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PubMed2000

Uptake of albendazole and albendazole sulphoxide by Haemonchus contortus and Fasciola hepatica in sheep.

The pattern of in vivo uptake of albendazole (ABZ) and its major metabolite, ABZ-sulphoxide (ABZSO), by Haemonchus contortus and Fasciola hepatica recovered from ABZ-treated sheep, was investigated. Concentration profiles of both compounds were simultaneously measured in target tissues/fluids from the same infected sheep. In addition, the proportion of the (+) and (-) ABZSO enantiomers was determined in plasma, bile and F. hepatica recovered from treated sheep. Sheep naturally infected with H. contortus were intraruminally (i.r.) treated with ABZ (micronized suspension, 7. 5mg/kg) and the plasma concentrations of ABZSO and ABZ-sulphone (ABZSO(2)) determined in addition to the concentration of ABZ and ABZSO in H. contortus, abomasal mucosa and fluid content samples. In addition, F. hepatica artificially infected sheep were treated i.r. with the same ABZ suspension (7.5mg/kg), and samples of blood, bile, liver tissue and adult flukes were collected and analysed by HPLC to determine the concentrations of ABZ and both enantiomers of ABZSO. ABZSO and ABZSO(2) were the analytes recovered in plasma with ABZ and ABZSO present in H. contortus. ABZ was the analyte recovered at the highest concentration in H. contortus and abomasal mucosa, whereas higher concentrations of ABZSO were measured in abomasal fluid content. Only low concentrations of ABZ were detected in F. hepatica and bile, but markedly higher concentrations of ABZ were measured in liver tissue. ABZSO was the main molecule recovered in F. hepatica, plasma and bile samples collected from ABZ-treated sheep. The (+) enantiomer of ABZSO was recovered at a higher proportion in plasma (75%), bile (78%) and F. hepatica (74%) after ABZ administration to infected sheep.

Abomasum↗

PubMed2005

Prevalence of Fasciola hepatica in dairy herds in England and Wales measured with an ELISA applied to bulk-tank milk.

An ELISA developed to diagnose Fasciola hepatica infection in cattle by detecting serum antibodies was adapted and validated for use with samples of bulk-tank milk. The prevalence of the infection in 61 dairy herds was established by using serum antibody levels or faecal egg counts measured in a proportion of the cows in each herd. The correlation between the results of the ELISA and the herd seroprevalence was 0.83. Using a cut-off value of 27 per cent positive, the bulk-tank ELISA identified herds in which more than 25 per cent of the cows were infected with a diagnostic sensitivity of 96 per cent (95 per cent confidence interval 89 to 100 per cent) and a diagnostic specificity of 80 per cent (95 per cent confidence interval 66 to 94 per cent). By applying the ELISA to 623 herds in England and 445 herds in Wales, the prevalence of F hepatica infection in England was estimated to be 48 per cent (95 per cent confidence interval 46 to 54 per cent), and in Wales 86 per cent (95 per cent confidence interval 84 to 90 per cent).

Animals↗

PubMed2002

Aspects of the maintenance of the life cycle of Fasciola hepatica in Lymnaea columella in Minas Gerais, Brazil.

Fascioliasis is a parasitic disease of domestic ruminants that occurs worldwide. The lymnaeid intermediate hosts of Fasciola hepatica include Lymnaea columella, which is widely distributed in Brazil. A colony of L. columella from Belo Horizonte, MG, was reared in our laboratory to be used in studies of the F. hepatica life cycle, the intermediate host-parasite relationship and development of an anti-helminthic vaccine. In the first experiment 1,180 snails were exposed to miracidia of F. hepatica eggs removed from the biliary tracts of cattle from the State of Rio Grande do Sul. In the second and third experiments the snails were exposed to miracidia that had emerged from F. hepatica eggs from Uruguay, maintained in rabbits. The rates of infection in the first, second and third experiments were 0, 42.1 and 0% respectively. Over 15,806 metacercariae were obtained and stored at 4 degrees C. Four rabbits weighing 1.5 kg each were infected with 32-44 metacercariae and two with 200. Three rabbits begin to eliminate eggs of the parasite in the feces from 84 days after infection onwards. The biological cycle of F. hepatica in L. columella and the rabbit was completed within 124 days.

Animals↗

PubMed1988

Fasciola hepatica: perturbation of secretory activity in the vitelline cells by the sodium ionophore monensin.

The effect of the sodium ionophore monensin on the vitelline cells of Fasciola hepatica has been determined both in vitro and in vivo by means of transmission electron microscopy. In intact flukes in vitro, vacuolation of the Golgi complex of the intermediate, shell protein secreting vitelline cells is evident after 1.5 hr incubation in monensin (1 X 10(-6) M). The vacuolation becomes progressively greater with time, eventually spreading to the late stem cells and mature cells. In addition, there is a block in the normal migration of the shell protein globules to the periphery, the shell globule clusters becoming very loosely packed and empty, and distended single globules accumulate in the perinuclear region of the cell. Disruption of the nurse cell cytoplasm is apparent from 6 hr onwards, giving the follicle a less compact appearance. Morphological changes induced by higher concentrations of monensin (up to 1 X 10(-4) M) followed a similar time course and pattern to that described for 1 X 10(-6) M) followed a similar time course and pattern to that described for 1 X 10(-6) M. In tissue-slice material (1 X 10(-6) M) these effects of monensin are evident more rapidly, and to a far greater extent: the condition of the vitelline cells in slices after only 1.5 hr resembles that reached in intact flukes after more than a 12-hr incubation. Incubation in ouabain, an inhibitor of Na+/K+-ATPase activity, has little effect on vitelline morphology over a 6-hr period (1 X 10(-3) M), although brief (0.5 hr) exposure to ouabain followed by monensin treatment (1 X 10(-4) M, 3 hr) does lead to gross vacuolation of the intermediate cells, the condition resembling that in tissue-slice material. In contrast, in vivo treatment of infected laboratory rats (1 X 5 mg/kg) only leads to a transient effect on the ultrastructure of the intermediate vitelline and nurse cells. The specific perturbation of the Golgi complex and secretory traffic in the vitelline cells of F. hepatica by monensin follows the classic pattern observed in other cell types.

Animals↗

PubMed2001

Ex vivo diffusion of albendazole and its sulfoxide metabolite into ascaris suum and Fasciola hepatica.

The current experiments compare the pattern of ex vivo uptake (diffusion) of albendazole (ABZ) and albendazole sulfoxide (ABZSO) by Ascaris suum and Fasciola hepatica. Specimens of A. suum and F. hepatica were collected from untreated animals (pigs and sheep, respectively) and incubated with either ABZ or ABZSO for different time periods (5-180 min). After incubation. the parasite material was analysed by HPLC to quantify the amount of ABZ and/or ABZSO. The parent drug and its active ABZSO metabolite were recovered from the parasites after ex vivo incubation for different time periods throughout the assay. Total drug availability in A. suum, expressed as area under the concentration versus time curve (AUC) over 180 min of incubation, was significantly greater (P<0.05) for ABZ parent drug (AUC = 4.19 +/- 0.59 microg x h xg(-1)) compared with the more polar ABZSO metabolite (AUC = 0.25 +/- 0.01 micro x h x g(-1)). Similar results were observed after the incubation of both molecules with F. hepatica, where the AUC values obtained were 10.6 +/- 0.28 microg x h x g(-1) and 2.04 +/- 0.33 microg x h x g(-1) for ABZ and ABZSO, respectively. The greater diffusion and availability of ABZ in both helminths correlate with the higher lipophilicity of the parent drug, compared with its sulfoxide metabolite. The amount of both molecules measured in A. suum was significantly lower (P<0.05) than that recovered in F. hepatica. The complexity of the histological structure of the nematode cuticle compared with the external tegument of the trematode may account for such a difference in drug diffusion between the species. These findings complement previous observations on the patterns of in vivo uptake of ABZ by different helminth parasites, contributing to the understanding of the pharmacological anthelmintic action of these moieties.

Albendazole↗

PubMed1998

Evaluation of a diagnostic monoclonal antibody-based capture enzyme-linked immunosorbent assay for detection of a 26- to 28-kd Fasciola hepatica coproantigen in cattle.

OBJECTIVE: To develop a monoclonal antibody-based capture ELISA for detection of a 26- to 28-kd coproantigen of Fasciola hepatica in the feces of infected cattle. ANIMALS: 27 crossbred yearling calves, 2 New Zealand White rabbits. PROCEDURE: A capture ELISA that uses a previously described monoclonal antibody (MAB) M2D5/D5F10 was developed. The MAB was used to capture the antigen from the feces, and hyperimmune rabbit serum raised against the purified 26- to 28-kd glycoprotein was used to detect the coproantigen. This test was used for the detection of the antigen in the feces of 27 experimentally infected calves with known numbers of flukes. Fecal specimens obtained before infection from the same calves were used as negative controls. RESULTS: The assay results identified all calves infected with more than 10 flukes at necropsy, and as little as 300 pg of coproantigen/ml of fecal supernatant was detected. The assay results correlated well with the number of flukes, suggesting that it is possible to estimate fluke burden. Infections as early as 6 weeks duration were detected, before flukes mature to adults and start to shed eggs. CONCLUSIONS: In experimentally infected calves, the coproantigen capture ELISA was more sensitive and easier to perform than microscopic examination for the diagnosis of F hepatica infection; moreover, 6-week-old prepatent infections were detectable. CLINICAL RELEVANCE: This capture ELISA containing an F hepatica 26- to 28-kd coproantigen is a quantitative assay that is more sensitive than fecal egg counting. In addition, the assay is rapid, easy to perform and lends itself well to large numbers of samples. Because it is antigen based, the ELISA may be useful for diagnosis of F hepatica infection in other species, including human beings.

Animals↗

PubMed2005

Molecular and electrophysiological characterization of nucleotide-sensitive chloride current-inducing protein of Fasciola hepatica.

Nucleotide-sensitive chloride current regulating proteins (ICln's) of the chloride channels have been characterized from man and animals. An ICln of Fasciola hepatica (ICln-Fh) consisting of 231 amino acids revealed high similarities to both consensus domain of ICln's and two acidic residue-abundant patches in its C-terminus. Native ICln-Fh protein was confirmed present in F. hepatica soluble extract by immunoblotting. The recombinant ICln-Fh protein expressed in collagenase-defolliculated Xenopus oocytes induced fast rising and outward rectifying Cl- currents (I(Cln-Fh)). The recombinant ICln-Fh protein, however, did not trigger cell swelling-induced Cl- currents (I(Cl-swell)). The I(Cln-Fh) currents were significantly reduced by substituting external Cl- with gluconic acid and by externally adding cAMP. Collectively, these results suggest that ICln-Fh protein is an inducer of Cl- currents in F. hepatica.

Amino Acid Sequence↗

PubMed1995

Seroprevalence of Fasciola hepatica infection in sheep in northwestern Spain.

To estimate the prevalence of Fasciola hepatica infection in sheep in the León province (northwestern Spain), we conducted a survey between October 1992 and May 1993. A total of 767 samples of feces and serum were collected from sheep over 1 year of age belonging to 152 flocks randomly selected from the 4 natural regions of León province. Samples were analyzed by a standard coprological sedimentation method and an indirect enzyme-linked immunosorbent assay (ELISA) using excretory-secretory products from F. hepatica as the antigen. The results showed the feasibility of using the indirect ELISA to facilitate the serodiagnosis of ovine fasciolosis in seroepidemiology studies (95% sensitivity and > 99% specificity). No serological cross-reaction with infection by the trematode Dicrocoelium dendriticum was found. Furthermore, a statistically significant association was demonstrated between the mean flock prevalence results as determined by ELISA (77.6%) and by coprological examination (23.7%; P < 0.001). Differences in the results obtained by the two diagnostic methods could have been due to fluctuations in the numbers of fluke eggs detected in feces and to the persistence of specific antibodies in serum after an efficacious fasciolicide treatment. The median number of F. hepatica eggs detected per gram of feces was 10 (range, 5-450 eggs/g feces). The geographical distribution of F. hepatica infection in León province was similar in all natural regions, probably due to the observation that meteorological conditions are not limiting for the maintenance of the parasite life cycle in any area of the province and to the abundance of irrigated areas together with the lack of planned control strategies.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed2002

Performance characteristics of an enzyme-linked immunosorbent assay for the detection of liver fluke (Fasciola hepatica) infection in sheep and cattle.

AIM: To validate an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against liver fluke (Fasciola hepatica) in sheep and cattle sera. METHODS: Gold-standard sera from sheep and cattle of known infection status, i.e. sera from non-infected animals and from animals known to be infected with F. hepatica were assayed with a commercially available ELISA and results analysed by ROC analysis. RESULTS: The ROC analysis suggested cut-offs that were considerably lower than those suggested by the manufacturer, yet the ELISA performed with high sensitivity and specificity, 98 to 100%, respectively for sheep and cattle sera. For bovine sera, particularly good discrimination between positive and negative sera was observed. Infection in experimentally infested animals could be demonstrated 7-8 weeks earlier than with classical parasitological techniques. CONCLUSIONS: The analysis of the ELISA's performance demonstrated high sensitivity and specificity. ROC analyses optimised the cut-off point suggested by the manufacturer of the commercial diagnostic assay. Diagnosis of infection with F. hepatica was achieved much earlier than is possible with current parasitological techniques. This could help with the control of fasciolosis, enabling treatment before clinical manifestation of the disease.

Animals↗

PubMed1979

The effect of route of infection on acquired resistance to Fasciola hepatica in the rat and sheep.

Rats and sheep were infected with Fasciola hepatica by subcutaneous, intramuscular or oral (intraruminal--sheep) inoculation with encysted metacercariae and the development of flukes from these primary infections assessed. Additional rats and sheep similarly infected were challenged with encysted metacercariae orally (rats), or intraruminally (sheep) to assess the level of acquired resistance generated by previous infection by each of the three routes of administration. In both rats and sheep 0--5 per cent, 0--5 per cent and 20--25 per cent of metacercariae given by the subcutaneous, intramuscular or oral (intraruminal--sheep) routes were recovered as flukes in the liver four and 12 weeks after infection of rats, or 17 weeks after infection of sheep. Primary infection of rats with F hepatica by each of the three routes generated strong protection against oral challenge whereas no protection was generated in sheep, irrespective of route of infection.

Administration, Oral↗

PubMed1981

Successful passive transfer of resistance to Fasciola hepatica infection in rats by immune serum and transfer factor.

Resistance to Fasciola hepatica was shown by a 64 per cent reduction in fluke recovery compared with control rats was recorded in rats inoculated with serum from sheep experimentally infected with F hepatica. A comparable reduction (55.6 per cent) was obtained using transfer factor prepared from lysed leucocytes of infected rats. No significant reduction was obtained using transfer factor prepared from sheep or calves.

Animals↗

PubMed1990

Eosinophil responses to Fasciola hepatica in rodents.

Qualitative and quantitative cellular changes in the peripheral blood and bone marrow of resistant (rat) and susceptible (mouse) hosts of Fasciola hepatica have been examined. Eosinophil numbers in the peripheral blood and bone marrow of both hosts increased almost immediately following infection. Rats responded more rapidly than mice. Bone marrow colony formation in both rats and mice was greatly enhanced following F. hepatica infection. Injection of excretory/secretory (E/S) antigens of the fluke into rats and mice caused peripheral eosinophilia. Eosinophil levels in mice dropped by day 7 post-injection, but those in rats remained high. Eosinophil precursors in the bone marrow of injected animals also rose. Bone marrow colony formation in antigen-injected mice peaked sharply at day 7 but then fell rapidly. Rats injected with E/S antigens had about twice the level of bone marrow colonies as controls, 12 days post-injection. For most parameters measured, the magnitude of the responses of rats was greater than mice, which may be significant in the context of the rat's ability to acquire resistance to reinfection.

Animals↗

PubMed1982

Diagnosis of Fasciola hepatica infections in sheep by an enzyme-linked immunosorbent assay.

An enzyme-linked immunosorbent assay (ELISA) was evaluated for diagnosis of Fasciola hepatica infections in sheep maintained under isolation and drylot conditions. Sheep were exposed to various numbers (100 to 500) of metacercariae, and blood was collected for assay at regular intervals thereafter. The ELISA values of controls were low and varied little. Although the ELISA detected antibodies to the flukes as early as postinoculation week (PIW) 2, ELISA values suggested as diagnostic (1.75 times control values) did not occur until PIW 6 to 8. Positive diagnosis of the infections by fecal examination for fluke eggs was not established until after PIW 10. Results also indicated that ELISA values were generally not correlated with magnitude of infection, concurrent nematode infections, or husbandry practices.

Animals↗

PubMed1980

Fasciola hepatica: influence of thymus function on the course of infection in mice.

The mouse has been shown to be a suitable experimental host in studies of the resistance to Fasciola hepatica infection. The present experiment compares the course of infection in two different strains of mice, including immunodeficient (congenitally athymic (nude)--(nu/nu)) mice. It was found that the infection was lethal in nu/nu mice, whereas nu/nu mice reconstituted with a neonatal thymus graft reacted as intact mice. The white blood cell picture of nu/nu mice remained unchanged during the infection and between day 24 and 30 after inoculation nearly all nu/nu mice died. Thymus-bearing mice showed blood leucocyte increase and neutrophilia from week 3 after primary inoculation. Furthermore, a marked peritoneal cell reaction occurred 2--4 weeks after primary inoculation. The total number of cells in peritoneal cavity increased approx. 10--15 times and a very remarkable eosinophil peak was seen in week 3 after primary inoculation. In nu/nu mice the peritoneal cell increase was approx. twice that of uninoculated mice and no eosinophil response was observed. When results obtained with the various groups of mice are compared, it appears that resistance to F. hepatica during infection is dependent on a functional thymus.

Animals↗

PubMed2003

Efficacy of an experimental fasciolicide against immature and mature Fasciola hepatica in artificially infected calves.

The efficacy of 5-chloro-2-methylthio-6-(1-naphthyloxy)- 1H-benzimidazole, called "alpha", was tested against Fasciola hepatica. Fluke-free calves ( n=32) were divided into 8 groups and infected with 150 metacercariae per animal. All animals subsequently received a second infection with another 150 metacercariae, given at different time intervals aimed to produce flukes of differing ages within the experimental animals. When the flukes reached the required age in the animals, four groups were treated with a single oral dose of 12 mg/kg of compound alpha and the remaining ones served as non-treated controls. Two weeks after treatment the animals of all groups were sacrificed and the livers were removed to determine the numbers of parasites present in the treated and untreated controls. In the treated groups the fluke reduction for the 3 day/2 week group was 100%, for the 3 week/4 week group it was 96.4%, for the 6 week/8 week group it was 99.2% and for the 10 week/12 week group it was 100%.

Animals↗

PubMed1995

Fasciola hepatica cathepsin L proteinase cleaves fibrinogen and produces a novel type of fibrin clot.

A cathepsin L proteinase secreted by the parasitic helminth Fasciola hepatica can cleave fibrinogen and produce a fibrin clot with a specific activity of 4.7 National Institutes of Health thrombin-equivalent U/mg. This is the first report of a fibrinogen-clotting activity aside that of thrombin and the snake venom proteinases, which are all serine proteinases. Clot formation by cathepsin L is not inhibited by the thrombin inhibitor hirudin or by the anti-polymerant H-Gly-Pro-Arg-Pro-OH. The enzyme exerts its activity on fibrinogen in a unique manner. Although the cleavage of fibrinogen may involve the initial removal of fibrinopeptides, additional proteolysis of the alpha, beta and gamma fibrinogen polypeptides takes place. SDS/PAGE analysis of the cathepsin-L-produced clots revealed that cleavage of the alpha polypeptide (66 kDa) precedes that of the beta (52 kDa) and gamma (46.5 kDa) polypeptides. Concurrent with the cleavage of these polypeptides is the appearance of components of 120, 100 and 25 kDa. The appearance of higher molecular-sized components in the cathepsin L clots suggests that polymerisation involves the formation of molecular interactions that are resistant to boiling in mercaptoethanol and SDS.

Animals↗

PubMed1977

The penetration of Fasciola hepatica miracidia into the snail host Fossaria bulimoides. A scanning electron microscope study.

Snails of the species Fossaria bullimoides were exposed to miracidia of Fasciola hepatica for given periods of time. The course of penetration was followed by means of scanning electron microscopy. Topographical features of miracidial morphology were studied with emphasis on epidermal plates and body shape. Comparisons are made on the rate of penetration into this snail host.

Animals↗

PubMed2002

Immunization with cathepsin L proteinases CL1 and CL2 secreted by Fasciola hepatica elicit a preferential type 1 response based on IgG2a antibodies in rats.

Cathepsin L proteinases (CL1 and CL2), the major components of Fasciola hepatica excretion/secretion products (E/S) are considered potential antigens of a vaccine against fascioliasis. The humoral response elicited by CL1 and CL2 in rats either immunized with the enzymes or infected with F. hepatica has been analysed, examining specific IgE and IgG subclass dynamics. The experiment was continued for 10 weeks and peripheral blood eosinophilia was also determined. Infected rats presented peaks of eosinophilia at weeks 3 and 7 post-infection, while those immunized with CL1 and CL2 were no different from controls. Total IgE in infected rats increased up to week 5, reaching 30 microg(-1) in some cases, then decreased slowly and rising again towards the end of the experiment. Determination of specific IgE, carried out in sera previously absorbed with Protein G-Sepharose, reached a peak in infected rats between weeks 2 and 5, depending on the individual. In immunized rats both total and specific IgE levels remained around the pre-immunization values. With regard to the IgG subclass responses to E/S products, in infected rats IgG1 predominated over IgG2a, and the reverse was true in rats immunized with CL1 and CL2 and tested against the respective antigens. In all cases an increase in IgG1 and IgG2a antibody titres was seen, with maximum levels being reached later (weeks 6-7) in infected rats than in immunized ones (weeks 4-5). No IgG2b or IgG2c responses were detected in any of the groups studied.

Animals↗