Disruption of egg formation by Fasciola hepatica following treatment in vivo with triclabendazole in the sheep host

Eight indoor-reared cross-bred sheep with no prior exposure to Fasciola hepatica were infected by oral gavage with 200 metacercarial cysts of the triclabendazole (TCBZ)-susceptible Cullompton isolate of F. hepatica. Twelve weeks after infection, sheep were treated with 10 mg/kg triclabendazole. Two sheep were euthanised per time period; at 48 h, 72 h and 96 h post-treatment (pt). Two untreated control sheep were euthanised at 96 h pt. Flukes were recovered from the liver and, if present, from the gall bladder of the sheep. They were processed for whole mount analysis, histology and transmission electron microscopy of the female reproductive system; specifically, the uterus, vitelline follicles. Mehlis' gland and ovary.

Comparison of two assays, a faecal egg count reduction test (FECRT) and a coproantigen reduction test (CRT), for the diagnosis of resistance to triclabendazole in Fasciola hepatica in sheep

A sheep trial was performed to evaluate two diagnostic assays, a faecal egg count reduction test (FECRT) and a coproantigen reduction test (CRT), for the diagnosis of resistance of Fasciola hepatica to triclabendazole (TCBZ). The FECRT defines successful TCBZ treatment as a 95% or greater reduction in fluke faecal egg counts (FECs) at 14 days post-treatment (dpt). The CRT defines effective TCBZ treatment as faeces negative for Fasciola coproantigens at 14 dpt, as measured by the commercial BIO K201 coproantigen ELISA (Bio-X Diagnostics, Jemelle, Belgium).

Development of an egg hatch assay for the diagnosis of triclabendazole resistance in Fasciola hepatica: proof of concept.

The aim of this study was to develop an Egg Hatch Assay (EHA) test for the detection of triclabendazole (TCBZ) resistance in Fasciola hepatica. A number of fluke isolates were used, of differing sensitivity to TCBZ. Eggs were exposed to solutions of triclabendazole sulphoxide (TCBZ.SO) for 14 days, then triggered to hatch. Egg development was divided into 6 distinct and easily identifiable stages: dead, empty, unembryonated, cell division, eye spot and hatched. The number of eggs reaching those stages was recorded. Initially, the discriminating dose (1% hatch) was determined for the Cullompton isolate, used as TCBZ-susceptible (TCBZ-S) standard. Once this concentration had been resolved, the response of different isolates to this concentration was examined. The hatch rate of the Fairhurst isolate was not significantly different from that of the Cullompton isolate, confirming its TCBZ-S status. The Patagonia isolate has not been exposed to TCBZ in the field and should be TCBZ-S: the results of the EHA supported this. The egg hatch response of the Oberon and Dutch isolates differed significantly from that of the Cullompton isolate; the former isolates are regarded as TCBZ-resistant (TCBZ-R) and the results confirmed this. Another isolate, the Leon isolate, was originally described as being TCBZ-R, but has since been shown to be TCBZ-S. There was no difference in its response to TCBZ.SO in the EHA from the Cullompton (and Fairhurst and Patagonia) isolate(s), further indicating its TCBZ-S status. The impact of TCBZ.SO treatment on the component stages of egg development was determined and revealed differences between the isolates. In conclusion, the results of the study have shown that it is possible to discriminate between TCBZ-S and TCBZ-R isolates of F. hepatica on the basis of the response of their eggs to an EHA and the test could be used to evaluate the TCBZ sensitivity of unknown field isolates

THE ULTRASTRUCTURE AND IMMUNOGOLD LABELING OF PANCREATIC POLYPEPTIDE-IMMUNOREACTIVE CELLS ASSOCIATED WITH THE EGG-FORMING APPARATUS OF A MONOGENEAN PARASITE, DICLIDOPHORA-MERLANGI

An electron microscopical examination has been made of the fine structure and disposition of pancreatic polypeptide immunoreactive cells associated with the egg-forming apparatus in Diclidophora merlangi. The cell bodies are positioned in the parenchyma surrounding the ootype and taper to axon-like processes that extend to the ootype wall. The terminal regions of these processes branch and anastomose and, in places, the swollen endings or varicosities form synaptic appositions with the muscle fibres in the ootype wall. The cells are characterized by an extensive GER-Golgi system that is involved in the assembly and packaging of dense-cored vesicles. The vesicles accumulate in the axons and terminal varicosities, and their contents were found to be immunoreactive with antisera raised to the C-terminal hexapeptide amide of pancreatic polypeptide. It is concluded that the cells are neurosecretory in appearance and that, functionally, their secretions may serve to regulate ootype motility and thereby help co-ordinate egg production in the worm.

Low-dose hypersensitivity in Chinese hamster V79 cells targeted with counted protons using a charged-particle microbeam

The Gray Laboratory charged-particle microbeam has been used to assess the clonogenic ability of Chinese hamster V79 cells after irradiation of their nuclei with a precisely defined number of protons with energies of 1.0 and 3.2 MeV. The microbeam uses a 1-mum. silica capillary collimator to deliver protons to subcellular targets with high accuracy. The detection system is based on a miniature photomultiplier tube positioned above the cell dish, which detects the photons generated by the passage of the charged particles through an 18-mum-thick scintillator placed below the cells. With this system, a detection efficiency of greater than 99% is achieved. The cells are plated on specially designed dishes (3-mum-thick Mylar base), and the nuclei are identified by fluorescence microscopy. After an incubation period of 3 days, the cells are revisited individually to assess the formation of colonies from the surviving cells. For each energy investigated, the survival curve obtained for the microbeam shows a significant deviation below I Gy from a response extrapolated using the LQ model for the survival data above 1 Gy. The data are well fitted by a model that supports the hypothesis that radioresistance is induced by low-dose hypersensitivity. These studies demonstrate the potential of the microbeam for performing studies of the effects of single charged particles on cells in vitro. The hypersensitive responses observed are comparable with those reported by others using different radiations and techniques. (C) 2001 by Radiation Research Society.

Modulation of the motility of the vagina vera of Ascaris suum in vitro by FMRFamide-related peptides

Ascaris suum contains a large number of FMRFamide-related peptides (FaRPs) of which KNEFIRFamide (AF1), KHEYLRFamide (AF2) and KSAYMRFamide (AF8, also called PF3) have been extensively studied and are known to exert actions on somatic muscle strips of the worm. In the present study, the effects of AF1, AF2 and AF8 on the activity of the vagina vera of female A. suum have been examined in vitro. The vagina vera is a muscular tube connecting the uterus and vagina uteri to the gonopore and is probably involved in regulating egg output. The tissue exhibited spontaneous, rhythmic contractions in vitro, which were modulated by each of the FaRPs tested. The effects of each of the peptides were qualitatively and quantitatively different, and in each case were reversible. AF1 (1 mu M) caused a biphasic response in the form of a transient lengthening of the preparation, followed by a shortening; contractions were initially inhibited but resumed 5 min post-addition of the peptide. Lower concentrations (less than or equal to 0.1 mu M) induced a less marked effect, with rhythmic contractions returning 5 min post-addition. AF2 and AF8 reduced contraction frequency at concentrations greater than or equal to 0.1 mu M. Both peptides also caused the tissue to shorten, although the effects of AF8 on baseline tension were inconsistent. The apparent potencies of AF1 and AF8 on contraction frequency of the vagina vera were 10-fold greater than AF2 and, unlike their actions on A. suum somatic body wall muscles, the actions of AF1 and AF2 were qualitatively different. Indeed, the effects of each of these FaRPs on the vagina vera were markedly different from those observed on the somatic muscle.

Kinetics of semicarbazide and nitrofurazone in chicken eggs and egg powders

The accumulation, depletion and partitioning of semicarbazide (SEM) and its parent compound nitrofurazone (NFZ) in eggs were studied using hens fed NFZ at therapeutic and sub-therapeutic levels. Dietary NFZ correlated strongly with NFZ and total SEM in eggs, while 28% of observed SEM was present in the form of parent NFZ. Depletion half-life in eggs was 2.4 days for SEM and 1.1 days for NFZ. NFZ accumulated preferentially in yolk (57-63%) as opposed to albumen, while 71-80% of SEM was found in yolk. In whole egg, 29% of SEM was present as tissue-bound residues compared with 80% in breast muscle. Whilst NFZ and SEM were partly degraded by pasteurization and spray drying, sufficient NFZ remained to suggest it might be detectable in egg powders when SEM is observed at low µg kg -1 concentrations. NFZ was detectable in whole eggs during ingestion of only 0.1% of the therapeutic NFZ dose, making detection of intact NFZ in eggs a feasible means to prove conclusively the administration of this banned compound.