The immunogenetics of resistance toTrichostrongylus colubriformis and Haemonchus contortus parasites in sheep

Three possible immunogenetic markers for resistance to intestinal parasitesin sheep have been studied. Allotypes of the major histocompatibility complex (MHC) of the sheep have been investigated as markers, using serological typing or gene probes, for associations between allotypes and resistance to parasites in selected high responder and low responder lines of sheep. Only the serologically-determined class I ovine leucocyte antigen (OLA) types SY 1a and SY 1b have been found to be consistently associated with increased resistance to Trichostrongylus colubriformis, but this association has not extended to the immunological distinct Haemonchus contortus parasite. Gene probes of the sheep DRB, DQB and DQA MHC class II loci have detected animals with increased susceptibility to T. colubriformis. Eosinophilia was investigated as a marker and found to be associated with increased resistance to parasites in lines of Australian Merinos and New Zealand Romneys selected for resistance on the basis of low faecal egg count. Blood eosinophilia was distinct from eosinophil infiltration of the gut which was poorly associated with resistance. The mechanism of parasite resistance appeared to involve the release of vasoactive amines and leukotrienes into intestinal mucus, since the selected high responder sheep to T. colubriformis and H. contortus had significantly increased amounts of these agents in their gut mucus, compared with selected low responder or random-bred sheep. Antibodies to T. colubriformis and H. contortus have also been used as markers to select high responder sire groups of lambs in contact with the parasites, for the first time, at weaning. This assay had the advantage of detecting distinct antigens for the two parasites, which would allow resistance to the species of parasite to be selected in the lambs. Vaccines have been developed against H. contortus using 'novel' gut antigens from the parasite, but variable responsiveness of the host sheep seemed to result in varying degrees of protection which were stimulated by these vaccines

The use of embryo genotyping in the propagation of genes involved in the immune response

Multiple ovulation and embryo transfer (MOET) now enables researchers to produce identical twin animals, to obtain progeny from pre-pubertal females and to obtain more offspring from valuable animals. MOET and sexed semen have produced genetic progress of up to 60% of milk production. The oestrous cycles of animals are synchronized with progestagens before superovulation with gonadal hormones, pregnant mare serum gonadotrophin and follicle stimulating hormone. Surgical, non-surgical and laparoscopic methods are applied to recover and transfer embryos. Sexing and genotyping of the pre-implantation embryos is a key step in improving the management and breeding programmes for livestock, as well as in the human for the prenatal diagnosis of genetic disorders. Several serological and physiological methods have been used to determine the sex of the pre-implantation embryos; none has had satisfactory results in terms of time and accuracy. Sexing by polymerase chain reaction (PCR) using male-specific chromosome sequences alone or with female-specific chromosomal DNA probes simultaneously has been sufficient to identify the sex of the embryos with 100% accuracy. However, caution should be taken against sources of the contamination. The MHC class I, class II and background genes have been implicated in resistance to internal parasites in animals. Biotechnological methods such as screening of embryos prior to transfer using PCR and primer extension pre-amplification have already made it possible to, detect transgenic or genetically disordered embryos and could be applied to select those embryos bearing immunological genotypes of interest, such as resistance to internal parasites. Ultimately, cloning and nuclear transplantation would provide the possibility of isolating these resistance genes and to transfer them to livestock pre-implantation embryos to propagate these desirable traits

The PCR typing of MHC-DRB genes in the sheep using primers for an intronic microsatellite: Application to nematode parasite resistance

The strong association between polymorphisms in an intronic microsatellite and the coding sequences for (BoLA)-DRB3 genes, previously described for demonstrating alleles of class II major histocompatibility complex (MHC) in the cow, was examined in sheep to see if similar polymorphisms could be demonstrated in the DRB region of the MHC. The bovine primers LA53 and LA54, previously used to amplify the bovine DRB3 microsatellites, were used with DNA from Australian sheep, eight DRB alleles were identified by length polymorphisms of polymerase chain reaction (PCR) products amplified from the DRB microsatellite region. Incomplete amplification of both alleles was sometimes found for sheep DNA samples using bovine primers, so a modified primer (LA53b) was used, and found to amplify the microsatellite next to intron 2 of the MHC more reliably than the LA53 primer. Two additional primers (LA31 and LA32), used in amplification of the exon 2 region of bovine DRB3, were used in the sheep, and the PCR products were analysed by single-stranded conformation polymorphism (SSCP). These primers successfully amplified the variable region of the ovine DRB region coded by exon 2, and the SSCP technique demonstrated polymorphisms with sheep DNA. Family studies demonstrated the segregation of alleles, by amplification both of intronic microsatellites and of the exon 2 variable region. Close correspondence was found between the two regions for several alleles, suggesting that the intronic microsatellites were closely linked to DRB-variable region alleles. Three families of Merino sheep with different antibody responses to intestinal nematode parasites were examined. The sire group with the highest antibody levels possessed two microsatellite alleles of closely similar length (alleles 3 and 4) inherited from the sire and present in high frequency in the lambs. In contrast, the other two sires did not possess these two alleles and the alleles were in low frequency in their progeny. Further studies are required in unrelated sheep to confirm whether these two alleles are associated with resistance to nematode parasites

Variation in immune responsiveness of sheep to the antigens of intestinal nematodes and blowfly larvae

The total and IgG antibody responses to the intestinal nematode parasites Haemanchus contortus and Trichostrongylus colubriformis were measured in the serum of 160 lambs, 4 months of age. These autibodies had developed as the result of natural exposure to the parasites on pasture. Three sires were examined and strong sire effects on half-sib progeny were found. Plotting of ELISA antibody results in two dimensions revealed clustering of responses within sire groups. Bimodal antibody distributions were also observed within sire groups and the whole population for T. colubriformis. A bimodal distribution of antibodies to H. contortus was found for one sire group but not for the whole population. The injection of blowfly larvae (Lucilia cuprina) extract into 42 160 lambs at a later age (12 months) was followed by increased antibodies to L. cuprina and an apparent increase in antibodies to T. colubriformis. A bimodal distribution for antibodies to L. cuprina was found in one sire group and in the whole population. These bimodal distributions of antibodies to L. cuprina did not coincide with the distribution of antibodies to T. colubriformis or H. contortus, measured on the same serum samples. It was concluded that high and low responder sire groups could be differentiated in lamb populations for all three parasites. These effects persisted during lamb maturation and appeared to be genetic effects. Finally, crossreacting antibodies between L. cuprina and T. colubriformis appear to be stimulated by injection of L. cuprina antigens