Adult Haemonchus Contortus Research Articles

Anthelmintic activity of extracts of Artemisia absinthium against ovine nematodes

The increasing prevalence of anthelmintic resistant strains of helminths, drug residues in animal products and high cost of conventional anthelmintics has created an interest in studying medicinal plants as an alternative source of anthelmintics. Artemisia absinthium Linn. (Tethwen) is used traditionally by people as a vermifuge in addition to its other livestock uses. The objective of this study was to evaluate the anthelmintic efficacy of crude aqueous extracts (CAE) and crude ethanolic extracts (CEE) of the aerial parts of A. absinthium in comparison to albendazole against the gastrointestinal (GI) nematodes of sheep. To fulfill the objectives, the worm motility inhibition assay was utilized in order to investigate the direct effects of plant extracts on the survival of the adult nematodes under in vitro conditions and faecal egg count reduction assay to investigate the effects on faecal egg output of GI nematodes under in vivo conditions. Significant anthelmintic effects of CAE and CEE on live adult Haemonchus contortus worms ( P < 0.005) were observed in terms of the paralysis and/or death of the worms at different hours post-treatment (PT), however, CEE were more efficacious than CAE. The oral administration of the extracts in sheep was associated with significant reduction in faecal egg output by the GI nematodes. The CEE was as effective as the reference drug-albendazole and demonstrated faecal egg count reduction (FECR) of 90.46% in sheep at 2.0 g kg −1 body weight on day 15 PT followed by 82.85% FECR at 1.0 g kg −1 bw on day 15 PT. The CAE showed less activity and resulted in maximum of 80.49% FECR at 2.0 g kg −1 bw. Dosage had a significant ( P < 0.05) influence on the anthelmintic efficacy of A. absinthium. The better activity of CEE can be attributed to the greater concentration of alcohol soluble active anthelmintic principle/s and a more rapid transcuticular absorption of the CEE into the body of the worms when compared with the CAE. The results of the present study suggest that A. absinthium extracts are a promising alternative to the commercially available anthelmintics for the treatment of GI nematodes of sheep.

Isolation and immunolocalization of a putative protective antigen (p26/23) from adult Haemonchus contortus

The putative protective antigen, p26/23, from adult Haemonchus contortus was isolated. A soluble extract from adult helminths obtained from the abomasa of hyperinfected (12,000 infective larvae) female Manchego lambs and treated with a mixture of protease inhibitors was subjected to affinity chromatography (hexylgluthatione) to eliminate the enzyme gluthatione S-transferase. The eluate was analyzed by electrophoresis under denaturing and reducing conditions (sodium dodecyl sulfate polyacrylamide gel electrophoresis), electrotransferred to nylon membranes, and assayed by Western blot with sera from immunized lambs. The bands recognized by the lambs' sera corresponding to proteins with a molecular weight of 23-26 kDa (p26/23) were excised, eluted, and separated by reverse-phase chromatography. This allowed the isolation of a single protein, which was expressed in both infective larvae (L3) and the adult stage of the parasite. The first 20 amino acids of the N-terminal end of the purified protein were determined. The partial amino acid sequence revealed a 100% identity with the N terminus of one of the peptides present in the p26/23 immunoprotective fraction previously tested by us against sheep haemonchosis. No significant homology with any reported sequence was found except for the deduced sequence of a hypothetic H.contortus protein (HCC00515). Immunolocalization studies showed that the protein was expressed in the hypodermic chords of the nematode.

Direct and indirect anthelmintic effects of condensed tannins in sheep

Anthelmintic activity of condensed tannins (CT) was evaluated both in vitro and in vivo. In vitro tests included egg hatch test and paralysis/mortality assay on adult Haemonchus contortus. In vivo anthelmintic effect was determined by faecal egg count reduction test in lambs. To this end, 18 lambs were divided into three groups (low tannin, high tannin and control). The lambs of low and high tannin groups were fed diets containing 2 and 3% CT while the control group was fed on diets without CT. In vitro trials showed a dose-dependent inhibition of nematode egg hatching; whereas, there was no effect of CT on adult H. contortus. In vivo trials indicated reduction in faecal egg counts in lambs fed diets containing CT. Feed intake and nutrient digestibility of CT-fed sheep was lower and nitrogen balance was higher as compared to control. Maximum weight gain was observed in animals fed diets containing 3% CT. The direct anthelmintic effect of CT, therefore, was evidenced by inhibited egg hatching; whereas, faecal egg counts reduction in sheep was through improved nutrient utilization.

Identification of a 66 kDa Haemonchus contortus excretory/secretory antigen that inhibits host monocytes

A 66 kDa adult Haemonchus contortus excretory/secretory (E/S) antigen was identified in Western blot by reaction with sera from the infected goats. The protein was purified from the adult worm extract and E/S products by anion exchange and ConA-Sepharose chromatography. The purified protein inhibited monocyte function in vitro as judged by decreased production of hydrogen peroxide and nitric oxide in the culture medium. The protein also caused proliferation of peripheral blood mononuclear cells. The absence of protein in the free living L3 larvae suggests that the expression of this protein coincides with the adaptation to the parasitic life. A correlation of antibody titre and worm burden was observed in the infected goats with higher antibody levels in high worm burdened animals. Anti-protein antibody caused loss of adult worm motility in vitro resulting in the death of the parasite. The fact that the protein is recognized by the host together with in vitro killing of adult parasites by antibodies makes this protein a promising candidate for vaccination trial.

Production of eosinophil chemoattractant activity by ovine gastrointestinal nematodes

Eosinophilia is a well documented feature of helminth infections but the precise nature of the interaction between parasite and eosinophil remains an enigma. This paper describes experiments demonstrating that ruminant gastrointestinal trichostrongyles produce potent chemoattractant activity for ovine bone marrow-derived eosinophils in vitro. This activity was initially identified as a constituent of whole worm extracts of third and fourth larval (L3, L4), and adult stages of Teladorsagia circumcincta, and adult Haemonchus contortus. Similar activity was detected in excretory/secretory (E/S) material derived from live T. circumcincta L3. Subsequently, by adapting the assay technique to incorporate live worms directly into the system, it was shown that L3 of both T. circumcincta and H. contortus produced eosinophil chemoattractant activity. In contrast, neither whole worm extracts, or E/S preparations from mixed stages of the free-living nematode Caenorhabditis elegans contained eosinophil chemoattractant activity, and there was no evidence of chemoattractant production by live C. elegans. The results described are challenging to the traditional dogma that eosinophils are host-protective effector cells, and raise the intriguing possibility that ovine nematodes actively encourage recruitment of eosinophils. Local eosinophil-mediated mucosal damage, comparable to that seen in the asthmatic lung, may then provide a permissive local microenvironment for the parasite. Moreover, if they prove important for pathogenicity, nematode chemoattractants could offer future potential as novel therapeutic targets.

Protection induced by vaccinating sheep with a thiol‐binding extract ofHaemonchus contortusmembranes is associated with its protease components

Previous work has shown that a protein extract enriched for cysteine protease activity (TSBP) prepared from adult Haemonchus contortus using thiol sepharose affinity chromatography confers substantial protection against a single challenge infection. TSBP comprised proteases and other proteins. Here, TSBP were further fractionated using anion-exchange chromatography and fractions pooled on the basis of containing (1) protease activity, (2) a prominent non-protease peptide and (3) material which did not bind to the column. A protection trial showed that only the protease-enriched material conferred protective immunity and this was comparable to that observed in a TSBP-immunized positive control group. Immunization stimulated a marked IgG response with the IgG2 isotype predominating.

Presence of the LDNF glycan on the host‐protective H‐gal‐GP fraction from Haemonchus contortus

Immunization of sheep with the gut membrane-associated protein complex H-gal-GP of adult Haemonchus contortus induces high levels of protection against a homologous challenge infection. Protection is correlated with a systemic IgG response against the antigen. Analysis of the antibody response showed that the majority of the antigen-specific IgG was of the IgG2 isotype. A substantial proportion (74%) of this response was directed against the glycan component of H-gal-GP. The high immunogenicity of the H-gal-GP glycans may be due to the presence of the fucosylated LacdiNAc (LDNF) antigen. 2D electrophoresis, Western blotting and mass spectrometry analysis of H-gal-GP showed that this glycan epitope was specifically located on a metalloendopeptidase, MEP3. MEP3 is the most abundant protein in H-gal-GP and has been identified as one of the most likely protective components of the complex. Here, we present evidence that the LDNF glycan does not contribute to the protective capacity of H-gal-GP. Animals vaccinated with reduced and denatured H-gal-GP are not protected against subsequent infection, although the antibody response against the LDNF glycan is very similar to that of animals vaccinated with the native H-gal-GP. In addition, an alternative version of H-gal-GP, H-sialgal-GP, which is equally protective, but isolated by affinity chromatography on jacalin lectin rather than peanut lectin, contains a MEP3 component which has no detectable LDNF glycan.

In vitro effects of four tropical plants on three life-cycle stages of the parasitic nematode, Haemonchus contortus

Alcoholic extracts of four tropical plants ( Zanthoxylum zanthoxyloides, Newbouldia laevis, Morinda lucida and Carica papaya) were screened in vitro for potential anti-parasitic effects against eggs, infective larvae and adult Haemonchus contortus. Significant effects were obtained with all four plants but differences were observed depending on the parasitic stage. The effects of the four plant extracts were similar on egg hatching and were dose dependent. In contrast, no dose–response relationship was found for infective larvae and adult worms, although more potent effects were usually observed with the highest concentrations. Using a larval inhibition migration test, extracts of fagara ( Z. zanthoxyloides) were found to be less active against Haemonchus infective larvae than were the other plants. N. laevis was found to be highly and rapidly effective against adult worms. Overall, these in vitro results suggest that these four plants, traditionally used by small farmers in Western Africa, do possess anti-parasitic properties. These effects remain to be confirmed through in vivo studies.

Characterization of proteinases in different isolates of adult Haemonchus contortus

A high degree of intra- and inter-geographical variation has been demonstrated previously in the excretory/secretory proteinases released by adult Haemonchus contortus. Proteinase activity has also been associated with host-protective 'hidden' antigens isolated from the gut of adult H. contortus. If similar geographical strain variation also exists within the gut-associated proteinases, this will have important implications for the development of a globally effective vaccine. The proteinases active in integral-membrane protein extracts from 3 different strains of adult H. contortus were characterized on the basis of their pH optima and molecular size. Although enzyme activity was detected over a wide pH range, the majority of proteinase activity was detected at acidic pH. Differences in specific activity and size of enzymes were observed between the 3 different parasite strains at different pH values. A high degree of conservation in reactive peptides was observed when protein extracts were probed with antisera raised to the protective hidden gut-antigen complexes isolated from the Moredun strain of H. contortus, or to bacterially expressed subcomponents thereof. Therefore, despite the observed differences in membrane-bound proteinase profiles, the similarity of the immunogenic response against these hidden antigens may be sufficient to prove protective against different geographical isolates of H. contortus.

Protection studies in sheep using affinity-purified and recombinant cysteine proteinases of adult Haemonchus contortus

Vaccination with a membrane-bound thiol Sepharose-binding fraction (TSBP) of adult Haemonchus contortus has been shown to confer significant levels of protection against homologous challenge in sheep. This fraction is greatly enriched for cysteine proteinase activity. Following fractionation of TSBP by anion-exchange chromatography on MonoQ, protection was found to partition with those fractions further enriched for cysteine proteinase activity. In this study, the cysteine proteinases of adult H. contortus TSBP were specifically purified by affinity chromatography using recombinant H. contortus cystatin, a potent cysteine proteinase inhibitor. Although only 1–1.5% of total TSBP bound to cystatin-Sepharose, this fraction contained 100% of the cysteine proteinase activity, as determined by gelatin substrate gel analysis. When used to immunise sheep, less than 3 μg per dose of this cysteine proteinase fraction was found to confer a substantial and repeatable level of protection against homologous challenge infection, reducing faecal egg counts by 48 and 28% and worm burdens by 44 and 46% over two trials. Host serum immunoglobulin levels and abomasal mast cell and eosinophil numbers were evaluated, although no correlation with protection was observed. Three cathepsin B-like cysteine proteinases present in TSBP (hmcp1, 4 and 6) have been identified previously by cDNA library immunoscreening. The predicted mature forms of these three cysteine proteinases were expressed in bacteria as insoluble, GST-fusion proteins. Following solubilisation in urea/DTT, the protective capacity of a cocktail of recombinant proteins was evaluated in sheep. Although no reduction in faecal egg counts was observed, sheep vaccinated with recombinant cysteine proteinases showed a highly significant 38% reduction ( P < 0.01) in worm burdens.

Haemonchus contortus: in vitro drug screening assays with the adult life stage

Motility and feeding assays were assessed as in vitro systems for screening of novel compounds for anthelmintic activity against adult Haemonchus contortus. The study aimed to develop an assay with the parasitic adult stage of this species that could be used in conjunction with, or as an alternative to, the free-living larval stage screens commonly used for drug discovery with many parasitic nematode species. The feeding assay showed limitations due to the apparent continuation of a significant degree of feeding in worms showing greatly reduced motility in the presence of some drugs. Hence, it appeared most likely that the feeding assay would underestimate the toxicity of these drugs. The motility assay was able to detect toxicity of known anthelmintics, including the ‘slow-acting’ benzimidazoles. A small-scale screening exercise used the motility assay to detect toxicity towards adult parasites in 10 compounds out of a group of 200 chemicals (selected due to known toxic effects in larval development assays). The motility assay appeared suitable for drug screening against adult H. contortus. The use of the adult stage for drug screening in this way ensures that the drug is toxic towards the parasite life stage to be targeted in vivo. A lack of activity in subsequent in vivo trials could, therefore, be most likely attributable to host pharmacokinetic factors rather than an intrinsic lack of activity of the drug towards the adult parasite. Index Descriptors and Abbreviations: Haemonchus, Nematode, Drug screening, Motility, Inulin

Vaccination against the nematode Haemonchus contortus with a thiol-binding fraction from the excretory/secretory products (ES)

Fractionated excretory/secretory products (ES) of adult Haemonchus contortus were evaluated as protective antigens. The proteins were successively eluted from a Thiol Sepharose column using 25mM cysteine followed by 25mM Dl-dithiothreitol (DTT). Sheep were vaccinated three times and challenged with 5000 third stage infective larvae (L3) of H. contortus. Highest level of protection was found in sheep vaccinated with the DTT-eluted fraction in which egg output and worm burden were reduced by 52 and 50%, respectively, compared to the adjuvant control group. There was a positive correlation between fecundity (number of eggs per female) and the cumulative EPG or worm burden. Serum and mucus antibody levels of ES-specific immunoglobulins increased after immunizations and after challenge for IgG, IgA and IgE. The harvesting of H. contortus from animals clustered per group revealed the presence of cysteine protease activity in the ES of all groups but in addition to that, metalloprotease activity was also detected in the groups vaccinated with the DTT-eluted fraction, total ES and adjuvant only, in contrast to previous batches of ES (completely inhibited by E64) obtained from non vaccinated animals.

Effects of condensed tannins on goats experimentally infected with Haemonchus contortus

Although the use of tanniferous plants or condensed tannins as an alternative to anthelmintics to control gastrointestinal nematodes has been largely documented in sheep, studies remain scarce in goats. The objective of this study was therefore to assess the possible impact of condensed tannins in goats infected with adult Haemonchus contortus. Two groups of cull goats were experimentally infected with 10.000 L3 of H. contortus. After 4 weeks, quebracho extracts, representing 5% of the diet DM, were administered for 8 days to one of the two groups. Goats of the second group remained as controls. One week after the end of quebracho administration, the goats were euthanised. Individual egg excretion and pathophysiological parameters were measured weekly during the study. At the end of the study, worm counts were assessed and histological samples from the abomasa were taken to count the numbers of mucosal mast cells, globule leukocytes and eosinophils. The administration of tannins was associated with a significant decrease in egg excretion, which persisted until the end of experiment. This reduction was not associated with any difference in worm number but with a significant decrease in female fecundity. No significant changes in the mucosal density of the three inflammatory cell types were detected between the two groups. These results indicate that the major consequence of tannin consumption in goats is a reduction in worm fecundity and egg output, which does not seem related to significant changes in the local mucosal response.

Immunization against ovine haemonchosis with three low molecular weight somatic antigens of adult Haemonchus contortus.

Individual bands (15) from electroblotted soluble extracts of adult Haemonchus contortus were excised and three peptides of molecular weight ca. 56 (F4), 39 (F8) and 18.5 kDa (F14) used to vaccinate 4-4.5-months-old lambs against the nematode. Immunizing doses from each peptide were administered in 1 ml Freund complete adjuvant (first 50 microg injection) and 1 ml Freund incomplete adjuvant (second and third 50 microg injections) to six lambs. Two weeks after last immunization, animals were challenged with 300 L-3/kg live weight (LW). Lambs were slaughtered 34 days after challenge. Immunization induced a strong antibody response estimated by enzyme-linked immunosorbent assay, whereas no peripheral lymphoproliferative response was observed. Lambs in the F8-vaccinated group showed on average delayed pre-patent period, lower faecal egg counts and reduction of abomasal worm burdens, although the differences were not statistically significant.

Excretory/secretory products of Haemonchus contortus inhibit aminopyrine accumulation by rabbit gastric glands in vitro

Abomasal parasites inhibit gastric acid secretion and reduce the number of acid secreting parietal cells either through physical contact with gastric tissue, the release of inhibitory excretory/secretory (ES) products or by initiating the host inflammatory response. To examine the role of parasite ES products, adult Haemonchus contortus were incubated in a medium based on Hank’s balanced salt solution and three incubates were tested for the presence of inhibitors of acid secretion by dispersed rabbit gastric glands in vitro, using the intracellular accumulation of 14 C -aminopyrine (AP) as an indirect measure of acid secretion. Acceptably sensitive rabbit preparations (80%) for testing ES products showed increased 14 C -AP accumulation with either 10 −5 or 10 −4 M histamine. Sheep glands proved unsuitable as a test system as only two of six preparations responded weakly to histamine. Inhibitory activity was demonstrated in all the three parasite incubates, although responses varied quantitatively between tests, even with the same batch of incubate. In single tests, 16% ES products significantly reduced the 14 C -AP ratio ( P<0.05) of 10 −4 M histamine-stimulated glands (15 of 19 tests with Batch 1 and two of six with Batch 3). Even at 1 and 5%, ES products (Batch 2) were inhibitory for 10 −5 M histamine-stimulated glands: at 1% the mean reduction was 26.0% (range 7.8–54.8%, n=10), four of which were statistically significant and at 5% was 17.6% (range −15.3–53.8%, n=17), four of which were statistically significant. ES products were overall inhibitory (−12%, P<0.05), when tested on glands exposed to increasing histamine concentrations from 10 −6 M to 10 –3 M (which increased the 14 C -AP ratio ( P<0.001)). Responses by individual gland preparations differed ( P<0.001). The active component(s) of the parasite ES products were less than 5000 molecular weight. Ammonium chloride, tested over the range of concentrations of ammonia present in worm incubates (0.2–1 mM, final concentrations in the gland incubations of 0.005–0.1 mM), had variable, but overall inhibitory effects on 10 −5 M histamine-stimulated glands ( n=3). When tested with a range of histamine concentrations, 0.01 mM NH 4Cl overall reduced the response by 8.6% (P<0.05) (n=4) . The similarity of responses of rabbit glands to parasite ES products and to NH 4Cl suggests that ammonia may be the small molecular weight ES product of adult H. contortus, which inhibited acid secretion in vitro.

Haemonchus contortus utilises catalase in defence against exogenous hydrogen peroxide in vitro

The toxicity of activated oxygen species towards adult Haemonchus contortus nematodes was examined in in vitro assays using ingestion of [ 3H]inulin to assess nematode viability. Both glucose/glucose oxidase (generation of hydrogen peroxide) and xanthine/xanthine oxidase (generation of superoxide anion) systems showed concentration-dependant toxicity to the nematodes. Both adult and larval Haemonchus contortus enzyme preparations showed significant catalase activities. Adult nematodes exposed to aminotriazole for 24 h showed catalase activities reduced to less than 20% of controls. Aminotriazole-treated nematodes exposed to a glucose/glucose oxidase system were significantly more susceptible to the toxic effects of the oxidant-generating system than controls (no aminotriazole pre-treatment). The concentration of glucose oxidase required to inhibit feeding by 50% was decreased 33-fold in aminotriazole-treated nematodes compared with controls. The effect of aminotriazole pre-treatment implicates hydrogen peroxide as a significant toxic agent in the glucose/glucose oxidase system. It is apparent that inhibition of Haemonchus contortus catalase increases the susceptibility of the parasite to the toxic effects of hydrogen peroxide, demonstrating a protective role for this enzyme. This suggests that catalase has the potential to play a significant role in the defence of this parasite against hydrogen peroxide produced as part of the respiratory burst of activated phagocytes within the host during its response to nematode infection.

Cathepsin B-like cysteine proteases and Caenorhabditis elegans homologues dominate gene products expressed in adult Haemonchus contortus intestine

Proteins expressed by nematode intestinal cells are potential targets for parasite control by immune or chemical based strategies. To expand our knowledge on nematode intestinal proteins, expressed sequence tags were generated for 131 cDNA clones from the intestine of adult female Haemonchus contortus. An estimated 55 distinct protein genes or gene families were identified. Predicted proteins represented diverse functions. Several predicted polypeptides were related to H. contortus proteins implicated in inducing protective immunity against challenge infections of this parasite. The dominant intestinal transcripts were represented by cathepsin B-like cysteine protease genes ( cbl) (17% of protein coding expressed sequence tags (ESTs) analyzed). An estimated 11 previously undescribed cbl genes were identified, doubling the recognized members of this gene family. Multiple C-type lectin sequences were identified. Other notable sequences included a predicted Y-box binding protein, serine/threonine kinases and a cyclin E-like sequence. Predicted protein homologues were found in Caenorhabditis elegans for all but one H. contortus sequence (99%), while fewer homologues from other parasitic nematodes were found. Many of the proteases, lipase and C-type lectin homologues in C. elegans had apparent signal peptides, suggesting that they are secreted. Several gene products had no obvious similarity outside the phylum Nematoda. The ESTs identified intestinal genes with potential application to immune control, understanding of basic intestinal regulatory processes and refinement of nematode genomic resources.

Preliminary observations on the potential of gut membrane proteins of Haemonchus contortus as candidate vaccine antigens in sheep on naturally infected pasture

In a trial lasting 11 months in South Africa, faecal egg counts and haematocrits of sheep vaccinated with gut membrane proteins of adult Haemonchus contortus were compared with unvaccinated controls grazing pasture contaminated with the parasite. Vaccination reduced egg output by >82% on average during one 4 month period of the trial and simultaneously significantly reduced the degree of anaemia and deaths due to haemonchosis. Although vaccine immunity was not sufficiently long lasting to prevent a surge in egg output which occurred after the onset of a period of irrigation, re-vaccinating the sheep at this point cleared their newly acquired infection and rapidly restored protection to approximately the level observed beforehand. It was clear that a vaccine based on parasite gut membrane proteins could offer substantial benefits in the control of natural haemonchosis.

Serodiagnosis of haemonchosis with a somatic antigen (Hc26) in several breeds of sheep.

Sera from 53 sheep belonging to Castellano, Churro, Manchego, and Merino breeds were analyzed to test the diagnostic value of a 26-kD antigen from adult Haemonchus contortus at prepatency and early and late patency of experimental haemonchosis. Animals that received zero, 1, or 2 infections with the parasite were tested. In addition, sera from 20 experimentally infected and 10 noninfected Texel sheep were used to test the antigen. Sera from 37 infected animals at prepatency as well as at patency in primary and secondary infection were found positive with the 26-kD antigen. However, sera from 10 animals with the lowest worm burdens (second infection) did not recognize the antigen during early patency (day 28 postinfection). IgG1 was the only isotype implicated in antigen recognition because IgG2, IgA, and IgM, in the same sera, showed no reactivity with the peptide. Antigen specificity was confirmed because hyperimmune sera against infective larvae and adult stages of the most common gastrointestinal nematodes found in natural infections in sheep (Trichostrongylus colubriformis and Teladorsagia circumcincta) did not recognize this peptide. The antigen was recognized only by anti-adult H. contortus hyperimmune sera and appeared to be absent in the L3 parasite stage. In addition, the partial N-terminal amino acid sequence of the diagnostic peptide is reported.

Vaccination of Manchego lambs against Haemonchus contortus with a somatic fraction (p26/23) of adult parasites.

A low molecular weight fraction from adult Haemonchus contortus containing two peptides (p26/23) was used to vaccinate Manchego female lambs between 3.5 and 5 months of age. Immunizing injections were given three times on days 0, 14 and 28 of the experiment. On day 43, lambs were challenged with 400 third stage larvae/kg live weight. Vaccination induced a lengthening of prepatent periods, significant reduction (> 60%) in mean faecal egg counts and smaller variations in packed cell volume values. At necropsy, average worm burden in the vaccinated lambs was significantly lower (61.6%) than that found in unvaccinated challenged animals. A clear correlation was found between protection and serum antibody response in immunized lambs.