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PubMed1994

Cell-mediated immunity in Merino lambs with genetic resistance to Haemonchus contortus.

Lymphocyte proliferation and skin hypersensitivity responses to parasite antigens were measured as indicators of cell-mediated immunity in genetically resistant and random-bred lambs following infection with Haemonchus contortus. Responses of PBMC to the mitogens PHA, PWM and LPS were also assessed to determine if infection was associated with suppression of general immune function. All lambs demonstrated significant antigen-specific lymphocyte proliferation prior to experimental infection. The responses did not differ between resistant and random-bred lambs and the correlation between pre-infection stimulation indices and resistance to challenge infection was also not significant. Following infection, PBMC from resistant lambs consistently exhibited higher blastogenic responses to both larval and adult antigens than PBMC from random-bred lambs. Addition of cyclosporin A to the antigen-stimulated cultures resulted in significant inhibition of blastogenesis, suggesting that the cells responding to parasite antigens were predominantly of the T-helper cell phenotype. Proliferation responses of resistant and random-bred sheep were similar for all mitogens tested. However, a transient suppression of lymphocyte reactivity to PHA and PWM was noted in all infected lambs. Responses to LPS remained unaffected during infection. There was no difference in the immediate hypersensitivity responses to intradermal inoculation of parasite antigens between the genotypes. However, the delayed-type hypersensitivity responses to parasite antigens were significantly greater in resistant lambs than those in random-bred lambs. Together, these data suggest that resistant lambs have an enhanced ability to mount parasite-specific cell-mediated immune responses when compared with random-bred lambs and that this may contribute to differences in the susceptibility of two genotypes to haemonchosis.

Animals↗

PubMed1995

The effect of reduced feed intake on the efficacy of oxfendazole against benzimidazole resistant Haemonchus contortus and Trichostrongylus colubriformis in sheep.

Within 12 h of reducing the daily intake of a 50:50 lucerne and wheaten hay ration from 800 to 400 g, there was an increase in digesta marker concentration indicating a reduction in the flow rate of digesta fluid and particulates through the abomasum of sheep. The efficacy of the recommended dose of oxfendazole (OFZ) was then tested against benzimidazole resistant Trichostrongylus colubriformis and Haemonchus contortus in sheep whose feed intake was halved 36 h before and after drug treatment. In animals fed the reduced ration there was a 60% reduction of T. colubriformis and 94% reduction of H. contortus compared with a 19% and 60% reduction respectively on the higher feed intake. It is therefore suggested that greater anthelmintic efficacy can be achieved in sheep by temporary feed reduction.

Animals↗

PubMed1995

Levamisole resistance in Haemonchus contortus selected at different stages of infection.

Resistance to the anthelmintic levamisole is common amongst most genera of trichostrongylid nematodes of sheep, but is relatively rare in the species Haemonchus contortus. The level of resistance in at least one strain of this species varies during the life cycle. Investigation of this phenomenon, especially in parasitic stages of the parasite may reveal information on resistance mechanisms and factors affecting the selection of resistance. Compared with adult worms of the same resistant strain, immature worms are more susceptible to levamisole both in in vitro contraction assays and an in vivo efficacy experiment. To ascertain if immature parasitic worms of a susceptible strain could be selected for resistance, 3 lines of worms were selected with levamisole for 9 generations. Lines were selected with the same dose rate (2 mg kg-1) at the 4th and 24th day of infection. In addition, because levamisole is less active against immature worms a further line was selected at the 4th day with a higher dose (8 mg kg-1) but similar efficacy to the selection at day 24. In in vitro assays the line selected at day 4 with the lower dose of levamisole showed no resistance compared with an unselected line. The other 2 lines developed high levels of resistance indicating that resistance can develop after selection at either stage. In addition these data directly show that selection pressure is important in determining the rate of development of resistance.

Animals↗

PubMed1995

Progress on vaccination against Haemonchus contortus.

Control of Haemonchus contortus at present is largely by the use of anthelmintics, assisted in some regions by management programs. Widespread development of resistance, particularly in South Africa and Australia, and concerns associated with the manufacture and use of chemicals have led to increasing interest in vaccination as an alternative means of control. Vaccination strategies basically fall into 2 categories, 'hidden' antigens (usually derived from the gastrointestinal tract of the adult parasite), or 'natural' antigens (those exposed to the immune system of the host during the course of infection, usually derived from the infective larval stage). Particularly promising results have been obtained using the hidden gut antigen H11, or H110D, and more recently with another hidden antigen, H-gal-GP. The use of a natural antigen vaccine, however, would provide advantages such as boosting of the immune response by field challenge. This article will review recent developments in both types of vaccines against H. contortus and consider the advantages and disadvantages of the 2 approaches.

Amino Acid Sequence↗

PubMed1996

Influence of organic acid excretion on cuticle pH and drug absorption by Haemonchus contortus.

To determine if a cuticle microenvironment pH is maintained by adult Haemonchus contortus, organic acid excretion kinetics and absorption kinetics of selected model weak acids and a weak base were measured in incubation media that varied in buffer capacity (0.25-20 mM HEPES or 5 mM glycine) and initial pH (7.5 or 3.5). To evaluate the importance of the cuticle as a pathway for organic acid excretion and drug absorption the pharynx was paralyzed with 1 nM ivermectin. H. contortus changed the media pH from initial values of 7.5 or 3.25 to an asymptotic value of approximately 5.6. The rate of pH change depended on the buffer capacity, but was not affected by chemical ligation with ivermectin. The intrinsic rate of excretion of organic acids (0.045 +/- 0.016 micromol/cm2 x h) was constant during the first 8-12 h of incubation and was independent of initial pH, buffer capacity or ivermectin ligation. The rates of absorption of the model weak acids, benzoic acid and p-nitrophenol, and the model weak base, aniline, were not affected by initial pH, buffer capacity or ivermectin ligation. These results suggest that H. contortus excretes organic acid endproducts of carbohydrate metabolism across its cuticle, and that these acids maintain a microenvironment pH within the water-filled pores of the cuticle that controls the rate of adsorption of weakly acidic or basic drugs.

Absorption↗

PubMed1995

Immune responses of sheep to microdissected parts of Haemonchus contortus.

Adult specimens of Haemonchus contortus were microdissected into four body fragments: oesophagus, cuticle of oesophagus, gut and cuticle with adjacent muscle layer. The antigenicity of these different body fragments was analysed in comparison to total (whole) worm extracts with immunoblotting and ELISA using sera of H. contortus-infected sheep. In particular, oesophagus-derived antigens appeared to be specifically recognized and may prove valuable in diagnostic assays.

Animals↗

PubMed1996

Lectin staining of trichostrongylid nematode eggs of sheep: rapid identification of Haemonchus contortus eggs with peanut agglutinin.

The binding of 10 different lectins to the surface of trichostrongylid nematode eggs of sheep were investigated. Osage orange seed agglutinin bound to eggs from Haemonchus contortus, Ostertagia circumcincta and Trichostrongylus colubriformis, jack bean agglutinin to H. contortus and O. circumcincta eggs and peanut agglutinin to H. contortus eggs only. Staining of egg mixtures with fluorescein-labelled peanut agglutinin showed that the lectin correctly identified the percentage of H. contortus eggs present. Identification of eggs by lectin staining correlated well with results from larval differentiation. The findings suggest that peanut agglutinin can be used in routine diagnostic parasitology for the rapid identification of H. contortus eggs.

Animals↗

PubMed1996

Reduction of Haemonchus contortus infective larvae by Harposporium anguillulae in sheep faecal cultures.

Harposporium anguillulae is a common nematophagous fungus which colonizes cattle pats deposited in Brachiaria decumbens pasture in Brazil. To assess its possible effect on free living stages of trichostrongylid nematodes, the reduction of Haemonchus contortus infective stages in sheep faecal culture was evaluated in vitro. Addition of 300,000 conidia per gram of faeces caused a reduction of 99.5% compared to the control group. This result indicates that H. anguillulae could be a candidate for the development of strategies for the biological control of trichostrongylid nematodes.

Animals↗

PubMed1991

Fecundity of anthelmintic resistant adult Haemonchus contortus after exposure to ivermectin or benzimidazoles in vivo.

A strain of Haemonchus contortus resistant to the benzimidazoles, ivermectin and salicylanilides was used to study the effect of exposure to ivermectin or oxfendazole on the number of eggs produced by adult female parasites. After anthelmintic administration there was a reduction in the faecal egg count of the infected lambs. Ivermectin caused a reduction in the number of eggs within adult parasites for up to 72 hours but this returned to higher than pre-treatment levels at 168 hours after treatment. After oxfendazole treatment there was an increase in the number of eggs stored in the adult parasites at 24 hours and then a significant reduction in the number of eggs for over 168 hours. This suggested that both ivermectin and oxfendazole have a deleterious effect on the reproductive potential of female H contortus although the parasites are resistant to these drugs. These results confirm the importance of allowing up to 10 days to elapse between the time of administration of an anthelmintic and the collecting of faecal samples when using the faecal egg count reduction test to identify anthelmintic resistance, especially when using benzimidazoles. The more prolonged effect on fecundity produced by oxfendazole may reflect the antimitotic action of the benzimidazole anthelmintics.

Animals↗

PubMed1992

Effects of dietary molybdenum on nematode and host during Haemonchus contortus infection in lambs.

The addition of molybdenum (0.05 mmol kg-1 dry-matter) to the diet of lambs given a trickle infection of Haemonchus contortus larvae (500 third stage larvae d-1 over six weeks) reduced mean faecal egg counts (epg) from 3952 to 2312 +/- 402 by 32 days (P less than 0.02) and greatly reduced the mean number of worms recovered from the abomasum 14 days after infection ceased (907 compared with 4167: P less than 0.01). Infection reduced haemoglobin concentrations less in lambs given molybdenum although the difference was small relative to the reduction in worm burden. Lambs not given molybdenum had low intraepithelial mast cell counts in the abomasal mucosa and less abomasal hypertrophy than expected from abomasal parasitism. Molybdenum did not consistently reduce the copper status of the host or the parasite. Previous exposure to molybdenum greatly reduced protein but not proteinase activity in, or secreted by, adult worms cultured for eight hours. It is suggested that molybdenum either increased the inflammatory response which preceded worm rejection or that it indirectly enhanced that reaction by reducing the effectiveness of copper-dependent, anti-inflammatory enzymes in the gastrointestinal mucosa.

Abomasum↗

PubMed1993

Protection in lambs immunised with Haemonchus contortus gut membrane proteins.

Substantial protection against Haemonchus contortus was induced in lambs by hyperimmunising them with an integral membrane extract of intestines dissected from adult worms. The procedure was effective in young growing animals but did not afford any cross protection against either Ostertagia circumcincta or Nematodirus battus. A serum transfer experiment suggested that the effector mechanism was serum antibody which bound to the brush border membrane of the parasites intestinal cells. Sheep which had acquired immunity to H contortus by previous exposure to a trickle infection did not recognise the gut membrane proteins, which suggested that these are normally hidden from the host.

Animals↗

PubMed1993

Evaluation of aspects of the protection afforded to sheep immunised with a gut membrane protein of Haemonchus contortus.

Experiments were undertaken to evaluate the type of protection conferred against Haemonchus contortus by immunising sheep with H11, a 'hidden' integral gut membrane protein isolated from this parasite. A serial kill experiment showed that worms began to be lost from immunised lambs between seven and 14 days after challenge with a single dose of larvae and adult female worms were more susceptible than male worms. Seven-day-old juveniles survived immunisation even though their intestinal cells became coated with sheep antibody. Immunisation was equally effective against both benzimidazole-resistant and susceptible strains of H contortus. When immunised lambs were subjected to a trickle infection, they were largely protected against the anaemia and egg output observed in the challenge controls. Moreover, they grew as fast and as efficiently as uninfected lambs fed on the same high protein diet and acquired a natural immunity during the course of the trickle infection.

Animals↗

PubMed1993

Survival and development of ivermectin-resistant or susceptible strains of Haemonchus contortus under field and laboratory conditions.

The free-living development of three strains of Haemonchus contortus was studied in two experiments. Day 21 faecal samples were collected from lambs infected with either a susceptible strain, a laboratory-selected ivermectin (IVM) resistant strain or a South African field strain showing multiple anthelmintic resistance, which included IVM. No eggs hatched in samples cultured at 4 or 10 degrees C. At 22 degrees C the laboratory-selected strain showed the highest rate of development while at 27 degrees C the susceptible strain produced the highest yield of third stage larvae (L3): at both temperatures the field strain showed the lowest percentage development to L3. The second experiment was a field study carried out in southern Brazil. Faeces containing either an IVM-susceptible or an IVM-resistant strain of H contortus were placed in two series of grass plots during each of three summer months. Soil subsequently yielded more larvae than did grass suggesting migration or mechanical transport into the soil. For plots contaminated during the first two months there was no significant difference in recovery rate between the two strains (P > 0.05). When contamination occurred during the third month, the IVM-resistant strain produced significantly higher recovery rates (P < 0.05) from both pasture and soil.

Animals↗

PubMed1993

The same fractions of Haemonchus contortus soluble antigen induce lymphocyte responses in naive lambs and immune sheep.

Soluble antigen extracted from third stage larvae of Haemonchus contortus was divided into fractions based on molecular weight by sodium dodecyl sulphate polyacrylamide gel electrophoresis and electrotransferred to nitrocellulose. Infection immunised adult sheep and H contortus naive lambs were used to examine the lymphocyte responses to these fractions in vitro. The results demonstrate that the same two fractions induced the strongest responses in both groups of animals.

Animals↗

PubMed1994

Serum antibody responses of Texel sheep experimentally infected with Haemonchus contortus.

The primary and secondary serum antibody responses of Texel sheep to infective larvae (L3) and adult worms of Haemonchus contortus were studied. Ten-month-old sheep were infected with 20,000 H contortus L3, treated with ivermectin seven weeks later and, after four weeks, reinfected with 10,000 L3 once a week for six weeks. Faecal egg counts were significantly lower during the secondary infection than during the primary infection, but both infections induced antibody responses, as demonstrated by an enzyme-linked immunosorbent assay (ELISA). The primary antibody response developed rather slowly, but the secondary response developed more rapidly and the IgA responses against L3 antigens and the IgG1 and IgG2 responses against adult antigens were twice those observed during the primary infection. These accelerated and enhanced responses after the reinfection suggest an immunological memory for H contortus antigens.

Animals↗

PubMed1977

A helical, polymeric extracellular protein associated with the luminal surface of Haemonchus contortus intestinal cells.

The structure of the intestinal cells of the parasitic nematode Haemonchus contortus is described. The cells have numerous microvilli about 0-09 micron in diameter; most being 5-5-7-5 micron in length. The microvillar (plasma) membrane is coated with a layer of amorphous material (glycocalyx) about 60 A thick which is electron dense in sectioned preparations. Associated with the surface of this material, and filing the spaces between the microvilli, are filaments in the form of helices about 400 A in diameter and of variable pitch. The helices appear to be flexible but they are aligned approximately with the long axes of the microvilli. There are up to ten helices per microvillus; they extend beyond the tips of the microvilli and are up to 10 micron long. The material has been obtained nearly pure in small amounts. It is primarily protein and it is proposed that it should be called contortin. The monomeric form (of molecular weight about 60,000) has been identified with a Y-shaped structure with arms about 45 A long and 25 A wide seen in negatively stained preparations. The helical filament appears to be formed by lateral polymerization of patirs of these units.

Animals↗

PubMed1992

Localization of immunoglobulin-containing cells in the abomasum of sheep following infection with Haemonchus contortus.

Abomasal cannulation followed by serial collection of biopsies was used to study the kinetics of appearance of IgA-, IgG1-, IgG2- and IgM-containing cells in the abomasum of sheep following infection with Haemonchus contortus. Very few immunoglobulin-containing cells (ICC) were found in the abomasum of sheep before infection. Following H. contortus infection, there were increased numbers of ICC in the submucosa of the abomasum. Seven days after infection, the numbers of IgA-, IgG1- and IgM-containing cells were six times greater than for uninfected control animals. The numbers of ICC continued to rise as the infection progressed, and the peak response was observed between 21 and 28 days after infection. IgA-containing cells (68-84%) were the most frequent cell types at all the observation times, followed by IgG1 and IgM. IgG2-containing cells were minimal throughout the experiment. As there were no significant changes in the numbers of ICC in the abomasum of uninfected controls it is concluded that H. contortus stimulated a local immune response in the abomasum of parasitized sheep.

Abomasum↗

PubMed1995

Humoral and cellular responses following local immunization with a surface antigen of the gastrointestinal parasite Haemonchus contortus.

A purified, larval specific antigen of the abomasal parasite Haemonchus contortus was used to immunize sheep. In an attempt to induce a local immune response in the abomasum, the antigen was injected twice into the abomasal wall after one peripheral immunization. Serum antibody responses were boosted by each intra-abomasal immunization but not by the challenge infection given 3.5 weeks after the last immunization. Examination of the specific antibody secreting cells (ASC) recirculating in the peripheral blood indicated that there was an increase in blood ASC 5 days after local stimulation. This increase was maintained only after immunization and not after infection, probably reflecting the different responses induced when antigen is presented by injection in an adjuvant or by the parasite during infection. High proliferative T cell responses in the abomasal lymph nodes were only observed in one of the five sheep immunized with antigen; this was also the only sheep in this group to maintain an adult parasite burden at postmortem corresponding with the lowest antibody response. Peak faecal egg counts after infection were reduced by 54% in the immunized group compared to control sheep. Egg counts in the control sheep were, however, variable and dropped quickly, probably as a consequence of the inflammatory response induced by the injection of aluminium hydroxide into the abomasal wall.

Abomasum↗