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  • 1
    ISSN: 1365-2761
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Abstract. Various antiparasitic drugs belonging to several pharmacological groups were tested by bath administration for in vivo activity against a natural infestation of rainbow trout. Oncorhynchus mykiss (Walbaum), by he flagellate protozoan Ichthyobodo necator. Fish were also monitored for signs of drug toxicity. Complete elimination of infestation in all fish was achieved only by bithionol (25 mgl-1 for 3h on two consecutive days). Ketoconazole, toltrazuril, amprolium, sulphaquinoxaline, quinacrine, N-metylglucamine, chloroquine, 1,3-di-6-quinolylurea, diminazene aceturate and paromomycin were not effective. Diminazene aceturate (100 mgl-1, 3h) was the only drug which was clearly toxic. The comparison of the response of symptomatic and asymptomatic infested fish to a formaldehyde bath indicates that poor health caused by I. necator may greatly increase the susceptibility of fish to the toxic effects of a drug or chemical.
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  • 2
    ISSN: 1365-2761
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Abstract. The humoral immune response of turbot, Scophthalmus maximus (L.), to antigens from the microsporean parasite Tetramicra brevifilum Matthews & Matthews, 1980, was studied. Thirty days after intraperitoneal immunization with whole T. brevifilum spores in Freund's complete adjuvant, double indirect ELISA indicated that initial production of antibodies to parasite surface antigens was considerably higher than production of antibodies to the antigens contained in a crude extract (CE) of spores. Following re-immunization without adjuvant on day 30, levels of antibodies to surface antigens gradually declined, whilst levels of antibodies to CE antigens increased. The antibody response of intraperitoneally immunized fish was characterized by Western blotting of total soluble antigens obtained by heating and reduction of T. brevifilum spores at 95–100°C in Tris-HCl buffer containing SDS and dithiothreitol: a series of bands with molecular weights between 20 and 53 kDa was recognized by immunized turbot sera. Four additional bands (with molecular weights between 15 and 18kdA) were recognized by serum from re-immunized fish. ELISA studies of sera from naturally infected fish revealed a surprisingly low incidence of strong T. brevifilum seropositivity (61% individuals); antibodies to surface antigens predominated in seropositive individuals. The low background response levels and high sensitivity of the ELISA used in this study indicate that the assay is of value for the monitoring of serum antibody levels in turbot. However, given the relatively low seropositivities observed in naturally infected turbot, particularly to CE antigens, the use of anti-T. brevifilum serum antibody levels for the diagnosis of infection by this parasite may lead to false negative results.
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  • 3
    ISSN: 1365-2761
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The commercial furunculosis vaccine Aquavac Furovac 5 and an autogenous vaccine, based on the challenge strain, induced immune protection in turbot, Scophthalmus maximus (L.), as shown in challenge tests 120 days post-immunization by injection (relative percentage of survival, RPS = 72–99%). This protective effect lasted for at least 6 months post-immunization at appreciable levels (RPS = 50–52%). Neither the autogenous vaccine nor the commercial vaccine was able to induce significant levels of protection against Aeromonas salmonicida in turbot when administered by immersion. Antibody levels were high or moderate in fish vaccinated by injection with the different vaccines and very low in fish vaccinated by immersion. The field results show that delivering an oral boost after the primary vaccination by injection did not enhance protection of turbot against furunculosis and that water-based (autogenous vaccine) and oil adjuvanted (Alpha Ject 1200) vaccines administered by injection conferred similar levels of protection (RPS 〉 80%) in turbot.
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  • 4
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract This work investigated the location on the parasite ofTrichinella antigens recognized by the mouse immune system and the question as to which of them bear the epitope phosphorylcholine (PC). Wheatley's trichrome stain (initially developed for faecal smears) proved to be excellent for visualization ofTrichinella structures, enabling four types of stichocyte to be distinguished. By applying this stain on infected muscle sections after immunocytochemistry using (a) anti-PC BH8 monoclonal antibodies, (b) serum from mice that had been infected twice in the presence of 0.05% thiabendazole (to prevent reproduction by adult females) and then bled on day 7 post-reinfection, (c) serum from infected mice that were bled on day 14 postinfection, or (d) serum from infected mice that were bled on day 42 postinfection, we found (1) that PC is an abundant structural epitope on the hypodermis/muscle, genital primordium and intestinal tract but is absent from the cuticle and stichosome; (2) that the principle secretory cells of adult worms are delta- and beta-stichocytes, whereas those of migrating and encysted L1 larvae are alpha-stichocytes; and (3) thatTrichinella antigens recognized in the encysted phase of the parasite's life cycle are present in parasitized myofibres in the sarcoplasmic matrix and in the nucleoplasm of hypertrophic nuclei. The significance of these findings is discussed.
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  • 5
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Mice primed withTrichinella spiralis orT. pseudospiralis and reinfected with either the homologous or the heterologous species produced high levels of IgE antibodies that cross-reacted with the non-inducing strain in passive cutaneous anaphylaxis assays. Crossreactive antibodies were not induced by primary infection. Cross-reactivity persisted for more than 6 months following secondary infection or destruction of encysted larvae with mebendazole. Both the prevention of larviposition by thiabendazole and the interruption of infection using naphthalophos indicated that the presence of the pre-adult stage alone provided sufficient priming for the induction of detectable levels of cross-reactive IgE by subsequent reinfection. These results suggest the existence of two sets ofTrichinella allergens, one comprising species-specific major allergens (MAs) and the other comprising minor allergens (mAs) evoking a crossreactive IgE response that occurs to a detectable extent only when the response to MAs has reached its ceiling. These findings are relevant to the design of experiments investigating the role played by IgE antibodies in protection against reinfection in rodents.
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  • 6
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  We used ELISA and immunoblotting to investigate antigenic cross-reactivity in mice between third-stage larvae of Anisakis simplex and five other nematodes: the ascaridoids Ascaris suum, Toxocara canis and Hysterothylacium aduncum, and the nonascaridoids Trichinella spiralis and Trichuris muris. Two sera were raised against each species (including A. simplex, but excluding A. suum), by infection or by immunization with somatic antigens. Serum against A. suum was raised by immunization only. The reactivities of each serum with A. simplex somatic antigens (SA), excretion-secretion antigens (ES), pseudocoelomic fluid antigens (PF) and cuticular antigens (CA) were investigated. The results of ELISA indicated high antigenic cross-reactivity between A. simplex and the remaining ascaridoid nematodes, confirming that there is extensive antigenic similarity within this group of nematode parasites. Immunoblotting again confirmed the high degree of cross-reactivity between the SA of A. simplex and SAs of the other ascaridoids, although several A. simplex SA components in the 11–18 kDA range were only recognized by sera from mice infected with A. simplex. In addition, two A. simplex PF components of 22 and 27 kDA, were recognized only by sera from mice infected with, or immunized with the SA of, A. simplex. Finally, the anti-phosphorylcholine monoclonal antibody BH8 recognized only a small number of A. simplex antigens, indicating that phosphorylcholine epitopes are not significant contributors to the observed cross-reactivity with the other nematodes.
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  • 7
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
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  • 8
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Although the presence of phosphorylcholine (PC) inTrichinella is well established, the structures of the TSL-4 antigens that bear this epitope are unknown. A subset of TSL-4 antigens (TSL-8 antigens) has been reported to be absent from the surface of first-stageT. spiralis larvae. We report experiments with a monoclonal antibody (mAb US2) developed in mice with a relative inability to produce antibodies to PC. In immunoblotting, mAb US2 and anti-PC mAb (BH8) showed apparently identical binding patterns. In addition, we used an immunogold double-labeling technique to study the anatomical distribution of the epitopes recognized by these mAbs; the results obtained indicate close colocalization of epitopes for BH8 and US2 in tissues ofT. spiralis first-stage larvae. On the basis of these results, we suggest that US2 probably binds to allT. spiralis TSL-4 antigens, including TSL-8 antigens. We also clarify some conflicting previous reports on the distribution of PC immunoreactivity in first-stage larvae ofT. spiralis.
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  • 9
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The effect of the adult worms and migrating L1 larvae of Trichinella spiralis on the production of specific IgE antibodies was determined in BCF1 mice. To achieve this, we combined the effect of two anthelminthics: thiabendazole, to produce chemosterilization of adult females, and napthalophos, to expel adult worms from the intestine of infected mice on the desired day. Our results demonstrate that when the natural route of infection is used the production of IgE antibodies is not dependent on the infection dose or the number of migrating L1 larvae, and that both intestinal worms and migrating L1 larvae contribute to the production of reaginic antibodies. In addition to this, an extended period of antigenic stimulation (10–12 days) is required for the induction of a detectable, specific IgE response by adult worms. Finally, our results seem to indicate that although the effects of adult worms and migratory L1 larvae on the IgE production are not additive, the presence of adult worms in the intestine of mice may stimulate a secondary exposure to common antigens released by the migrating L1 larvae.
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  • 10
    ISSN: 1432-1955
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  The avidin-biotin molecular recognition system is widely used in parasite immunology. However, the presence of biotin and/or biotin-containing molecules (BCMs) in samples may lead to erroneous results. In the work reported herein we investigated the extent to which biotin and BCMs present in helminth extracts may interfere in avidin/biotin-based immunoassays and developed an enzyme-linked immunosorbent assay (ELISA) for quantification of these components. In avidin-based ELISA using antinematode monoclonal antibodies, an extract of the nematode Anisakis simplex showed very high background reactivity due to biotin/BCMs, whereas the background reactivity in an extract of the nematode Trichinella spiralis was negligible. To investigate interspecies differences further, we performed Western-blot analyses (with avidin as the detector) of extracts from seven nematodes (A. simplex, Ascaris suum, Toxocara canis, Hysterothylacium aduncum, T. spiralis, and Trichuris muris) and the cestode Bothriocephalus scorpii. Even within superfamilies there was considerable variation in the banding patterns obtained. The above-mentioned results confirm that biotin and BCMs may be a significant source of interference in ELISA and immunoblotting, two of the techniques most widely used in parasitological immunodiagnosis. A competition ELISA designed to allow accurate quantification of biotin and BCMs in helminth extracts likewise indicated very considerable interspecies variation. Both A. simplex and H. aduncum had very high biotin/BCM contents. Microdialysis of extracts in the presence of dimethylsulfoxide to remove free biotin prior to ELISA indicated that the high biotin/BCM content of the H. aduncum extract (but not the A. simplex extract) was very largely due to free biotin. Taken together, these results indicate that extreme caution should be exercised in the use of avidin/biotin-based immunoassays for the detection of helminth antigens and that in many cases it may be better to use an alternative recognition system.
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