Haemonchus contortus microtubules are cold resistant.

Haemonchus contortus is an important nematode of livestock that is present in most parts of the world. The life cycle comprises free living stages (egg, L1, L2 and L3 larvae), and parasitic stages (L4, adult and egg) in a ruminant. Microtubules are filamentous structures which are made from polymerization of α- and ß-tubulin. In vitro polymerization of α- and ß-tubulin can be achieved by increasing the temperature to 37°C under certain conditions. As part of its normal functioning, in mammals, the microtubules can be depolymerized when the temperature is reduced to 0°C. However, interestingly the microtubules of H. contortus are cold resistant i.e. they do not depolymerize at 0°C. Moreover these microtubules did not depolymerize even in the presence of 5 mM CaCl2 or 50 µM colchicine. These interesting findings may explain how larvae in the free living stages may survive cold temperatures over winter.

Ultraviolet light increases mortality of nematode larvae and can explain patterns of larval availability at pasture.

Despite its documented effects on the viability of living organisms, the impact of ultraviolet (UV) light on the survival of parasitic nematode larvae has received surprisingly little attention. Infective L3s of the trichostrongyloid nematodes Haemonchus contortus, Teladorsagia circumcinta and Nematodirus battus, suspended in water, were exposed to direct UV irradiation in two experiments. In the first, during 6 days of constant illumination with UVA lamps at intensities simulating sunlight at ground level, the mortality rate was increased up to 100-fold compared with controls sheltered by UV-impermeable perspex. Significant differences in mortality rates were detected between the three species, with H. contortus the least sensitive. In the second experiment, larvae were exposed to natural sunlight during the temperate spring and summer, for 24-h periods on seven separate days representing a range of weather and UV doses. Mortality was again increased by UV exposure in all species, but was less in H. contortus than in T. circumcincta or N. battus. At higher daily UV doses, the mortality rate was on average 2.27 times higher in exposed larvae than in sheltered controls. Increased mortality caused by UV irradiation could help to explain patterns of abundance of infective stages at pasture, especially pronounced population declines in spring when solar radiation rises rapidly and temperature is still low. Implications for the epidemiology of gastrointestinal nematode infection in ruminants, and for trade-offs in parasite life history, are discussed.

Radiolabelled adult Haemonchus contortus obtained from eggs voided in the faeces of a sheep injected with 59Fe.

The infective larvae of Haemonchus contortus (L3) obtained from an artificially infected sheep injected with 125 microCi of 59Fe, incorporated the isotope and retained it in sufficient quantities to be detectable by autoradiography in the adult worms which developed. This is apparently the first report of incorporation of 59Fe into adult helminths, of which the L3 originated from an animal that was injected with the isotope.

A possible role of caesium radiations in the control of propagation of hookworm.

Faeces charcoal mixtures containing eggs of sheep hookworm, Haemonchus contortus were exposed to caesium radiations (23 r to 1656 r) from 1st to 7th day of incubation. Each plate was exposed only once. A dose related inhibition of transformation of eggs to infective larvae was observed. The maximum effect was observed on 2nd day of incubation. The radiation has maximum effect on the 1st stage larvae followed by 2nd stage. The 3rd stage or ineffective larvae are relatively resistant to the effect of radiation.

Influence of dietary protein on the immune responsiveness of sheep to Haemonchus contortus.

Twenty helminth-free lambs were fed diets containing either 169 g crude protein (CP) kg-1 dry matter (DM) or 88 g CP kg-1 DM from the age of seven months. One month later five lambs from each dietary group were vaccinated against Haemonchus contortus by the oral administration of 10,000 irradiated larvae on two occasions, four weeks apart. Four weeks following the administration of the second dose of irradiated larvae both the vaccinated and unvaccinated lambs were exposed to an experimental infection of 10,000 non-irradiated H contortus larvae Faecal egg output and haematological changes were monitored throughout the study. The lambs were slaughtered 28 days after challenge when worm burdens were assessed. Vaccination was equally successful in inducing a strong resistance to the challenge infection regardless of dietary status. It was concluded that dietary protein does not influence the response to vaccination with irradiated H contortus larvae of lambs more than seven months old.

Studies on the abomasal pathology of immunized and non-immunized sheep infected with Haemonchus contortus.

Five adult ewes and five lambs were repeatedly immunized with weekly doses of 10,000 irradiated Haemonchus contortus L3 before challenge with 100,000 and 10,000 normal larvae respectively. Two groups of non-immunized ewes and lambs were similarly challenged and one animal from each of the four groups killed on days 3, 5, 7, 10 and 24 post-challenge. The cellular changes in the abomasal mucosa were less marked in the non-immunized groups than in the immunized animals and appeared later in the lambs than in the ewes. Thus, in the immunized ewes increases in the numbers of mast cells and eosinophils were evident within five days of challenge whereas similar changes appeared later in the immunized lambs. Also marked lymphoid aggregates at the base of the mucosa and in the submucosa were detected only in the immunized ewes. However, both immunized and non-immunized ewes showed rises in the numbers of IgA plasma cells after challenge which were not evident in either group of lambs.

Some factors influencing the immunisation of sheep with irradiated Haemonchus contortus larvae.

Over the last 6 years a number of experiments in which sheep were successfully immunised with 2 doses of 10 000 Haemonchus contortus larvae, irradiated at 60 krad in a gamma-source, have been reported by the authors. In this paper, the failure of such a regimen of immunisation, using the same strain of larvae, is reported together with an investigation of the possible causes. Technical errors were eliminated as a cause, on the grounds that the failure occurred in 2 separate laboratories using different 60Co sources, as was increased radiosensitivity of the continuously passaged strain of H. contortus as measured by the percentage development of irradiated larvae in 6 naive lambs. From experiments utilising 18 lambs and using both irradiated and normal larvae, it was postulated that the strain had lost its original degree of immunogenicity. Subsequently, in 2 experiments involving 27 lambs, it was shown that irradiation at 40 krad was completely successful in restoring immunogenicity and producing a degree of protection against challenge similar to that previously reported with 60 krad. It was concluded that, with the current strain of H. contortus, 60 krad is now too high a level of irradiation of larvae to produce a consistently high degree of protection against challenge.

Use of coproantibody measurement to assess the influence of diet on local immune responses of sheep vaccinated against Haemonchus contortus.

Local antibody responses to challenge with 10,000 normal Haemonchus contortus larvae were evaluated in vaccinated and control Blackface sheep fed either a high or low protein diet. Anti-parasite antibody activity was found in faecal extracts of both vaccinated and control animals after infection. The coproantibody levels showed some correlation with abomasal mucosa antibody levels and faecal antibody activity was mostly associated with IgA. The low protein diet reduced local antibody responses in vaccinated and control sheep. However despite lower antibody levels the efficacy of vaccination was not diminished in sheep fed a low protein diet.

Radiolabeling of infective larvae of Haemonchus contortus (Nematoda: Trichostrongyloidea) with 75Se-methionine and their performance as tracers in sheep.

Haemonchus contortus infective larvae incorporated between 5 and 12 pCi/larva for each muCi of 75Se-methionine added per gram of fecal sediment. Thorough admixture of 75Se-methionine and fecal sediment was necessary to obtain approximately normal distribution and low variance of individual larval radioactivities. Ecdysis induced by treatment with 0.025% HClO in vitro resulted in loss of approximately 40% of the 75Se label of infective larvae. Loss of 75Se by parasitic larvae and adult H. contortus in vivo conformed to a two-component negative exponential function with half lives of 3.1 and 56 days acting on compartments representing 90% and 10%, respectively, of the 75Se label remaining after ecdysis. Labeled and unlabeled worms were readily distinguished by autoradiography 37 days after infection. No effect of gamma radiation arising from decay of 75Se in the range 130 to 1,300 pCi/larva could be measured in terms of survival or sex ratio of worms recovered at 17 days PI.

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.

Investigation of Haemonchus contortus infections in sheep. Comparison of irradiated larvae and transfer factor treatment.

A comparison is made between the resistance conferred to Haemonchus contortus challenge by irradiated larval treatment and transfer factor (TF) treatment in four-month-old lambs and seven-month-old lambs. As in previous investigations the irradiated larval treatment failed to confer resistance to the four-month-old lambs challenged with 10,000 third stage larvae while similar irradiated larval treatment in seven-month-old lambs reduced the worm burdens by 40 per cent compared to controls. The TF treatment produced a 34 per cent reduction in the challenge infection in the four-month-old lambs and a 45 per cent reduction in the seven-month-old lambs compared to the control lambs. It is concluded that TF activity operates independently of immune competence.