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PubMed1994

New morphological characters for identifying individual specimens of Haemonchus spp. (Nematoda: Trichostrongyloidea) and a key to species in ruminants of North America.

The large stomach worms Haemonchus contortus, Haemonchus placei, and Haemonchus similis are important pathogens of cattle and sheep. This paper describes characteristics of surface cuticular ridges (synlophe), which for the first time provide morphological criteria for identifying individual adult specimens of either sex. The diagnostic patterns of the synlophe on the anterior half of specimens can be observed at 400x in temporary mounts on glass slides. The synlophe can be studied in cleared preserved specimens or in living or freshly thawed frozen specimens mounted in water. The synlophe of H. contortus has 30 ridges in the region of the posterior half of the esophagus, 4 fewer than H. placei and H. similis. The 4 extra ridges of H. placei and H. similis are consistently located bilaterally to the 3 ventralmost and the 3 dorsalmost ridges. The 4 extra ridges of H. similis extend to the end of the synlophe posterior to midbody, but in H. placei they extend posteriorly only to the end of the anterior quarter of the nematode. A key is included to the 3 species of Haemonchus parasitic in domestic sheep and cattle using characteristics of spicules, female reproductive system, female tail, and the synlophe.

Animals↗

PubMed1999

Description of Haemonchus placei (Place, 1893) (Nematoda, Trichostrongylidae, Haemonchinae), identification and intra-specific morphologic variability.

Haemonchus placei in cattle has never been completely described, possibly due to great morphological similarity with small ruminants Haemonchus contortus. It is newly described on one isolate from Argentina. It has clear distinct morphological features from sheep and goats Haemonchus contortus and presents only two female morphotypes (linguiform and knobbed) instead of three recorded in H. contortus. A key is proposed to identify females. Female as well as male Haemonchus placei from New World (Argentina, Mexico, USA) are morphologically different from those of Old World (Africa: Burkina-Faso, Mauritania and Ivory Coast) or Australia, possibly due to local evolution since their introduction several centuries ago from Africa or India. We propose to differentiate three sub-species, H. placei placei in Australia, H. placei africanus in western Africa and H. placei argentinensis in the New World.

Animals↗

PubMed1994

Cloning and characterization of ribosomal RNA genes from three species of Haemonchus (Nematoda: Trichostrongyloidea) and identification of PCR primers for rapid differentiation.

Genomic DNAs prepared from adult worms of Haemonchus contortus, Haemonchus placei, and Haemonchus similis were used to clone and map complete ribosomal RNA gene repeats from each species. The lengths of the repeating units were estimated at 7.2, 7.2, and 7.4 kb, respectively, with a second and distinct repeat 6.9 kb in length identified from H. contortus only. Restriction enzyme recognition sites within all ribosomal RNA genes were fully conserved except for the loss of a SalI site within the large subunit rRNA gene of the 6.9-kb H. contortus repeat where only minor differences were observed in the restriction enzyme recognition sequences within the external spacer DNAs. Sequence analysis of the subcloned small subunit ribosomal DNAs from each species demonstrated 100% conservation within the 1758-bp fragments with only limited sequence variability observed within the adjacent 5' external-transcribed spacer. Enzymatic amplification of external spacer sequences using primers complementary and proximal to the 3'-end of the large subunit and the 5'-end of the small subunit rDNAs enabled rapid differentiation of individual worms of H. contortus from H. placei by utilizing the size variability within this region of the repeat.

Animals↗

PubMed2000

Immunisation of lambs with drug-abbreviated Haemonchus contortus infections: protection against homologous and heterologous challenge.

Haemonchus contortus is known as a prolific parasite, producing high numbers of eggs. It could therefore be used as a cheap source of larvae for immunisation of lambs. The value of immunisation would be improved if the immunity produced gave protection against not only homologous but also heterologous infections. Because antibody cross-reactivity between Haemonchus and Ostertagia has been well established, we wanted to know whether drug-abbreviated infections of H. contortus would stimulate production of antibodies that would cross-react with Trichostrongylus colubriformis. The results obtained from these trials indicated that H. contortus drug-abbreviated infections produced significant immunity against not only Haemonchus but also O. circumcincta. Immunisation increased the level of immunoglobulin E (IgE) and IgE-specific antibodies against T. colubriformis, but the differences between experimental and control animals were not statistically significant. Significantly higher levels of IgG-specific antibodies against T. colubriformis were observed.

Animals↗

PubMed1995

Protection against multiply drug-resistant and geographically distant strains of Haemonchus contortus by vaccination with H11, a gut membrane-derived protective antigen.

H11 is a "hidden antigen" derived from adult Haemonchus contortus which is capable of conferring a high degree of protection against subsequent challenge. To be commercially useful, any vaccine based on this antigen must protect against field strains and drug-resistant worms. The results show that vaccination with H11 affords high levels of protection against the multiply drug-resistant (MDR) Lawes strain of Haemonchus contortus. Moreover, there was no difference in protection whether the H11 was extracted from drug-resistant or susceptible worms, indicating that multiple drug-resistance does not significantly alter the immunogenicity of H11. Ideally, a commercial vaccine based on this antigen should also be capable of protecting against strains from geographically separate areas, so that it is not necessary to manufacture "region-specific" vaccines. Vaccination with H11 purified from either Australian or U.K. Haemonchus contortus was effective in protecting against subsequent challenge with Australian larvae, indicating that one type of H11 should have world-wide application.

Animals↗

PubMed1992

Efficacy of moxidectin against an ivermectin-resistant strain of Haemonchus contortus in sheep.

The efficacy of moxidectin was determined against ivermectin-susceptible and resistant strains of Haemonchus contortus. At the onset of the trial, 40 lambs were each infected with 5000 third stage larvae of one of two strains of Haemonchus contortus. The lambs were randomly sorted into eight treatment groups 28 days post-infection and were treated as follows: Group 1, susceptible strain with no treatment; Group 2, resistant strain with no treatment; Group 3, susceptible strain treated with 0.2 mg moxidectin kg-1 body weight; Group 4, resistant strain treated with 0.2 mg moxidectin kg-1; Group 5, resistant strain treated with 0.4 mg moxidectin kg-1; Group 6, susceptible strain treated with 0.2 mg ivermectin kg-1; Group 7, resistant strain treated with 0.4 mg ivermectin kg-1; Group 8, resistant strain treated with 0.8 mg ivermectin kg-1. The lambs were killed 1 week post-treatment. Comparisons were made among groups based on the number of eggs per gram of feces on the day of treatment and the numbers of worms recovered from each lamb. Both moxidectin and ivermectin were effective in removing susceptible Haemonchus with efficacies of 100% and 99.7%, respectively. The efficacy of moxidectin against the resistant strain was 99.9% and 100% at 0.2 mg kg-1 and 0.4 mg kg-1, respectively, whereas there were only 38.8% and 53.1% efficacies in the lambs treated with 0.4 mg ivermectin kg-1 and 0.8 mg kg-1 body weight, respectively.

Animals↗

PubMed2006

In vitro and in vivo anthelmintic activity of crude extracts of Coriandrum sativum against Haemonchus contortus.

In vitro anthelmintic activities of crude aqueous and hydro-alcoholic extracts of the seeds of Coriandrum sativum (Apiaceae) were investigated on the egg and adult nematode parasite Haemonchus contortus. The aqueous extract of Coriandrum sativum was also investigated for in vivo anthelmintic activity in sheep infected with Haemonchus contortus. Both extract types of Coriandrum sativum inhibited hatching of eggs completely at a concentration less than 0.5 mg/ml. ED(50) of aqueous extract of Coriandrum sativum was 0.12 mg/ml while that of hydro-alcoholic extract was 0.18 mg/ml. There was no statistically significant difference between aqueous and hydro-alcoholic extracts (p>0.05). The hydro-alcoholic extract showed better in vitro activity against adult parasites than the aqueous one. For the in vivo study, 24 sheep artificially infected with Haemonchus contortus were randomly divided into four groups of six animals each. The first two groups were treated with crude aqueous extract of Coriandrum sativum at 0.45 and 0.9 g/kg dose levels, the third group with albendazole at 3.8 mg/kg and the last group was left untreated. Efficacy was tested by faecal egg count reduction (FECR) and total worm count reduction (TWCR). On day 2 post treatment, significant FECR was detected in groups treated with higher dose of Coriandrum sativum (p<0.05) and albendazole (p<0.001). On days 7 and 14 post treatment, significant FECR was not detected for both doses of Coriandrum sativum (p>0.05). Significant (p<0.05) TWCR was detected only for higher dose of Coriandrum sativum compared to the untreated group. Reduction in male worms was higher than female worms. Treatment with both doses of Coriandrum sativum did not help the animals improve or maintain their PCV while those treated with albendazole showed significant increase in PCV (p<0.05).

Albendazole↗

PubMed2001

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-dependent 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.

Amitrole↗

PubMed2003

Pyrosequencing analysis identifies discrete populations of Haemonchus contortus from small ruminants.

The genus Haemonchus consists of blood-sucking parasitic nematodes in the abomasum of ruminants. Members of this genus are responsible for extensive production losses, particularly of small ruminants in the tropics but are also found in temperate regions. In this study, we examined the internal transcribed spacers-1 and -2 of rRNA in Haemonchus spp. The rRNA region spanning the internal transcribed spacers-1, -2 and the 5.8S rRNA gene was amplified by PCR from each of 10 worms from Swedish sheep, a Swedish goat and Kenyan sheep. The fragments were sequenced and examined with respect to genetic differences fixed among the three isolates. These and additional worms were further analysed with Pyrosequencing technology. This technique allowed us to rapidly analyse 110 individuals in three putative polymorphic nucleotide positions that were initially identified with dideoxy sequencing. The geographical isolates could to a large extent be genetically distinguished, but none of the polymorphic positions were consistent among all individuals within each isolate. Furthermore an alignment of our sequences and a consensus sequence published for Haemonchus contortus revealed two differences in positions 123 and 196 in internal transcribed spacers-2. Although these positions were previously reported as heterogenic, no polymorphism was detected among the 30 worms sequenced in the present study. Modelling of the internal transcribed spacers-2 secondary structure based on our data also identified a new putative long-range interaction. The isolates are best described as populations. In conclusion, consistent differences were not identified and the studied isolates are therefore best described as discrete populations. This study also reveals for the first time the potential of Pyrosequencing technology as a tool in the analysis of nematode population genetics.

Abomasum↗

PubMed1994

Immune responses of Texel sheep to excretory/secretory products of adult Haemonchus contortus.

The excretory/secretory (E/S) products of adult Haemonchus contortus comprise of at least 15 polypeptides with molecular weights ranging from 10 to > 100 kDa. These E/S products induce an immune response in infected Texel sheep, as demonstrated by specific IgG1 levels and a significant lymphocyte proliferation index. Moreover, immunoblotting analysis revealed that sera of primary H. contortus-infected sheep specifically recognize a 24 kDa E/S product. In addition, sera of challenged sheep react strongly with a 15 kDa E/S product. The other E/S products of H. contortus showed immunoreactivity with serum samples of Haemonchus-infected sheep as well as with samples of sheep harbouring other trichostrongylid infections. These cross-reacting epitopes are the main cause of the lack of specificity of an E/S material-based ELISA. This ELISA can differentiate Haemonchus infections from Nematodirus battus infections, but not from Ostertagia circumcincta or Trichostrongylus colubriformis infections.

Animals↗

PubMed2000

Evaluation of immunization with gut membrane glycoproteins of Ostertagia ostertagi against homologous challenge in calves and against Haemonchus contortus in sheep.

Peanut and ConA lectins were used as ligands to isolate glycoproteins from detergent extracts of adult Ostertagia ostertagi membranes. As judged by their profiles following SDS-PAGE, these fractions closely resembled the equivalents from Haemonchus contortus which are derived from the nematode intestinal cell microvillar membranes and which are highly protective when used as antigens. Groups of calves were immunized with the peanut and ConA binding fractions of Ostertagia, either as separate or pooled antigens mixed with QuilA as adjuvant. All calves, including controls immunized with adjuvant only, were challenged with a single dose of infective Ostertagia larvae and faecal egg counts were monitored for 5 weeks. In two experiments where the antigen fractions were pooled, moderate (30-50%), but statistically significant reductions in egg output were observed, but the number of worms was not diminished. No significant protection was observed in a third trial where groups of calves were immunized with peanut or ConA binding proteins given separately. Two further trials were conducted in sheep immunized with the same Ostertagia fractions but challenged with Haemonchus. Irrespective of whether they were administered separately or together, the Ostertagia antigens cross protected efficiently against Haemonchus reducing egg counts by between 81% and 97% and worm numbers by between 57% and 84%.

Adjuvants, Immunologic↗

PubMed1993

Isotype-specific antibody responses to Haemonchus contortus in genetically resistant sheep.

The kinetics of anti-Haemonchus antibody responses in serum and faecal extracts of pasture-reared, genetically resistant and random-bred sheep infected with Haemonchus contortus were examined using an isotype-specific ELISA. Anti-Haemonchus antibodies of IgA, IgG1, IgG2 and IgM isotypes were detected in serum and faecal extracts of both resistant and random-bred sheep after challenge infection. Serum IgG1 and IgA levels in resistant sheep were significantly higher than in random-bred sheep between 10 and 31 days after infection. However, there were no differences in IgG2 and IgM antibody responses between the two genotypes. Faecal antibody responses to H. contortus showed a clear genetic effect with resistant sheep exhibiting higher IgA levels throughout infection and higher IgG1 levels between 24 and 31 days after infection. Furthermore, serum IgG1 and IgA, and faecal IgA responses were negatively correlated with faecal egg counts in both genotypes on 17, 24 and 31 days after infection. Together, these results are taken to indicate that anti-parasite IgA and IgG1 antibodies may play an important role in genetically determined resistance of sheep to haemonchosis.

Analysis of Variance↗

PubMed1994

Protection studies with integral membrane fractions of Haemonchus contortus.

Techniques for targetting glycoproteins integral to the luminal membrane of the intestinal cells of Haemonchus contortus were used to isolate fractions of whole parasites with protective antigen potential. Sheep immunization trials with various candidate fractions revealed one which selectively bound to lectins with specificity for N-acetylgalactosamine and which reduced mean challenge worm burdens by up to 72% and mean faecal egg counts by up to 93%. The luminal surface of the intestines of the Haemonchus recovered from sheep immunized with this antigen were coated with host immunoglobulin, suggesting that the protective effect was due to antibodies interfering with the function of the gut. Further biochemical characterization of this fraction, which has been termed Haemonchus galactose-containing glycoprotein complex (H-gal-GP complex), showed that it could be distinguished from previously described protective antigens of this parasite and that it was only detectable in detergent extracts of the worms.

Acetylgalactosamine↗

PubMed1999

Variation in resistance to haemonchosis: selection of female sheep resistant to Haemonchus contortus.

Seventy female lambs (6-7 months old) which were exposed to natural infections of Haemonchus contortus were designated as responders or non-responders on the basis of 10 weekly cumulative faecal egg counts. Selected responder and non-responder lambs were treated with ivermectin, housed separately and 6 weeks post-housing, seven lambs from each group were given a trickle infection of Haemonchus contortus at 1000 L3 daily for 5 days per week up to 2 weeks and examined weekly for 10 weeks after first infection. Analysis of data revealed significantly lower mean faecal egg counts and non-significantly less weight loss in responder than non-responder lambs. Mean values of haemoglobin, packed cell volume, total serum protein and peripheral eosinophil counts were significantly higher in responders than non-responders. In contrast, serum pepsinogen concentration was significantly less in responders than in non-responders. At 10 weeks post-infection, there were fewer pathological lesions and significantly lower worm burdens in responders than in non-responders. These results demonstrate a distinct resistance in responders to Haemonchus contortus infection.

Animals↗

PubMed1987

Seasonal changes in the prevalence of Haemonchus and Trichostrongylus hypobiotic larvae in tracer goats in Malaysia.

An investigation into the seasonal changes in the population structure of Haemonchus and Trichostrongylus in tracer goats was conducted over 12 months at Serdang, an area in southern West Malaysia with year-round tropical rainfall. Successive groups of parasite-free tracer goats were grazed for a month alongside naturally infected adult goats and necropsied for worm counts 14 days after their removal from pasture. No hypobiotic larvae of Trichostrongylus were recovered. Hypobiotic larvae of Haemonchus were evident during each month of the year but accounted for only a very small proportion of the total Haemonchus burden. Very low levels were encountered from December through to June. Comparatively higher levels of hypobiosis were observed thereafter with a peak of 7.4% in September. The factors responsible for hypobiosis were not clearly defined but the phenomenon was associated with increasing levels of soil moisture storage. Host resistance, adult worm population of tracers and population of ingested L3 were ruled out as possible inducing factors.

Animals↗

PubMed1990

Seasonal changes and hypobiosis in Haemonchus contortus infection in the West African Dwarf sheep and goats in the Nigerian derived savanna.

A survey was conducted in the Nigerian derived savanna (from August 1987 to July 1988) on the seasonal fluctuations in the composition of Haemonchus contortus burden of naturally infected West African Dwarf sheep and goats. During this period 117 abomasa collected from locally slaughtered animals were processed. Male, female as well as immature worms were enumerated. The incidence of Haemonchus infection was high (77.8-100%) with no definite seasonal distinction. There was however, a clear seasonal trend in the worm burden of animals; higher burdens were evident during the rainy rather than the dry season. There was no significant hypobiosis; Haemonchus contortus survived in the host during the unfavourable dry season (November to March) as adults.

Abomasum↗

PubMed1999

Further immunization and biochemical studies with a protective antigen complex from the microvillar membrane of the intestine of Haemonchus contortus.

Immunization of sheep with a protective antigen complex from the intestinal cells of Haemonchus contortus in Freund's adjuvant stimulated individually variable antibody responses but still conferred significant protection against parasite infection. Correlation between antibody concentration and degree of protection was suggestive of antibody being the effector mechanism. The antigen is known as Haemonchus galactose-containing glycoprotein complex (H-gal-GP) because it binds to lectins with a specificity for N-acetyl-galactosamine. Polypeptide composition analysis by polyacrylamide gel electrophoresis indicated an apparent molecular weight of about 1000 kDa and SDS gels revealed four major polypeptides, containing between 2 and 5 disulphide linked subunits, nearly all being glycosylated. N-terminal amino acid sequence was obtained from 12 subunits, ten showing homologies with cDNAs from Haemonchus encoding either pepsin, metalloprotease or cysteine protease-like enzymes. pH optima, inhibitor and various substrate studies confirmed that the native complex possessed proteolytic activities in agreement with the sequence data. Although the cDNAs predicted water soluble enzymes, little of the complex was solubilized from worm membranes without the use of a detergent, such as Triton X-100. It is hypothesized that H-gal-GP is a gut membrane associated multiprotease complex which is involved in the digestion of the blood meal and which can be neutralized by specific antibodies with drastic consequences for the parasite.

Amino Acid Sequence↗

PubMed1992

The influence of challenge dose, duration of immunity, or steroid treatment on mucosal mast cells and on the distribution of sheep mast cell proteinase in Haemonchus-infected sheep.

The distribution of granule-specific sheep mast cell proteinase (SMCP), was assayed by immunocytochemistry and quantified by immunoassay in sheep immune to Haemonchus contortus. Repeated infection with Haemonchus larvae over 10-12 weeks induced a pronounced mucosal mastocytosis, including intraepithelial globule leukocytes (GL), which, 7 days after ceasing this dosing regime, was associated with the inability of incoming larvae to establish within the abomasal mucosa. Loss of this resistance, due to the cessation of stimulation with Haemonchus larvae 84 days previously or to treatment of sheep with corticosteroid, was associated with a marked decline in mast cell density and concentrations of SMCP in abomasal mucosal tissues. Nevertheless, larvae also failed to establish in immune sheep rested from challenge 42 days previously and in which mast cell counts were not significantly different from those of control sheep. A small, but significant, release of SMCP was demonstrated in gastric mucus from immune sheep following larval challenge, whereas little or no SMCP was detected in mucus from naïve animals.

Abomasum↗