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PubMed1981

Inheritance of cambendazole resistance in Haemonchus contortus.

Fourth-molt larvae (M4) of cambendazole-resistant (CBZ-R) and cambendazole-susceptible (BPL) strains of Haemonchus contortus were recovered from donor lambs and inoculated orally into source lambs to effect mating in 2nd lamb. Eggs produced by these parasitic matings were collected, cultured, and inoculated into lambs to compare the anthelmintic activity of cambendazole (CBZ), given at a dose rate of 20 mg/kg, against the F1 adults with its activity against the 2 parenteral strains (CBZ-R and PBL). In 2 replications, CBZ was 94% and 99% efficacious against the progeny of the cross matings and 91% and 95% against the BPL strain; CBZ was only 39% and 42% efficacious against the CBZ-R strain. In 1 test of F2 progeny of the reciprocal matings, CBZ was 81.2% efficacious against the F2 adults, whereas CBZ was 97.5% efficacious against the BPL strain and 37% against the CBZ-R strain. The results indicated that heredity of resistance to CBZ in H contortus is not sex-linked and is probably a result of a heterozygous recessive allele. The infective larvae of the CBZ-R groups in these 3 trials were from the 20th and 24th passages through lambs without exposure to any anthelmintics, yet the anthelmintic activity of CBZ ranged from 37% to 42%, indicating that there had been no reversion to susceptibility.

Alleles↗
PubMed1981

Prolonged anthelmintic effect of closantel and disophenol against a thiabendazole selected resistant strain of Haemonchus contortus in sheep.

Two long-acting anthelmintics, closantel and disophenol, were tested by treatment 90, 60 and 30 days before challenge with a thiabendazole selected laboratory strain of Haemonchus contortus. The sheep were slaughtered 28 days after infection for total worm counts. A significant reduction in the number of adults remaining at autopsy was recorded after treatment with both anthelmintics 30 and 60 days before challenge but there was zero efficacy when the animals were treated 90 days before challenge. No significant difference was exhibited between the two compounds. Variability in response was recorded between animals on the 60 day period of treatment, which for disophenol may be dose related because animals receiving the lower volumes showed higher worm counts. For both anthelmintics a variability in the rate of metabolism may be responsible. The possibility offered by both anthelmintics for treating benzimidazole resistant strains of H contortus is discussed.

Animals↗
PubMed1981

Efficiency of oxfendazole administered as a single dose or in a controlled release capsule against benzimidazole-resistant haemonchus contortus, Ostertagia circumcincta and Trichostrongylus colubriformis.

Laboratory strains of Haemonchus contortus and Ostertagia circumcincta selected for thiabendazole resistance were found to have a strong side resistance to a single dose of oxfendazole. The LD50 and LD95 in mg of drug per host liveweight were respectively 4.28 and 18.46 mg/kg for H contortus and 3.61 and 11.20 mg/kg for O circumcincta. A field strain of Trichostrongylus colubriformis that had not been selected with thiabendazole for seven years also had a strong side resistance to oxfendazole with approximately 50 per cent of its population resistant to the recommended dose rate of 5 mg/kg. Prolonged administration of oxfendazole by intraruminal controlled release capsules was found to be effective against both susceptible and resistant strains of the above parasites. The first observed effect of oxfendazole, from controlled release capsules, on resistant worms was a decrease in the percentage of eggs developing to third stage larvae. This was followed by a decrease in egg count and in worm numbers.

Animals↗
PubMed1981

Haemonchus contortus third stage larvae in sheep: kinetics of arrival into the abomasum and transformation during rumino-omasal transit.

The kinetics of passage of Haemonchus contortus third stage larvae through the omasoabomasal orifice were analysed in sheep for six hours after their administration by the nasooesophageal route. The larvae moved into the abomasum more slowly than the liquid phase of the ruminal contents. They arrived in three different forms: sheathed, exsheathed or coiled. A very small number of exsheathed larvae seemed to be necessary for inducing early modifications in permeability of the abomasal mucosa.

Abomasum↗
PubMed1981

Studies on Haemonchus contortus. IV. The effect of Trichostrongylus axei and Ostertagia circumcincta on challenge with H. contortus.

Worm-free Merino yearlings were dosed with either a mixture of infective larvae of Trichostrongylus axei and Ostertagia circumcincta or with O. circumcincta only, and challenged 90-93 days later with infective larvae of Haemonchus contortus. Neither of these methods protected sheep against challenged and slight protection was afforded sheep predosed with T. axei and O. circumcincta and challenged with a trickle dose of H. contortus.

Animals↗
PubMed1980

Studies on Haemonchus contortus. III. Titration of Trichostrongylus axei and explusion of H. contortus.

Groups of Merino weaners were dosed with infective larvae of Trichostrongylus axei in numbers ranging from 20 000-50 000 and challenged 3 months later with 50 000 infective larvae of Haemonchus contortus. When the dose of infective larvae of T. axei was given on Day 0 and the balance on Day + 14, efficacy against H. contortus was > 60% in > 60% of sheep (P < 0, 1). A single dose of 40 000 or 50 000 infective larvae of T. axei was > 80% effective against H. contortus in > 80% of sheep (P < 0, 01). Two doses of 20 000 infective larvae of T. axei followed by a challenge with H. contortus 31-33 days after the initial dose caused a reduction of > 50% in > 50% of sheep (P < 0, 1). This rose to > 60% in > 60% of sheep if the doses of 25 000 infective larvae of T. axei were followed by a challenge with H. contortus 45 days after the initial dose of T. axei. Most of the challenge doses of infective larvae of H. contortus were rejected within 3 days. Surviving worms were retarded in the 4th stage and only a few developed to the 5th or adult stage.

Animals↗
PubMed1980

Haemonchus contortus: attempts to immunise lambs with irradiated larvae.

Two doses of irradiated Haemonchus contortus larvae stimulated firm resistance to challenge in adult worm-free sheep but not in two-month-old lambs. Parenteral administration of larval antigens in adjuvant together with the two doses of irradiated larvae did not reduce the susceptibility of the lambs. However four doses of the attenuated larvae stimulated three of six lambs to resist the challenge. Similar concentrations of serum IgG and mucosal IgA and IgG antibodies were detected in the sheep which had been vaccinated with irradiated larvae only, regardless of their immune status. The lambs which had been injected with worm antigens had much higher levels of circulating and mucosal IgG antibodies, but mucosal IgA concentrations were similar to those in the other immunised groups. Histological examination of the abomasal mucosa did not reveal any differences between resistant and susceptible vaccinates.

Animals↗
PubMed1980

Benzimidazole-resistant Haemonchus contortus--the effect of cryopreservation on the resistance of successive generations.

Infective larvae (L3) of a strain of benzimidazole-resistant Haemonchus contortus were stored in the gas phase of liquid nitrogen, thawed and used to infest worm-free sheep from which, in turn, larvae were cultured, frozen and thawed. This cycle was repeated 5 times. Thereafter, the progeny of the cryopreserved larvae were compared with the progeny of the original untreated larvae for susceptibility to benzimidazole anthelmintics. Repeated freezing of consecutive generations of L3 in liquid nitrogen did not appear to affect their relative resistance to benzimidazoles and, although other strains must also be tested, it would seem that cryopreservation may be used for storing resistant strains.

Animals↗
PubMed1978

Physiological characteristics of free-living and parasitic stages of strains of Haemonchus contortus, susceptible or resistant to benzimidazole anthelmintics.

The effect of changes in genetic constitution (associated with developing benzimidazole resistance) on the physiological characteristics of Haemonchus contortus was investigated. Benzimidazole resistant H contortus were significantly more infective for sheep (20 per cent) than benzimidazole susceptible strains. Pathological changes were more severe in sheep infected with resistant H contortus as measured by packed cell volume, plasma protein concentration and haemoglobin levels. Faecal egg output was significantly higher for resistant strains as was the development and survival of eggs and free-living stages on pasture. Infective third stage larvae of benzimidazole resistant H contortus had a significantly faster exsheathment rate in rumenal fluid than did susceptible strains. These adaptive changes in response to continuing anthelmintic selection of resistant worm populations are discussed and the implications of these findings for worm control programmes indicated.

Animals↗
PubMed1994

Cloning of a putative inhibitory amino acid receptor subunit from the parasitic nematode Haemonchus contortus.

A cDNA, HGl, encoding an inhibitory amino-acid receptor subunit has been cloned from a mixed egg population of the parasitic nematode Haemonchus contortus. The predicted amino-acid sequence of the subunit shows 24% to 32% homology with other vertebrate and invertebrate GABAA and glycine receptor subunits and has all the expected motifs for a member of the ligand-gated ion channel superfamily. When expressed in Xenopus oocytes HGl gives a small response to 1 mM glycine, but not to 1 mM GABA, glutamate, taurine or L-alanine.

Alanine↗
PubMed1995

Developmentally regulated secretion of cathepsin L-like cysteine proteases by Haemonchus contortus.

Cysteine protease activity was present in media collected after 24 hr in vitro culture of adult Haemonchus contortus. The released cysteine protease hydrolyzed the fluorogenic 7-amino-4-trifluoromethyl coumarin (AFC)-substituted synthetic peptides Z-phe-arg-AFC and Z-ala-arg-arg-AFC, but not Z-arg-arg-AFC or Z-arg-AFC, characterizing this activity as cathepsin L-like. Within the parasite, cysteine protease activity was highest in extracts of intestinal tissue. Secreted cysteine protease inhibited the clotting of sheep blood and hydrolyzed hemoglobin, fibrinogen, collagen, and IgG; the IgG hydrolysis site was within the hinge region. Four proteases with M(r) values of 30, 34, 37, and 41 kDa were identified with biotinylated-phenylalanine-arginine-fluoromethyl ketone, a specific probe that binds to active cysteine proteases. Adult parasites cultivated in the presence of 0.1 mM levamisole released 50% less protease activity compared to control cultures; in the presence of rafoxanide (0.1 mM), protease was not detected. Cathepsin L-like cysteine protease activity was released also by L4, but not the L3 larval stage. The active and developmentally regulated release of cysteine proteases by H. contortus may have a critical function in worm nutrition, immune evasion, or both.

Amino Acid Sequence↗
PubMed1993

Protective immunity to Haemonchus contortus induced by immunoaffinity isolated antigens that share a phylogenetically conserved carbohydrate gut surface epitope.

Whole gut homogenates of the blood-sucking nematode Haemonchus contortus induce protective immunity in goats, and some of these gut Ag are conserved among related parasitic nematode species. To identify gut Ag that induce protective immunity and have phylogenetically conserved epitopes, mAb were made to gut-surface Ag of H. contortus. Forty-nine mAb reacted with microvilli of the parasite gut. Two of these mAb (42/10.6.1 and 42/53.3.5) were analyzed here. Both of the mAb bound to the microvillar surface of freshly isolated gut, and each mAb recognized carbohydrate epitopes, based on sensitivity to periodate oxidation. The 42/10.6.1 epitope occurred on at least 18 proteins in Western blots and in several H. contortus tissues. Proteins recognized by this mAb localized to membrane and excretory/secretory fractions of the worm. This epitope was also identified on the gut and other tissues and multiple proteins of related adult and larval nematodes, including larval Ancylostoma caninum and a mixed population of the free-living nematode Caenorhabditis elegans. In contrast, the 42/53.3.5 mAb bound to the gut surface and recognized proteins of 100 and 46 kDa from adult H. contortus gut. Four proteins of 100, 52, 46, and 30 kDa were isolated from the 42/53.3.5 immunoaffinity columns, and except for the 30-kDa protein, each was recognized by both the 42/10.6.1 and 42/53.3.5 mAb. Epitopes recognized by each mAb were distinct from one another and phosphorylcholine. When used to immunize goats, Ag isolated by both mAb induced protection that significantly (p < 0.05) reduced total worm counts after challenge infections compared with the control groups.

Animals↗
PubMed1995

High energy phosphate metabolites observed by NMR in infective larvae of Haemonchus contortus.

Phosphorus resonances consistent with phosphoarginine and ATP were observed in the in vivo 31P-nuclear magnetic resonance (NMR) spectra of infective larvae of Haemonchus contortus. The level of phosphoarginine quickly declined when nematode suspensions were purged with nitrogen and was restored upon return to aerobic conditions. Saturation transfer NMR demonstrated forward and reverse exchange of phosphorus between phosphoarginine and ATP.

Adenosine Triphosphate↗
PubMed1995

Characterization of local antibody responses to the gastrointestinal parasite Haemonchus contortus.

The ability to identify antigens associated with an infection has generally relied on the use of serum antibodies produced by infected or previously exposed individuals. A major drawback with the use of serum is that it does not necessarily reflect the local antibody response at mucosal tissue sites. This study describes an approach that allows the use of antibodies generated close to the infection site to detect the transient expression of stage-specific antigens during infection with the gastrointestinal parasite Haemonchus contortus. This was achieved by infecting immune sheep with H. contortus larvae and removing the abomasal lymph nodes draining the infection site shortly after the challenge infection. Antibody-secreting cell (ASC) probes were generated from these lymph nodes after short-term in vitro culture of cell suspensions, which allowed the accumulation of antibodies secreted by in vivo-induced ASC into the culture supernatant. Lymph node culture supernatants (= ASC probes) from immune sheep challenged 5 days previously were used to probe Western blots of third and fourth stage larval preparations, and revealed distinct reactivity to larval antigens. No antibody reactivity to larval antigen preparations was detected in sheep that were not challenged. The number of antigens identified using ASC probes was significantly restricted compared to either pre- or post-challenge sera. In contrast to the variability of the serum response, the specificity of ASC probes was highly repeatable between different sheep. ASC probes were also used to purify a H. contortus larval antigen by affinity chromatography, which allowed limited biochemical studies to be undertaken. The antigen(s) recognized by the ASC probes were shown to be expressed on the surface of the larvae. These studies illustrate the use of a novel means of studying the local antibody response close to a mucosal infection site in order to identify and isolate stage-specific antigens expressed during infection.

Animals↗
PubMed1994

Ivermectin binding sites in sensitive and resistant Haemonchus contortus.

Membranes from both ivermectin-sensitive and -resistant Haemonchus contortus L3 larvae were examined for the presence of high affinity [3H]ivermectin binding sites. Both tissue preparations displayed high affinity drug binding sites (Kd = 0.13 nM). Receptor density (Bmax = 0.4 pmol/mg) was the same in both the sensitive and resistant nematodes suggesting that target site modification was not involved in the development of drug resistance in this particular strain of H. contortus. The H. contortus ivermectin binding site appeared to be similar to the well characterized Caenorhabditis elegans ivermectin binding site with respect to affinity for ivermectin and receptor density.

Animals↗
PubMed1993

Comparison of the effects on milk production of concurrent infection with Haemonchus contortus and Trichostrongylus colubriformis in high- and low-producing dairy goats.

The impact of nematode parasitism of the digestive tract on milk output and milk quality was examined in dairy goats. In addition, the consequences of worm infection were compared in goats with different lactation performance (ie, with initial high or low milk production). Forty-eight goats in the second month of lactation were allotted equally to 2 groups. The first group was given 5,000 Haemonchus contortus and 20,000 Trichostrongylus colubriformis larvae. The 24 additional goats remained free of parasites. Parasitologic, serologic, and milk data were collected every 2 weeks for 5 months, and body condition of the goats was scored throughout the study. Results of strongyle egg count in feces, increase in pepsinogen values, and reduction in RBC count, PCV, and serum inorganic phosphate concentration indicated subclinical infection. This subclinical parasitism induced a decrease in body condition scoring and led to persistent decrease in milk yield, ranging from 2.5 to 10% reduction from control values. Changes in fat and protein contents were not detected. In contrast, the consequences of infection were more severe in the 6 goats with the highest milk production at the start of the study. Decrease in milk output ranged between 13.0 to 25.1%, and was associated with decrease in fat content. Comparison of the response to parasitism in the 6 goats with the highest lactation performance and the 6 goats with the lowest performance indicated differences between both subgroups. According to parasitologic and pathologic data, high-producer goats had less resistance and/or resilience to infection associated with more severe consequences on milk production.

Animals↗
PubMed1993

The pattern of faecal egg output in lambs infected with a multiple resistant strain of Haemonchus contortus after treatment with albendazole.

Worm-free lambs (n = 6) were infected with 5,000 L3 of an ivermectin/benzimidazole/closantel resistant strain of Haemonchus contortus. On Day 21 post infection animals were allocated, according to worm egg counts, to 2 groups of 3 animals each and on Day 29 one group was treated with albendazole at 3.8 mg kg-1, while animals in the other group remained untreated as controls. All animals were slaughtered on Day 44 when the treated group was found to have 2,083 +/- 1,718 worms and the controls 2,783 +/- 633, a reduction of 25.2% in worm burdens. The pattern of egg output was monitored and it was found that 2 d after dosing, worm egg counts had been suppressed by 89% in the treated group; thereafter the egg counts increased, but never returned to the pretreatment levels and by Day 15 post treatment, they were still 41.1% lower in the treated group than in the controls.

Albendazole↗
PubMed1993

Selection for a strain of Haemonchus contortus that exhibits periparturient egg rise in sheep.

Periparturient egg rise (PPR) is a principal means for some nematode parasites to survive over winter and to provide transmission of infective larvae from ewes to lambs during the spring. Routine laboratory propagation techniques probably have selected for those characteristics that would not promote PPR in conventional laboratory strains of Haemonchus contortus. An established isolate of H. contortus (BPL) was the source of the initial inoculum to select for a strain that readily exhibits characteristics of PPR. The selection process included inoculation of helminth-free pregnant ewes during mid-gestation, collection of nematode eggs during lactation, and storage of infective larvae at 4 C for 4 mo. After 10 generations, the 2 strains, BPL and PPR from lambs and pregnant ewes were compared for reproductive, morphological, and population differences in lambs and pregnant ewes. After lambing, ewes inoculated with the PPR strain had significantly higher fecal egg concentrations. Lambs inoculated with the PPR strain had higher egg concentrations, higher total daily egg production, fewer adult worms, larger female worms, and higher fecundity. Repeated selection in the appropriate host, after prolonged storage of the inoculum, produced a PPR strain of H. contortus morphologically and reproductively distinct from the initial source isolate.

Animals↗