Melanisation of Teladorsagia circumcincta larvae exposed to sunlight: a role for GTP-cyclohydrolase in nematode survival.

Trichostrongylid nematode parasites of livestock inhabit two very different niches during their life-cycle; within the host and free-living in the environment. UV radiation plays a significant role in the survival of free-living, pre-parasitic nematode larvae, with different species exhibiting differing levels of sensitivity. In many eukaryotes, melanisation is a key protective mechanism against UV damage, however there is little information about this process in parasitic nematodes. Caenorhabditis elegans cat-4 mutants, which are deficient in the enzyme guanosine triphosphate-cyclohydrolase I (GTP-CH), have both depleted levels of melanin in their cuticles and an increased sensitivity to anthelmintic drugs. Some parasitic nematodes have very high levels of GTP-CH transcript in their pre-parasitic stages, suggesting an important role for this biopterin synthetic enzyme. Here, we show that the Tci-cat-4 gene, which encodes GTP-CH in Teladorsagia circumcincta, has a role in melanisation and is also capable of rescuing C. elegans cat-4 mutants. In addition, following exposure of T. circumcincta L3s to sunlight, there is a 32% increase in GTP-CH enzyme activity (P=0.019), and a 21% increase in levels of melanin (P=0.031) compared with unexposed larvae. These data suggest that one explanation for the high level of GTP-CH present in pre-parasitic stages of trichostrongylid nematodes is to facilitate melanisation in response to UV exposure.

Vaccination of young lambs against infection with Nematodirus battus using gamma irradiated larvae.

Helminthologically naIve 6-week-old Suffolk lambs were given 1-3 doses of 20000 gamma-irradiated infective larvae (L3) of the nematode Nematodirus battus at weekly intervals. Following an anthelmintic drench they were challenged with 50000 viable L3 at 10 weeks of age. Nematode worm burdens 14 days post-challenge showed a significant (P<0.01) 66% reduction in the single vaccine dose group. The two and three dose groups had mean worm burdens which were 30 and 42% lower than controls, respectively, although these were not statistically significant. There was little measurable stimulation of the immune system in the vaccinated lambs, suggesting that the repeatedly dosed animals may have developed immunological unresponsiveness to the parasite.

Attempts to immunise sheep against Ostertagia circumcincta with irradiated larvae.

Mature sheep immunised with two doses of 10,000 irradiated Ostertagia circumcincta larvae were only slightly resistant to challenge with a single dose of 10,000 larvae. This slight resistance was abolished if the 'vaccine' worm population was removed one week before challenge. In contrast, seven of eight sheep immunised with irradiated Haemonchus contortus larvae were highly resistant to homologous challenge. Evidence of some cross protection against challenge with O circumcinta was observed in five of six sheep immunised with irradiated H contortus in that egg counts were lower and more worms were retarded as fourth stage larvae.

The effect of gamma irradiation on Nematospiroides dubius. Factors affecting the survival of worms in a primary infection in mice.

The infective larvae of Nematospiroides dubius were exposed to various levels (0-30 krad) of gamma irradiation by means of a Cobalt 60 source. Groups of mice were infected with these larvae and autopsied 5 weeks later for worm counts. It was found that male worms were more susceptible to irradiation than female worms. In both instances, however, the survival curve on a semi logarithmic plot was characterised by a shoulder at low doses and an exponential component at the higher levels of exposure. No male worms were recovered from mice infected with larvae given more than 12 krad but some female worms were capable of surviving 20 krad. The fecundity of female worms was reduced by 61% at 4 krad and totally ablated at 8 krad. Further experiments demonstrated that the survival of irradiated N. dubius in vivo was related to the extent of the damage caused at the time of irradiation and was not dependent on additional host parameters. Thus neither the number of irradiated worms inoculated nor the sex of the host radically altered the sex ratio or proportion of the worms lost as a result of irradiating the larvae. Furthermore, treatment with cortisone or sublethal irradiation of the host did not increase the proportion of surviving worms. It was, therefore, concluded that a host immune response was not involved.

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.