs' Ringer Tris (KRT) buffer (100, 75, 50, 25 and 0% of bile) containing either ABZ, FBZ or TCBZ at a final concentration of 5 nmol/ml. After the incubation time expired, the liver fluke material was chemically processed and analysed by high performance liquid chromatography (HPLC) to measure drug concentrations within the parasite. Additionally, the octanol-water partition coefficients (PC) for each molecule were calculated (as an indicator of drug lipophilicity) using reversed phase HPLC. The 3 BZD molecules were recovered from F. hepatica at both incubation times in all incubation media assayed. The trans-tegumental diffusion of the most lipophilic molecules ABZ and FBZ (higher PC values) tended to be greater than that observed for TCBZ. Interestingly, the uptake of ABZ by the liver flukes was significantly greater than that measured for TCBZ, the most widely used flukicidal BZD compound. This differential uptake pattern may be a relevant issue to be considered to deal with TCBZ-resistant flukes. Drug concentrations measured within the parasite were lower in the incubations containing the highest bile proportions. The highest total availabilities of the 3 compounds were obtained in liver flukes incubated in the absence of bile. Altogether, these findings demonstrated that the entry of the drug into a target parasite may not only depend on a concentration gradient, the lipophilicity of the molecule and absorption surface, but also on the physicochemical composition of the parasite's surrounding environment.

Albendazole↗

PubMed1992

The overwintering of Fasciola hepatica eggs under semi-arid and temperate Mediterranean climate (Madrid, Spain).

In this experimental work, conducted over two consecutive years (1988 and 1989), we have analyzed the overwintering possibilities of Fasciola hepatica eggs deposited in winter in a semi-arid and temperate Mediterranean area (Madrid, Spain). Eggs did not survive for more than a week in faeces at environmental moisture conditions. In contrast, over a 75% of the eggs maintained in permanently wet faeces survived winter temperatures, showing no high mortality until important thermic increment from June. The success of overwintering was higher in eggs arrested in wet faeces than included in water (55% "versus" 20% of hatching in spring), and it was unrelated to the deposition time (excretion in early or late winter). The hatching period of overwintered eggs started in May, at the same time as those excreted in early spring. Therefore, overwintering of eggs will have an effect on the level but not on the start of the spring contamination with miracidia. The dry winters of this semi-arid area greatly restrict the overwintering eggs contamination to soils next to streams, ponds or irrigation channels.

Animals↗

PubMed1990

Glutathione S-transferase. Novel vaccine against Fasciola hepatica infection in sheep.

The potential of GST as a vaccine candidate against liver fluke infection in ruminants was studied by vaccinating sheep (n = 9) with GST purified from adult worms of Fasciola hepatica and challenging with 500 F. hepatica metacercariae. The immunization induced a high antibody response to GST in contrast to the poor or undetectable response to this Ag observed in naturally infected sheep. Throughout the trial, the progress of the fluke infection was monitored by measuring RBC hemoglobin levels, the extent of liver damage and the fecal egg output in the sheep. This analysis indicated that a subpopulation (n = 4) of the GST vaccinated animals exhibited no anemia, reduced liver damage and a lower mean fecal egg count relative to the infected control group suggesting a lower fluke burden in these animals. Worm burdens in the livers of the GST vaccine group (107 +/- 22) were 57% lower than in the infected control group (250 +/- 25). The subpopulation of the GST vaccine group demonstrated a 78% reduction in mean worm burdens relative to the control group. These results show that GST of adult F. hepatica is a novel Ag that can significantly protect sheep against liver fluke infection. The results suggest that the immune response to GST is directed to the juvenile worm reducing the number of worms that can establish in the liver of the vaccinated animals.

Animals↗

PubMed1981

Stimulation of resistance to Fasciola hepatica infection in sheep by a regime involving the use of the immunomodulatory compound L tetramisole (levamisole).

The effect of prior nematode and cestode infection in sheep on the course of experimental Fasciola hepatica was studied. The cross immunising properties of these parasites per se and the modification of any protective effect conferred by the immunomodulatory compound L tetramisole (levamisole) were investigated. It was recorded that no protection was conferred on sheep against F hepatica infection by virtue of prior infection with any helminth species per se, including Cysticercus tenuicollis. However, a degree of immunity to F hepatica was recorded, for only the second time, in sheep primed with helminth infections before treatment with levamisole. This is the first demonstration of a beneficial effect conferred by levamisole by virtue of its immunomodulatory properties in ruminants infected with any helminth species. Resistance was shown by a 47 per cent reduction in flukes recovered from sheep treated in this way (P less than 0.05).

Animals↗

PubMed1987

Fasciola hepatica: tegumental surface alterations following treatment in vitro with the deacetylated (amine) metabolite of diamphenethide.

The effect of the deacetylated (amine) metabolite of diamphenethide (10 micrograms/ml) on the tegumental surface of Fasciola hepatica over a 24 h period in vitro has been determined by scanning electron microscopy. Blebbing begins around the oral sucker after 3 h and then passes backwards along the body, reaching the ventral sucker and midbody by 6 h, and finally the posterior end of the body (by 12 h). Initially, the blebs are small, the tegument surrounding the spines is swollen and the tegument generally has a smooth, swollen appearance. This submerges the spines below the body surface. At higher magnification the surface is seen to bear microvillous-like projections in addition to the blebs and surface pitting is deeper than normal. Later on, the blebs increase in size and burst, causing lesions and loss of spines. Lesions begin to appear on the oral cone and ventral sucker after 6 h, in the midbody by 12 h and on the dorsal surface of the posterior region after 24 h. By this time the damage is extensive: around the oral and ventral suckers, and over large areas of the oral cone and midbody region the tegument has been stripped off to expose the basal lamina beneath. The dorsal surface of the fluke is consistently more severely affected than the ventral surface.

Acetanilides↗

PubMed1978

Chaetogaster limnaei K von Baer 1872 on Lymnaea tomentosa: ingestion of Fasciola hepatica cercariae.

The fresh water mollusc, Lymnaea tomentosa, the intermediate host of the liver fluke is heavily infested under natural conditions with ectocommensal annelid, Chaetogaster limnaei. These annelids which destroy the larval stages of Fasciola hepatica have been observed in the laboratory. The association of C. limnaei with the snail intermediate host is of value in exploiting a control measure against economically important liver fluke disease.

Animals↗

PubMed1994

Vaccination of sheep against Fasciola hepatica with glutathione S-transferase. Identification and mapping of antibody epitopes on a three-dimensional model of the antigen.

The glutathione S-transferases (FhGST) of the liver fluke Fasciola hepatica have been identified as novel vaccine candidates that protect sheep against a fluke infection. With the use of overlapping peptides covering the predicted amino acid sequences of four FhGST cDNAs, we have defined the linear epitopes recognized by polyclonal antibody from sheep vaccinated with FhGST. Dominant and minor epitopes were found to be present on all four of the sequences although some epitopes were shown to be specific to particular FhGST. A high percentage of the FhGST peptides were found to be antigenic although considerable variability in response to the peptides was observed among the animals. This analysis was extended to the IgG1 and IgG2 response at the peptide level. Based on the recently solved crystal structure of the rat mu-class GST 3-3, a three-dimensional model of one of the FhGST sequences was generated that allowed the predicted spatial localization of defined epitopes. Most epitopes were localized on regions of high flexibility and accessibility. A comparison of epitopes on FhGST with the B cell epitopes on Sm28, a 28-kDa GST from Schistosoma mansoni, has found few similarities. There was no correlation between an antibody response to linear peptide epitopes and the level of protection induced in sheep by vaccination with FhGST.

Amino Acid Sequence↗

PubMed2000

Detection of Fasciola hepatica infection in a community located in the Ecuadorian Andes.

The presence of fascioliasis was assessed in four Andean communities using an enzyme-linked immunosorbent assay (ELISA) test to measure antibodies against Fasciola hepatica excretion-secretion antigens. Six percent (9 out of 150) of the individuals in one community were ELISA-positive for these antibodies. Fecal samples from two of the ELISA-positive individuals contained F. hepatica ova. All of the ELISA-positive cases, except for one, were children within the ages of 9 to 12 years.

Adult↗

PubMedDate not supplied

[Neurologic manifestations of hepatic distomiasis caused by Fasciola hepatica].

The hepatic distomiasis is a parasitic and cosmopolite affection caused by a trematode Fasciola hepatica. The clinical manifestation usually includes an invasion phase with toxi-infectious hepatitis and intermittent angiocholitis episodes during the chronic phase of the disease. Many authors insist for few years, on some revealing manifestations of the disease that are unusual in the classical forms, especially the neurological manifestations that are rare and of extreme polymorphism. Based on a 12 years old tunisian child case of distomiasis caused by F. hepatica associated with neurological manifestations and through the analysis of 25 literature cases, the symptomatological and physiopathological aspects of this clinical form that generally makes the physician perplex, are reviewed.

Child↗

PubMed1981

Fasciola hepatica: a technique for the study of gut penetration by juvenile flukes.

A method using light and electron microscopes is described which is suitable for the examination of gut penetration by juvenile Fasciola hepatica. It involved the ligation of small sections of the small intestine of rats and the introduction of artificially excysted flukes into these gut loops. By restricting the area of infection in this way it was possible to either recover flukes from the gut lumen or to prepare ultrathin sections for electron microscopy of flukes penetrating the gut wall. In addition, flukes were recovered from the body cavity at various times after preparation of loops in resistant and naive rats. It was found that more flukes reached the body cavity in naive rats than in resistant rats, demonstrating a resistance to infection in the gut loops of sensitized rats.

Animals↗

PubMed2000

DNA sequence polymorphisms in two strains of Fasciola hepatica from Egypt and Europe.

A new PCR based system was used that had broad detection capability among parasites based on a conserved region of the 18 S ribosomal DNA gene. Five samples each of European and Egyptian Fasciola hepatica of bovine origin were obtained and DNA was isolated. The target of the PCR primers was a fragment of approximately 600 nucleotides in length corresponding to a region of the 18 S rRNA gene. Sequences were compared over a 107 base pair region that identified polymorphism between the strains. All five Egyptian isolates were identical. Likewise all of the European isolates had identical sequence. There were polymorphisms between the two strains and also with F. hepatica isolates from North America. Both the European and Egyptian isolates have a single base addition in the polymorphic region. In addition there is a single base substitution in the Egyptian isolates when compared to the others. This region is a small target that can be used to identify the origins of different F. hepatica isolates.

Animals↗

PubMed1986

The effect of albendazole and triclabendazole on colchicine binding in the liver fluke Fasciola hepatica.

Albendazole (ABZ) and its sulfoxide (SX) and sulfone (SO) metabolites inhibit the binding of 3H-colchicine, a ligand with high affinity for tubulin to homogenate preparations of the liver fluke Fasciola hepatica. The relative potency of these compounds is SX greater than ABZ greater than SO. The benzimidazoles (cambendazole, parbendazole, oxibendazole and mebendazole), when tested at a concentration of 10 microM, also inhibited colchicine binding to fluke homogenates. However, a potent new benzimidazole flukacide, triclabendazole (TCB), was without effect on colchicine binding to F. hepatica homogenates. When intact flukes were exposed in vitro to 10(-5)M SX for as little as 5 min the subsequent binding of 3H-colchicine to fluke homogenates was significantly reduced. However, flukes recovered from sheep either 12 or 24 h after treatment with ABZ did not have a decreased ability to bind colchicine, although the non-specific binding was higher in flukes from treated sheep, suggesting some interaction of drug with tubulin in vivo. ABZ, SX and SO were effective in preventing embryonation of fluke eggs at doses as low as 0.01 microM, but TCB was without effect at concentrations as high as 10 microM. The results suggest that ABZ exerts at least part of its anthelmintic effect by interaction with fluke tubulin.

Albendazole↗

PubMed1983

Fasciola hepatica: a technique for monitoring in vitro motility.

An apparatus utilizing a force and displacement transducer is described for the direct and long-term recording of the motility in vitro of Fasciola hepatica. Normal movement is typically rhythmical, with bursts of more powerful contractions alternating with periods of lesser activity. Such rhythms and the overall level of activity are maintained for more than 30 hr. The fluke remains active for much longer periods of time: recordings of fluke movements have been made for up to 4 days. Potential damage to the fluke caused by the attachment system within the recording apparatus has been determined by the Evans' Blue Technique and scanning electron microscopy. It is restricted to the attachment sites, and does not spread to other parts of the body over the 30-hr normal activity period. Transmission electron microscope studies have shown that the tegument retains its structural and functional integrity over this period of time. There are advantages of the recording apparatus over previous kymographic methods for studying fluke motility.

Animals↗