Haemonchus contortus: Local and serum antibodies in sheep immunised with irradiated larvae

Smith W. D. and Christie M. G. 1978. Haemonchus contortus: lccal and serum antibodies in sheep immunised with irradiated larvae. International Journal for Parasitology8: 219–223. Two doses of Co⁶⁰ irradiated Haemonchus contortus protected 9-month-old sheep against a challenge infection of 10,000 normal larvae. This resistance was associated with increased concentrations of IgG antibodies in the serum as well as IgA and IgG antibodies in the abomasal mucosa.     

Effect of Selenium Yeast Supplementation on Naturally Acquired Parasitic Infection in Ewes

Gastrointestinal parasites cause substantial economic losses in pasture-based sheep production systems. Supranutritional organic selenium (Se) supplementation may be beneficial because it improves immune responses to pathogens. To evaluate the effect of Se-yeast supplementation on gastrointestinal parasite load, 30 ewes per treatment group were drenched weekly with no Se, 4.9 mg Se/week as Se yeast (maximum FDA-allowed concentration), or supranutritional concentrations of Se yeast (14.7 and 24.5 mg Se/week) starting early fall for 85 weeks. Fecal samples were collected at weeks 63, 66, 78, and 84 and counted for total trichostrongyle-type eggs and Haemonchus contortus eggs (in samples with ≥200 trichostrongyle eggs/g feces). During breeding season (fall), ewes were kept on pasture; ewes receiving 24.5 mg Se/week had lower fecal trichostrongyle egg counts (93?±?40 eggs/g feces) compared with ewes receiving no Se (537?±?257 eggs/g feces; P?=?0.007) or ewes receiving 4.9 mg Se/week as Se yeast (398?±?208 eggs/g feces; P?=?0.03). In winter, fecal trichostrongyle egg counts decreased, and group differences were not apparent. During lambing season (spring), ewes were kept in the barn and fecal trichostrongyle egg counts increased, although no group differences were observed. However, none of the ewes receiving supranutritional Se yeast, and with trichostrongyle egg counts ≥200 eggs/g of feces, but four of the ewes receiving lower Se dosages had H. contortus egg counts ≥1,000 eggs/g feces (P?=?0.04). Our results suggest that supranutritional Se-yeast supplementation may enhance resistance to naturally occurring H. contortus gastrointestinal parasitism in sheep.     

The mechanism of hatching of eggs of Haemonchus contortus

Fluid collected from hatching eggs of Haemonchus contortus contained a lipase which hydrolysed 2-naphthyl laurate (about 0·7 μmol naphthol freed /h/10⁶ eggs). The fluid also hydrolysed l-leucinamide (about 2·3 μmol leucine freed/h/10⁶ eggs). The fluid when added to normal or heated eggs caused ‘hatching’. ‘Hatching’ also occurred in exsheathing fluid from infective juveniles and in a preparation of pancreatic lipase containing leucine aminopeptidase. A purified mammalian leucine aminopeptidase in combination with several different lipases did not attack egg shells.The ‘spontaneous’ hatching of eggs of H. contortus was strongly inhibited by 1,10-phenanthroline, 10^?3M, and this inhibition was reversed by Zn²⁺. However, the inhibition of ‘hatching’ of eggs in externally applied hatching fluid, or the hydrolysis of leucinamide in hatching fluid was generally less marked.     

Time of onset and target of immune reactions in sheep with acquired immunity against Haemonchus contortus

Adams D. B. 1982. Time of onset and target of immune reactions in sheep with acquired immunity against Haemonchus contortus. International Journal for Parasitology12: 439–443. Nonimmune sheep and sheep rendered immune by infection with the abomasal nematode, Haemonchus contortus, were infected with the parasite and treated at various times with the glucocorticosteroid, dexamethasone. The results show that this immunosuppressant drug abolished acquired but not innate immunity to H. contortus and that acquired responses were not important in restraining the fecundity of adult worms during primary infection. By treating immune sheep with dexamethasone during reinfection, it was shown that the responses acting against the establishment of infection commence later than the fourth day after larval administration and are complete by the seventh day. H. contortus more advanced in development than the fourth-larval stage were relatively insusceptible to this manifestation of acquired immunity.     

The response of sheep to parenteral vaccination and immunizing infections against the abomasal nematode, Haemonchus contortus

Adams D. B., Beh K. J. and Davies H. I. 1982. The response of sheep to parenteral vaccination and immunizing infections against the abomasal nematode, Haemonchus contortus. International Journal for Parasitology12: 445–449. Subcutaneous injection of relatively large amounts of unfractionated homogenates of adults plus infective larvae of Haemonchus contortus emulsified in complete Freund's adjuvant produced a degree of protective immunity when challenge infection was given eight weeks after the first or only dose of vaccine. In an attempt to improve acquired immunity, parenteral vaccination was either followed or preceded by a short immunizing infection with H. contortus, which was terminated by anthelmintic before patency. This treatment aimed at stimulating general responsiveness to worm antigens and invoking mucosal immunity in the abomasum. Disparate results were obtained; immunizing infections either increased immunity or made sheep more susceptible to challenge infection. In this latter situation, the unresponsiveness associated with primary infection with H. contortus may have been increased.     

Immunity acquired by sheep from an experimental infection with haemonchus contortus

Adams D.B. and Beh K.J. 1981. Immunity acquired by sheep from an experimental infection with Haemonchus contortus. International Journal for Parasitology11: 381–386. A primary infection of sheep with a single dose of Haemonchus contortus larvae was traced by faecal egg counts until it had substantially declined after 55 weeks. These primed sheep were then given a sequence of two reinfections with the parasite. Comparison of faecal egg counts in primed sheep and in two separate groups of previously worm-free sheep showed that primary infection conferred significant immunity. This, however, was not sufficiently protective to prevent the development of further anaemia and faecal egg counts indicative of clinical haemonchosis. It is suggested that an adaptation in the host-parasite relationship which promotes the longevity of primary infection with H. contortus may also moderate the induction of acquired immunity.The titre of haemagglutinating antibody specific for H. contortus rose in serum during the course of primary infection, but the two reinfections did not stimulate a rise in titre. Titres of haemagglutinating antibody before reinfection did not correlate with subsequent faecal egg counts.     

Organic and inorganic acids as the stimulus for exsheathment of infective juveniles of nematodes

Petronijevic T., Rogers W. P. and Sommerville R. I. 1986. Organic and inorganic acids as the stimulus for exsheathment of infective juveniles of nematodes. International Journal for Parasitology 16: 163–168. A variety of organic and inorganic acids stimulated exsheathment of infective juveniles of Nematospiroides dubius over the range pH 2–7. Activity, in relation to pK values, suggests that the undissociated form of the acid was the active agent. Under normal conditions of exposure, exsheathment of infective juveniles of Haemonchus contortus was not obtained below pH 6, and at higher pH values activity was low except in the presence of CO₂ at pressures >9-5 kPa in the gas phase. Brief exposure to HCl at concentrations up to 2N did not induce exsheathment but in the presence of CO₂ in the gas phase, 95.3 kPa, activity was obtained in 0-O1N HCl when the times of exposure were 3–25 min.The differences in the responses of N. dubius and H. contortus to a stimulus at low pH values may be attributed to the stability of the exsheathing enzyme of N. dubius in acid media as well as to the relative sensitivity of the two species to stimuli generally.     

Optimum temperature for egg development of phenotypes in Haemonchus contortus cayugensis as determined by Arrhenius diagrams and Sacher's entropy function

Lejambre L. P. and Whitlock J. H. 1973. Optimum temperature for egg development of phenotypes in Haemonchus contortus cayugensis as determined by Arrhenius diagrams and Sacher's entropy function. International Journal for Parasitology3: 299–310. Oxygen uptake, maturation time and percentage hatch were determined on eggs from “wild-type” Haemonchus contortus cayugensis populations as well as those from the morph types smooth, linguiform B and linguiform A. Sacher's organizational entropy, which is essentially a sum of oxygen utilized by a system corrected for the number of viable units in the system, was used to determine the optimum temperature for development of the eggs. Although the eggs from “wild-type” H. contortus cayugensis hatched well across a broad temperature range; individual phenotypes showed a much narrower range. The optimum temperature for the development of eggs from smooth worms was approximately 30°C while linguiform A and B hatched best at 25°C. Linguiform A eggs had a broader temperature tolerance than linguiform B so that at temperatures of 17°C they hatched with a greater efficiency than did either smooth or linguiform B. It is argued that H. contortus follows a “strategy” of having many phenotypes, each one of which appears to be adapted to a different temperature. This would allow a population to maintain a broad range of optimal temperatures without the expense of maintaining the cybernetic machinery which a single individual would have to have if it were to tolerate the same range of temperature.     

Cimetidine inhibits the pentagastrin-induced release of calcitonin in normocalcaemic man

Pentagastrin stimulates the release of calcitonin from normal C-cells in the human thyroid. In the present investigation the effect of cimetidine on the liberation of calcitonin in response to intraarterial pentagastrin (0.6 μg · kg^?1) was studied in 14 normocalcaemic patients undergoing surgery for thyroid adenomas. Cimetidine was administered as a bolus injection of 200 mg followed by an intravenous infusion of 1.5 mg · kg^?1 · h^?1. In seven patients not given cimetidine, mean calcitonin concentration in the thyroid vein rose from 419 ± 58 to 2787 ± 645 pM in response to pentagastrin. In seven patients given cimetidine, mean calcitonin concentration only increased from 107 ± 33 to 166 ± 51 pM after pentagastrin. The difference between the two groups was statistically significant both during basal conditions (P < 0.001) and in response to pentagastrin (P < 0.01). The results suggest that pentagastrin affects normal C-cells via release of histamine and that cimetidine markedly interferes with this mechanism.     

Micrometeorological factors involved in development and survival of free-living stages of the sheep nematodesHaemonchus contortus andTrichostrongylus colubriformis. A review

Temperature and soil moisture are the most important factors affecting the development and survival ofHaemonchus contortus andTrichostrongylus columbriformis eggs and larvae on pasture. More than half of the eggs develop into infective larvae in the laboratory, but a very low percentage (0.03% forH. contortus) does so on pasture. There is a marked difference betweenH. contortus andT. colubriformis in survival of infective larvae.H. contortus larvae survived in the winter at Urbana poorly, whereasT. colubriformis did well. The former survived better than the latter in the spring and worse in the summer, while both survived equally well in the fall. Technics for larval recovery from pasture are not very efficient. Meteorologic conditions at ground level where the larvae exist are quite different from those in a standard weather shelter 1.6 m above the ground. Bioclimatographs in which mean monthly maximum temperatures are plotted against total monthly precipitation are fairly good in predicting the type of nematode liable to be important in a given region, but they are too simplistic to be relied on for more than approximations.     

Efficiency of a genetic test to detect benzimidazole resistant Haemonchus contortus nematodes in sheep farms in Quebec,Canada

Haemonchus contortus is a hemophilic nematode which infects sheep and causes anemia and death to lambs. Benzimidazole drugs are used to remove these parasites, but the phenomenon of resistance has arisen worldwide. A sensitive test to detect resistance before treatment would be a useful tool to enable farmers to anticipate the efficiency of the drug before drenching the flock. In this study, we compared a test for benzimidazole resistance based on detection of genetic markers in H. contortus before treatment with the common method of fecal egg count reduction test (FECRT). We recruited 11 farms from different regions of Quebec for this study. Fecal samples from animals were collected per rectum before and after treatment in control and treated groups (10 animals per group). The 10 sheep were treated with fenbendazole at the recommended dose rate. Among the 11 farms participating in the study, we found H. contortus in 8 of them and it was the most predominant nematode species detected by egg count. Using the genetic test, we found benzimidazole resistance in each of these 8 farms. In 5 of these 8 farms there were sufficient sheep with an egg count for H. contortus above 150 eggs per gram to allow the FECRT test to be conducted. Benzimidazole resistance was observed in each of these 5 farms by the FECRT. When we compared the results from the genetic test for samples off pasture and from individual sheep, with the results from the FECRT, we concluded that the genetic test can be applied to samples collected off pasture to estimate benzimidazole resistance levels before treatment for H. contortus infections.     

A combined microscopic-molecular method for the diagnosis of strongylid infections in sheep

We evaluated a combined microscopic-molecular approach for the diagnosis of key strongylid infections in sheep using panels of well-defined control and test samples. The method established is based on the separation of nematode eggs from faecal samples using a salt flotation procedure, the extraction and column-purification of genomic DNA, followed by real-time PCR and melting-curve analysis. Specific and semi-quantitative amplification from (a minimum of 0.1-2.0 pg) genomic DNA of Haemonchus contortus, Teladorsagia circumcincta, Trichostrongylus spp., Cooperia oncophora, Oesophagostomum columbianum, Oesophagostomum venulosum or Chabertia ovina is achieved using a specific, forward oligonucleotide primer located in the second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA (rDNA) together with a conserved reverse primer in the large subunit of rDNA. Using a panel of well-defined genomic DNA samples from eggs from sheep monospecifically infected with H. contortus or Te. circumcincta, there was a correlation between cycle threshold (Ct) values in the PCR and numbers of egg per gram of faeces, thus allowing the semi-quantitation of parasite DNA in faeces. The findings of the present study indicate that a microscopic-molecular approach provides a useful tool for diagnosis, for epidemiological and ecological surveys as well as for integration into parasite monitoring, drug resistance (i.e. ‘egg count reduction’) testing or control programmes, particularly following semi- or full-automation.     

Respiratory metabolism and thiabendazole susceptibility in developing eggs of Haemonchus contortus

Weston K. M., O'Brien R. W. and Prichard R. K. 1984. Respiratory metabolism and thiabendazole susceptibility in developing eggs of Haemonchus contortus. International Journal for Parasitology14: 159–164. The respiratory metabolism of and uptake of thiabendazole (TBZ) by unembryonated (8–16 cells) and embryonated eggs of Haemonchus contortus have been compared. Lipid, which forms the greatest energy reserve in the eggs, decreases during embryonation and seems to be the sole source of respiratory energy. Trehalose increases to the same extent as glycogen decreases during this development. Isocitrate dehydrogenase (NADP⁺) and lactate dehydrogenase were not detected in the unembryonated eggs, but were present after embryonation. In addition, the activities of citrate synthase and malate dehydrogenase significantly increased during embryonation. Respiratory enzymes involving cytochrome c oxidation and reduction were detected in eggs at both stages of development. However, in line with other results indicating an increased capacity for and utilization of aerobic metabolism, the rate of oxygen uptake more than tripled during development of the eggs. Although both unembryonated and embryonated eggs took up palmitate, its metabolism to CO₂ only occurred in the embryonated eggs.The unembryonated eggs, exposed to TBZ for 4 h, concentrated it 5.9 times and the embryonated eggs 2.1 times, which are in proportion to their respective lipid contents. Uptake of TBZ was dependent on the concentration in the incubation medium and appears to be a passive process.The studies indicate that the embryonated eggs have a greater capacity for, and do utilize aerobic metabolism to a greater extent than do unembryonated eggs. The reduced susceptibility of embryonated eggs to TBZ could be associated with this metabolic difference and/or with their reduced uptake of TBZ.     

Leucine aminopeptidase in exsheathing fluid of North American and Australian Haemonchus contortus

Rogers W. P. and Brooks F. 1978. Leucine aminopeptidase in exsheathing fluid of north American and Australian Haemonchus contortus. International Journal for Parasitology8: 55–58. Juveniles of Haemonchus contortus from north America and Australia produced exsheathing fluid containing leucine aminopeptidase when stimulated in tetraborate-carbon dioxide medium. Exsheathment in this medium was inhibited by 1, 10-phenanthroline, 10^?3M, and this inhibition was largely reversed by Zn²⁺, 10^?3M. This supports the view that the enzyme is produced by the juveniles and that it is concerned in exsheathment.     

Changes in blood leukocytes,bone marrow and lymphoid organs in sheep infected with Haemonchus contortus

Haemonchus contortus infection, which causes a blood loss anaemia in sheep, depleted leukocytes and produced leukopenia and a mild lymphopenia. Thymus atrophy, a decreased size of spleen and enlargement of adrenal glands occurred concomitantly in infected sheep which suggested that they were caused by the stress of infection. The overwhelming change in bone marrow of infected sheep was a 4-fold increase in erythroid series cells.Primary immunization with rat erythrocytes produced similar haemagglutinating antibody responses in infected sheep and in sheep allowed to recover from infection after treatment with thiabendazole. This suggests that extant infection with H. contortus was not associated with a secondary immunodeficiency. Blastogenic responses of blood lymphocytes from infected sheep to larval antigen correlated with faecal egg counts but not with haematocrit or leukocyte values. These results suggest that the unresponsiveness of lymphocytes to worm antigen which occur during infection with H. contortus is more closely related to contact with the parasite than to the pathological effects of infection.     

DNA sequences flanking the starts of the hsp 70 and alpha beta heat shock genes are homologous

To determine whether ribosomes have a role in the postfertilization activation of protein synthesis in sea urchin eggs, we measured the translational activity of ribosomes isolated from unfertilized eggs and embryos of Strongylocentrotus purpuratus. Numerous previous studies have indicated few if any differences in the activity of such ribosomes. However, by using improved physiological isolation and in vitro conditions, we have found important differences in the activities of egg and embryo ribosomes. Ribosomes obtained from blastula polyribosomes were active in translating reticulocyte mRNA in a ribosome-dependent cell-free translation system, whereas ribosomes obtained from unfertilized eggs became fully active only after a characteristic, reproducible delay of up to 15 min at 26°C. The extent of this delay varied with incubation pH, but not with concentrations of K⁺, Mg²⁺, initiation factors, or mRNA. However, at incubation pH between 6.90 and 7.65, the egg ribosomes were always less active than blastula ribosomes.     

Calcium influx following fertilization of Urechis caupo eggs.

Measurements of ⁴⁵Ca flux into and out of Urechis eggs indicate that, during the first 10 min after insemination, the eggs take up 0.24 pmole of Ca/egg. Total egg Ca measured by atomic absorption (AA) spectroscopy increased by 0.23 pmole of Ca/egg (0.56, 0.79, and 0.76 pmole of Ca/egg for unfertilized, 10-min fertilized, and 60-min fertilized eggs, respectively). Thus, the total change in egg Ca is accounted for by the influx even though the rate of efflux, measured as a release of ⁴⁵Ca from preloaded eggs, increases to twice the unfertilized rate by 15 min. The fertilization influx follows saturation kinetics (K_a = 1.3 mM). It is competitively inhibited by procaine, but is not inhibited by dinitrophenol, mersalyl acid, or ruthenium red. Ten percent of the total Ca influx has occurred by 10 sec, and it is, therefore, the most rapid response to fertilization yet known in these eggs. The influx is also observed in eggs partially activated by insemination in pH 7 seawater (SW); the other fertilization responses, except sperm penetration, do not occur in pH 7 SW. Although Ca influx alone is insufficient to activate the eggs, it may be a prerequisite for cytoplasmic activation and development, inducing other secondary responses which are prevented by low external pH.     

Proglumide: Selective antagonism of the rumination but not gastric motor effects induced by pentagastrin in sheep

The effect of intravenous (IV) vs intracerebroventricular (ICV) administrations of pentagastrin on gastro-intestinal motility and rumination were investigated by electromyography in sheep ; these effects were compared to those obtained after a previous IV or ICV injection of proglumide. When ICV administered at a dose of 20 ng.kg^?1, pentagastrin did not significantly affect the frequency of reticular and abomasal spiking activity but elicited a 13 to 37 min period of rumination after a delay of 23 ± 7 min. In contrast, when IV infused at a rate of 20 ng.kg^?1.h^?1 during 20 min, pentagastrin inhibited significantly the frequency of reticular and abomasal contractions for 30 to 40 min but did not induce rumination. Proglumide ICV administered (0.8 mg/kg^?1) abolished the rumination induced by central injection of pentagastrin whereas a 10 times higher dose administered systematically (8 mg.kg^?1 IV) did not block these effects. Both of ICV and IV administrations of proglumide at respectively 0.8 and 8 mg.kg^?1 were unable to antagonize the inhibitory effects of pentagastrin on reticulum and abomasum motility.     

The impact of refugia on the development of thiabendazole resistance in Haemonchus contortus

Martin P. J., Le Jambre L. F. and Claxton J. H. 1981. The impact of refugia on the development of thiabendazole resistance in Haemonchus contortus. International Journal for Parasitology11: 35–41. Beginning with a mixture of 5% thiabendazole-resistant and 95% non thiabendazole-resistant Haemonchus contortus larvae, worm free sheep were artificially infected with either 10,000, 9000, 7000, 2500 or zero larvae. Four weeks later, the infected sheep were treated with 44 mg/kg of thiabendazole. Seven days after anthelmintic treatment the same sheep were infected with zero, 1000, 3000, 7500 or 10,000 larvae respectively so that each sheep was given a total of 10,000 larvae. The larvae given after thiabendazole treatment are referred to as being in refugia (defined as an area where individual members of a population can escape exposure to a drug). This was repeated for six generations of parasites, maintaining the same proportion of larvae in refugia in each generation. The egg hatch assay for resistance indicated that refugia delayed the development of resistance in Haemonchus contortus. Where none or a small proportion of larvae were in refugia, a rapid increase in resistance occurred. With an increased proportion of larvae in refugia, resistance was slower to develop. Population size, as estimated by faecal egg counts done after anthelmintic treatment and corrected for the number of larvae given in each treatment, was lower at the higher levels of refugia. This was due to a lower level of resistance in the higher refugia lines and consequently a greater effect of the drug. However, the estimate of population size, from egg counts done after the larvae introduced from refugia had matured, were similar in all lines.     

Meiotic abnormalities in backcross lines of hybrid Haemonchus

Le Jambre L. F. and Royal W. M. 1979. Meiotic abnormalities in backcross lines of hybrid Haemonchus. International Journal for Parasitology10: 281–286. Hybrid worms from the reciprocal matings between Haemonchus contortus and H. placei were backcrossed to the parent species. Six of the possible eight backcrosses produced fertile eggs. The six backcross lines were passaged through sheep for four generations. The adults of this generation had various levels of meiotic abnormality as evidenced by abnormal variations in chromosome structure and aneuploidy. The eggs from backcross lines with a degree of meiotic abnormality had the lowest percentage hatch. Four of the six backcross lines required a higher temperature for larval development than did H. contortus. The size of the third-stage larvae varied from one with larvae larger than H. placei to one line with larvae smaller than H. contortus.