R.G. Windon

Eosinophil responses in sheep selected for high and low responsiveness to Trichostrongylus colubriformis.

A breeding programme, based on selection for faecal egg counts, has produced lines of sheep which demonstrate either increased resistance (high responder) or susceptibility (low responder) to challenge infection with T. colubriformis after vaccination with irradiated larvae. Circulating blood eosinophilia, a hallmark of helminth infections, was examined in third generation lambs from two separate selective matings and random bred control lambs. Numbers of eosinophils were higher in high responder lambs when compared to low responders after vaccination and challenge infections. Analysis of eosinophil counts confirmed a strong line effect and there was no evidence of a sex effect. Random bred lambs showed wide individual variations in eosinophil numbers and their response to infection. It was concluded that peripheral eosinophilia was more a measure of host responsiveness to infection than an indicator of helminthiasis. As such the eosinophil may serve as an indicator of the hosts ability to respond to T. colubriformis vaccination and infection.

Identification of immune genes and proteins involved in the response of bovine mammary tissue to Staphylococcus aureus infection

BackgroundMastitis in dairy cattle results from infection of mammary tissue by a range of micro-organisms but principally coliform bacteria and Gram positive bacteria such as Staphylococcus aureus. The former species are often acquired by environmental contamination while S. aureus is particularly problematic due to its resistance to antibiotic treatments and ability to reside within mammary tissue in a chronic, subclinical state. The transcriptional responses within bovine mammary epithelial tissue subjected to intramammary challenge with S. aureus are poorly characterised, particularly at the earliest stages of infection. Moreover, the effect of infection on the presence of bioactive innate immune proteins in milk is also unclear. The nature of these responses may determine the susceptibility of the tissue and its ability to resolve the infection.ResultsTranscriptional profiling was employed to measure changes in gene expression occurring in bovine mammary tissues sampled from three dairy cows after brief and graded intramammary challenges with S. aureus. These limited challenges had no significant effect on the expression pattern of the gene encoding β-casein but caused coordinated up-regulation of a number of cytokines and chemokines involved in pro-inflammatory responses. In addition, the enhanced expression of two genes, S100 calcium-binding protein A12 (S100A12) and Pentraxin-3 (PTX3) corresponded with significantly increased levels of their proteins in milk from infected udders. Both genes were shown to be expressed by mammary epithelial cells grown in culture after stimulation with lipopolysaccharide. There was also a strong correlation between somatic cell count, a widely used measure of mastitis, and the level of S100A12 in milk from a herd of dairy cows. Recombinant S100A12 inhibited growth of Escherichia coli in vitro and recombinant PTX3 bound to E. coli as well as C1q, a subunit of the first component of the complement cascade.ConclusionThe transcriptional responses in infected bovine mammary tissue, even at low doses of bacteria and short periods of infection, probably reflect the combined contributions of gene expression changes resulting from the activation of mammary epithelial cells and infiltrating immune cells. The secretion of a number of proinflammatory cytokines and chemokines from mammary epithelial cells stimulated by the bacteria serves to trigger the recruitment and activation of neutrophils in mammary tissue. The presence of S100A12 and PTX3 in milk from infected udder quarters may increase the anti-bacterial properties of milk thereby helping to resolve the mammary tissue infection as well as potentially contributing to the maturation of the newborn calf epithelium and establishment of the newborn gut microbial population.

The role of immunologically specific and non-specific components of resistance in cross-protection to intestinal nematodes.

Abstract Vaccination of 6–8-month-old Merino sheep with irradiated Trichostrongylus colubriformis larvae gave a high level of protection (81 %) against single-species challenge with normal infective larvae of the same species. The level of protection (34%) was substantially reduced against challenge with a closely related species ( T. vitrinus ) and no significant protection occurred against single-species challenge with a generically unrelated nematode (Nematodirus spathiger). These results suggest that antigen(s) which stimulate protective immunity are shared by the related Trichostrongylus species but not by N. spathiger . By contrast with the results obtained for single-species challenge, vaccination with irradiated T. colubriformis produced 98–100% protection against all 3 species in animals challenged simultaneously with infective larvae of the 3 species. Comparison of the levels of protection recorded following the 2 types of challenge indicate that although a specific antigenic trigger is required to provoke an appropriate response, the results obtained, particularly in the case of N. spathiger , suggest that the terminal effector mechanism is not immunologically specific. The implications of these conclusions are discussed in relation to theories of the mechanism of resistance to gastrointestinal nematodes and the potential efficacy of vaccination in the field.

An association between a lymphocyte antigen in sheep and the response to vaccination against the parasite Trichostrongylus colubriformis

Abstract Lymphocyte antigens were tested in sheep which had been selected for responsiveness to vaccination against the intestinal nematode Trichostrongylus colubriformis . These sheep had been bred in an assortative mating programme which produced offspring designated as either “high responders” or “low responders”, with highly heritable resistance or susceptibility. Ovine lymphocyte antigen (OLA) typing antisera were obtained from parous ewes in the course of matings which produced the high and low responder flocks. A particular antigen (SY1) was found to be present in high frequency on the lymphocytes of high responder (72·2%) and in lower frequency (21·9%) on the lymphocytes of low responder rams. In ewes, the frequency for high responders was 65·7% and for low responders it was 33·5%. A similar association between the SY1 antigen and low faecal egg count was found in random-bred sheep which had been vaccinated with irradiated larvae and challenged with normal larvae. The conclusion was drawn that this lymphocyte antigen was likely to be part of the sheep major histocompatibility complex which influenced the immune response of sheep to vaccination against the parasite.

The effect of sire selection on the response of lambs to vaccination with irradiated Trichostrongylus colubriformis larvae.

Abstract Dineen, J. K. and Windon, R. G. (1980). The effect of sire selection on the response of lambs to vaccination with irradiated Trichostrongylus colubriformis larvae. International Journal for Parasitology 10 : 189–196. Rams selected for responsiveness and unresponsiveness to vaccination with irradiated T. colubriformis larvae at an early age were mated to unselected random bred ewes. Progeny were vaccinated with 20,000 irradiated larvae at 8 and 12 weeks of age, given anthelmintic treatment at 16 weeks and challenged with 20,000 normai larvae at 17 weeks. The results, based on wether worm counts and ewe faecal egg counts, showed significant differences between responder and non-responder progeny. There was a significant correlation between worm counts and faecal egg counts of half-sibs from the same sire group. The occurrence of globule leucocytes was inversely related to worm burdens of wether progeny, however, no clear relationship was found with eosinophils. In vitro lymphocyte stimulation using T. colubriformis L 3 antigen, concanavalin A and lipopolysaccharide showed that statistically defined responder progeny, pooled from both responder and non-responder sire groups, gave higher responses than non-responder lambs after vaccination. The results confirm that genetically-determined factors are involved in the response of lambs to vaccination at an early age, and indicate that rapid genetic progress may be achieved in the type of mating usually carried out under fleld conditions.

The segregation of lambs into 'responders' and 'non-responders': response to vaccination with irradiated Trichostrongylus colubriformis larvae before weaning.

Abstract Random bred Merino ram and ewe lambs were vaccinated at 1, 2 and/or 3 months of age with irradiated T. colubriformis larvae. An exponentially increasing challenge of normal larvae was given to all groups including unvaccinated controls commencing at 1 month of age. The results, based on faecal egg counts, showed a dissociation into animals which responded to vaccination (geometric mean egg count 441) and those which did not (geometric mean egg count 1567). The proportion of responders was greatest in groups first vaccinated at the earliest age (1 month). Wool growth and liveweight gains showed severe depression corresponding to peak egg counts, however, responders were less affected than non-responders. There was no correlation between haemoglobin type and resistance to challenge. Faecal egg counts after impulse challenge with 10,000 normal larvae given at about 6 1 2 months of age showed a significant ranked correlation with those obtained during the primary exponential challenge. These results confirm that a proportion of young lambs respond to vaccination with irradiated larvae, and that genetically-determined factors are implicated in the ability of animals to respond to vaccination at an early age.

The effect of selection of both sire and dam on the response of F1 generation lambs to vaccination with irradiated Trichostrongylus colubriformis larvae

Abstract Windon R. G. and Dineen J. K. (1981). The effect of selection of both sire and dam on the response of F 1 generation lambs to vaccination with irradiated Trichostrongylus colubriformis larvae. International Journal for Parasitology 11 : 11–18. Rams and ewes, tested for responsiveness to vaccination with irradiated T. colubriformis larvae at an early age, were mated on the basis of responder × responder and non-responder × non-responder. Progeny were vaccinated at 8 and 12 weeks of age with 20,000 irradiated larvae, treated with anthelmintic at 16 weeks and challenged with 20,000 normal larvae at 17 weeks. Faecal egg counts of progeny from responder matings were significantly lower than progeny from non-responders, and within each mating type, ewe lambs had markedly lower egg counts than ram lambs. The level of circulating complement-fixing antibodies to T. colubriformis larval extract were inversely related to egg counts. Thus, ewe progeny from responder matings had the highest serum antibody levels, non-responder ram progeny had the lowest levels and responder rams and non-responder ewes had similar intermediate levels. In vitro responses of cells stimulated with T. colubriformis L 3 antigen were greater in progeny from responder matings, whereas responses to bacterial lipopolysaccharide were higher in progeny from non-responder matings. The results confirm that the response to vaccination at an early age is genetically determined, and show that the response of progeny is most vigorously expressed when both sires and dams have been selected.

The response of lambs to vaccination and challenge with Trichostrongylus colubriformis: Effect of plane of nutrition on, and the inter-relationship between, immunological responsiveness and resistance

Abstract Wagland B. M. , Steel J. W. , Windon R. G. and Dineen J. K. 1984. The response of lambs to vaccination and challenge with Trichostrongylus colubriformis : effect of plane of nutrition on, and the inter-relationship between, immunological responsiveness and resistance. International Journal for Parasitology 14: 39–44. Merino lambs weaned at 8 weeks of age were fed either ground and pelleted (high plane, HP) or chopped (low plane, LP) lucerne hay ad libitum to achieve an approximate 2-fold difference in liveweight gain. When aged 17 and 21 weeks, 15 of the 20 lambs in each diet group were vaccinated with 80,000 irradiated T. colubriformis larvae. At 25 weeks, vaccinated and unvaccinated lambs were treated with anthelmintic and one week later challenged with 30,000 normal larvae. Four weeks after challenge the animals were killed for worm counts. After vaccination HP lambs had higher titres of antibodies to the parasite and after challenge had lower worm egg outputs, and lower worm burdens than LP lambs. Immunological responsiveness (serum titre of complement-fixing antibodies against worm antigen) and manifestations of resistance (eggs produced per female worm per day and percent protection calculated from worm counts) were significantly correlated within dietary groups. Percent protection and egg production per female worm were highly correlated ( r = −0.81) in individual animals pooled over dietary groups, suggesting that both manifestations of resistance respond to essentially the same immunological mechanism. Failure to obtain significant correlation between weight gain pre-vaccination and immunological and resistance parameters indicated that animal production and resistance to infection are not genetically linked. Negative correlation of weight gain during the vaccination period with serum antibody titre at challenge suggests that the developing immune response competes with weight gain for limited physiological resources of the animal.

Cross-immunity between Haemonchus contortus and Trichostrongylus colubriformis in sheep

Cross-protective immunity between the nematode parasites, Haemonchus contortus and Trichostrongylus colubriformis, was examined in sheep vaccinated with irradiated larvae of either species. Secondary immunological responsiveness stimulated in this manner protected only against challenge infection with the species used for vaccination. Significant cross-protective immunity was not observed. Titres of serum antibody to an extract of adult but not infective larval T. colubriformis reflected the specificity for protective immunity. Immediate hypersensitivity skin reactions to nematode extracts did not reflect the antigen-specificity for protective immunity.

Histamine and leukotriene concentrations in duodenal tissue and mucus of lambs selected for high and low responsiveness to vaccination and challenge with Trichostrongylus colubriformis.

Lambs selectively bred for high responsiveness or low responsiveness to vaccination with irradiated Trichostrongylus colubriformis were vaccinated and challenged. Duodenal tissue histamine concentrations in both high and low responder lambs were lower at 3 days than at 28 days after challenge. At 3 days after challenge, histamine concentrations were higher in both male and female high responder lambs than in low responder lambs whereas at 28 days concentrations were increased only in high responder females. At 3 days after challenge, histamine concentrations were generally lower in mucus than in tissues, but levels were again higher in mucus from high responder groups. In duodenal tissue at 3 days after challenge, leukotriene C4 and B4 concentrations were similar in high and low responder animals. At the same time, concentrations of both leukotrienes were higher in mucus than in tissues, with high responder female lambs having the highest concentration. It is suggested that increased levels of histamine and leukotrienes in mucus and tissue are associated with larval rejection or exclusion.