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Agglutination of sheep erythrocyte-rabbit antibody complexes treated with sheep serum by a rabbit antiserum specific for the third component (C3) of sheep complement.

Sheep E-rabbit A incubated with sheep serum for up to 20 min at 39 degrees C and washed were agglutinated by a rabbit antiserum (anti-inulin-foetal lamb serum) that was specific for sheep C3. If incubated for longer than 20 min at 39 degrees C, there was less sensitisation. Such a complex was not readily generated at 17 degrees C whereas there was no reaction if incubated at 2 degrees C. Once the sheep E-rabbit A-sheep C complex had been generated at 39 degrees C and washed free of sheep serum, it could still be agglutinated after overnight storage at 4 degrees C. Addition of chelators of Ca2+ and/or Mg2+ or antrypol or cobra venom factor (a specific inhibitor of C3) prevented sensitisation of sheep E-rabbit A by sheep serum for agglutination. These results show that sheep serum C3 was taken up by sheep E-rabbit A and its presence on this indicator demonstrated by agglutination using a rabbit antiserum specific for sheep C3.

Animals↗

Within-flock transmission of sheep scab in naive sheep housed with single infested sheep.

The within flock transmission of sheep scab was studied in two trials, which lasted 14 and 12 weeks, respectively. A total of 40 non-infested, scab-naive sheep were divided into six groups of between 6 and 20 individuals. Each group was subsequently housed with a single infested index case, carrying scab lesions of different ages. The groups of sheep in both trials were video recorded continuously and all the sheep were individually examined on a weekly basis. The occurrence of transmission was measured in two ways: first, by the detection of early scab lesions during clinical examination and, second, by an increase in the rubbing of individual sheep as noted from the analysis of the video recordings. These two methods complemented each other in predicting the week on which transmission occurred. Overall, transmission was detected in 34 of the 40 scab-naive sheep. Transmission was shown to occur at about the time when the population of adult mites in lesions on index cases reached their peak numbers. The highest number of mites at this time, however, differed markedly between index sheep. There was no obvious relationship between the incidence of transmission and the time index sheep spent rubbing each day or other index sheep variables, including the age of the lesion or the time since the introduction to the scab free sheep.

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Differentiation of prion protein glycoforms from naturally occurring sheep scrapie, sheep-passaged scrapie strains (CH1641 and SSBP1), bovine spongiform encephalopathy (BSE) cases and Romney and Cheviot breed sheep experimentally inoculated with BSE using two monoclonal antibodies.

A panel of ruminant brain tissues were subjected to a Western immunoblotting technique using two monoclonal antibodies (mAbs). The resultant prion protein (PrP) glycoforms showed that three distinctions can be made between natural ovine scrapie cases and sheep experimentally inoculated with bovine spongiform encephalopathy (BSE). Differentiation between BSE-infected cattle and natural cases of sheep scrapie was also possible using these two antibodies. There were subtle differences in the molecular weight positions of the di-glycosylated, mono-glycosylated and unglycosylated forms of the abnormal PrP (PrP(Sc)) associated with these ruminant transmissible spongiform encephalopathies. In particular, a distinct difference for the unglycosylated protein band was observed. For ovine scrapie samples, this band was noticeably of a higher molecular weight than that found for brain samples from the Romney and Cheviot breed sheep infected with BSE and, to a lesser degree, higher than that observed for bovine BSE samples. Using the comparison of glycoform ratios, the technique provided a distinction between the sheep experimentally infected with BSE and natural cases of sheep scrapie but did not provide a distinction between natural cases of bovine BSE and ovine scrapie. The sheep-passaged CH1641 scrapie strain gave molecular weights similar to, but not identical to BSE, and a glycoform ratio similar to ovine scrapie cases. The SSBP1 experimental scrapie strain gave molecular weights that were akin to natural scrapie cases but the glycoform ratio was different to that found for all the other samples. When mAb P4 was substituted for mAb 6H4 in the technique, only the natural scrapie samples and SSBP1 gave strong signals. BSE in sheep and the CH1641 strain gave weak reactions and PrP(Sc) from BSE-infected cattle could not be detected at all. The results suggest that this combination of molecular weight and glycoform ratio analyses, and differentiation with two specific antibodies could be used to provide a possible screening test for BSE in the UK sheep flock, if confirmed as accurate by bioassay and lesion profile analysis in mice inoculated with brain tissue from suspect field cases.

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Suppression of sheep and goat lymphocyte proliferation by sheep, goat, and sheep x goat hybrid trophoblast tissue cultures.

Immunosuppressive activity of conditioned medium from cultured ovine, caprine, and hybrid trophoblast tissue was examined. Conceptuses were obtained from naturally mated donor ewes and does at d 20 of gestation and trophoblast tissue was cultured for 24 h in medium supplemented with 15% calf serum and 1% antibiotic/antimycotic. Conditioned medium was added to pokeweed mitogen-stimulated sheep and goat lymphocyte cultures. Quantification of [3H]thymidine uptake by cells was used to measure lymphocyte proliferation. Ovine, caprine, and hybrid conditioned medium effectively suppressed sheep and goat lymphocyte proliferation (P less than .01). There were no differences (P greater than .05) between the immunosuppressive activity of the three tissue types on either sheep or goat lymphocytes. For all treatment groups, sheep lymphocytes were suppressed more than goat lymphocytes (P less than .05). These results indicate that, at d 20 of gestation, sheep x goat hybrid trophoblast tissue is capable of suppressing pokeweed mitogen-stimulated lymphocyte proliferation.

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Lack of a specific immune response against a recombinant capsid protein of Jaagsiekte sheep retrovirus in sheep and goats naturally affected by enzootic nasal tumour or sheep pulmonary adenomatosis.

Enzootic nasal tumour (ENT) and sheep pulmonary adenomatosis (SPA) are two contagious adenocarcinomas of the respiratory tract of sheep and goats. Both diseases are associated with related, but distinct, type-D-retroviruses (ENTV and JSRV respectively). No evidence of circulating antibodies has been described in animals affected by either ENT or SPA using antigens from natural sources. We evaluated the usefulness of a recombinant JSRV capsid protein (JSRV-CA) as antigen to study the antibody responses of animals naturally affected by ENT or SPA, using immunoblotting. Positive reactions were detected in the sera of both affected and unaffected sheep and goats. The reactivity was abolished completely by absorption with the GST fusion partner but not by JSRV-CA, suggesting that it was not specific. The results support prior observations indicating that sheep and goats infected by JSRV and ENTV do not develop specific humoral responses to these retroviruses.

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[Major OLA histocompatibility complex of sheep. Frequency of factors in sheep affected by trembling (scrapie) or normal sheep].

The OLA histocompatibility complex of sheep was studied in a flock of Ile-de France breed affected with Scrapie. On the whole, gene frequencies of 9 out of 11 factors were increased in the non-affected sheep of this contaminated flock. In these sheep, frequencies of the three following genes appeared to be increased significantly: OLA-A4, OLA-A8, OLA-B6. When these genes were present in the sheep of this flock, the relative risk of Scrapie was estimated at 0.05, 0.10 and 0.25, respectively.

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Fatal Pasteurella haemolytica pneumonia in bighorn sheep after direct contact with clinically normal domestic sheep.

Six Rocky Mountain bighorn sheep were raised in captivity from birth (n = 5) or taken from the wild as a lamb (n = 1). After the bighorn sheep were in captivity for over a year, 6 clinically normal domestic sheep were placed on the 2 ha of pasture on which the bighorn sheep were kept. Nasal swab specimens were obtained from all sheep at the time the domestic sheep were introduced. Pasteurella haemolytica was isolated from swab specimens obtained from 4 of 6 domestic sheep, but not from specimens obtained from the bighorn sheep. All 6 bighorn sheep died of acute hemorrhagic pneumonia after exposure to domestic sheep. Death in the bighorn sheep occurred on days 4, 27, 27, 29, 36, or 71 after initial exposure to domestic sheep. Pasteurella haemolytica was isolated from respiratory tract tissue specimens of all bighorn sheep at the time of death. None of the domestic sheep were clinically ill during the study. At the end of the study, 3 of 6 domestic sheep were euthanatized, and at necropsy, P haemolytica was isolated from 2 of them. The most common serotypes in bighorn and domestic sheep were P haemolytica T-3 and A-2. Other serotypes isolated included P haemolytica A-1, A-9, and A-11 in bighorn sheep and A-1 in domestic sheep. On the basis of results of this study and of other reports, domestic sheep and bighorn sheep should not be managed in proximity to each other because of the potential fatal consequences in bighorn sheep.

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Susceptibility of phagocytes from elk, deer, bighorn sheep, and domestic sheep to Pasteurella haemolytica cytotoxins.

Alveolar macrophages and peripheral blood neutrophils from elk (Cervus elaphus), bighorn sheep (Ovis canadensis canadensis), and domestic sheep were exposed to culture supernatants from Pasteurella haemolytica isolated from bighorn sheep and domestic sheep. In a second experiment, peripheral blood neutrophils from mule deer (Odocoileus hemionus), elk, and bighorn sheep were exposed to culture supernatants from P. haemolytica isolated from elk, bighorn sheep and domestic sheep. Alveolar macrophages from elk, bighorn sheep and domestic sheep were resistant to killing by P. haemolytica supernatants from bighorn sheep and domestic sheep; susceptibility of neutrophils to cell death, as measured by release of lactate dehydrogenase, differed significantly (P < 0.05) between the four species tested. Bighorn sheep and domestic sheep neutrophils were susceptible to cytotoxin damage by the P. haemolytica isolates used; bighorn sheep neutrophils were four- to eight-fold more susceptible to cytotoxin damage than domestic sheep neutrophils. Neutrophils from deer and elk were resistant to killing by P. haemolytica cytotoxins from any species tested.

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Fatal pneumonia following inoculation of healthy bighorn sheep with Pasteurella haemolytica from healthy domestic sheep.

In a series of three experiments, isolates of Pasteurella haemolytica biotype A, serotype 2, ribotype reference WSU-1, from healthy domestic sheep, were inoculated intratracheally into eight bighorn sheep (Ovis canadensis canadensis) and seven domestic sheep with doses of bacteria ranging from 5.3 x 10(8) to 8.6 x 10(11) colony forming units. Seven of eight inoculated bighorn sheep died from acute pneumonia within 48 hr of inoculation, whereas all seven domestic sheep inoculated with comparable or greater doses of bacteria remained healthy. One contact control bighorn sheep also died 6 days after its penmates received P. haemolytica. Three other noncontact control bighorn sheep remained healthy during the experiments. Pasteurella haemolytica biotype A, serotype 2, ribotype reference WSU-1 in the inocula was recovered from one or more tissues from all bighorns that died; whereas, it was not detected in any bighorn sheep before inoculation. Three different ribotypes of P. haemolytica A2 were recovered from bighorn sheep; however, only the ribotype reference WSU-1 in the domestic sheep-origin inoculum was recovered from all dead bighorn sheep, and was not recovered from bighorn sheep that survived the experiments. Thus, a relatively nonpathogenic and common isolate of P. haemolytica from healthy domestic sheep was lethal in bighorn sheep under experimental conditions.

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Immune responses in Indonesian thin tail and Merino sheep during a primary infection with Fasciola gigantica: lack of a specific IgG2 antibody response is associated with increased resistance to infection in Indonesian sheep.

After a primary infection with Fasciola gigantica, the immune responses in a resistant (Indonesian thin tail) and a susceptible (Merino) breed of sheep were analysed. The number of adult flukes recovered from the livers of the Indonesian thin tail sheep were significantly lower than those found in the Merino animals. On days 8, 14 and 25 p.i., Indonesian thin tail sheep exhibited a significantly higher eosinophilia than Merino sheep, whereas neutrophilia was significantly elevated in the Indonesian thin tail sheep on days 36 and 48 p.i. Serum from both sheep breeds demonstrated IgM, IgG1 and IgE responses to F. gigantica. In contrast, the Indonesian thin tail sheep produced significantly lower levels of IgG2 antibodies relative to the high level detected in Merino sheep. The IgE response was biphasic in both sheep breeds with the first response detected by day 14 and the second response developing from days 30 to 60 p.i. Western blotting showed that a similar profile of adult fluke antigens was recognised by IgG1 and IgE antibodies in both the Indonesian thin tail and Merino sheep. The IgE response was directed to a major antigen at about 92 kDa. We postulate that IgG2 could act as a blocking antibody for protective effector responses against F. gigantica in sheep and that the Indonesian thin tail sheep, by downregulating IgG2 responses, have an enhanced capacity for killing F. gigantica in vivo.

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Mapping the sheep genome: production of characterized sheep x hamster cell hybrids.

Specific chromosomes of sheep have been selectively "captured" in hamster x sheep cell hybrids produced by fusing different Chinese hamster auxotrophs with lymphocytes from sheep carrying normal or Robertsonian translocation chromosomes. A minipanel has been established comprising sheep chromosomes 1; 3; t1 = rob(6;24); t2 = rob(9;10); and X in a predominantly monochromosomal state. Using this targeted cell fusion approach the nutritional markers, uridine monophosphate synthetase (UMPS), mapped onto human chromosome 3 (HSA3), phosphoribosylglycinamide synthetase, and phosphoribosylaminoimidazole synthetase (GART) (HSA21), are reported to be located on sheep chromosome 1q; and phosphoribosyl pyrophosphate amidotransferase (PPAT) (HSA4) has been assigned to sheep chromosome 6. By isozyme analysis and Southern hybridization, transferrin (TF) and superoxide dismutase 1 (SOD1), both in the bovine syntenic group U10, were assigned to sheep chromosome 1q; adenylate kinase 1 (AK1), in the bovine syntenic group U16, to sheep chromosome 3p; lactate dehydrogenase B (LDHB) and phenylalanine hydroxylase (PAH) on bovine chromosome 5 (BTA5) to sheep chromosome 3q; phosphoglucomutase 2 (PGM2) (bovine syntenic group 15) to sheep chromosome 6 (t1q); avian myelocytomatosis viral oncogene homolog (MYC) and Moloney murine sarcoma viral oncogene homolog (MOS) (BTA 14) to sheep 9 (t2q); and glutathione reductase (GSR) (bovine syntenic group U14) to sheep 24 (t1p).

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cDNA sequence of two sheep mast cell tryptases and the differential expression of tryptase and sheep mast cell proteinase-1 in lung, dermis and gastrointestinal tract.

BACKGROUND: Mast cell tryptases are a family of serine proteinases which are implicated in the proliferation of smooth muscle cells and fibroblasts, upregulation of interleukin-8 synthesis by endothelial cells, and recruitment of neutrophils and eosinophils. Trials in sheep showed that administration of a specific tryptase inhibitor reduced the late-phase response to inhaled allergen. OBJECTIVES: The aim of this study was to characterize the sequence and distribution of sheep tryptase(s), to validate the sheep model of allergic lung disease. METHODS: Reverse transcriptase PCR cloning was used to obtain cDNA sequences for two sheep tryptases. Lung and gut extracts were used as a source of tryptase for partial purification and characterization of the protein. The distribution of tryptase in skin, lung and gut was determined by immunohistochemistry, and compared with the distribution of sheep mast cell proteinase-1 (sMCP-1). RESULTS: Two highly similar cDNA sequences encoding sheep tryptase were found, indicating the presence of a 28 amino acid leader sequence, and a mature peptide of 245 amino acids. Partial purification of a putative sheep tryptase from lung and gut extracts was achieved using heparin-Sepharose affinity chromatography. Rabbit antihuman skin tryptase antiserum recognized the putative sheep tryptase on Western blot (approximate Mr 32-34 000) and paraformaldehyde-fixed tissue sections. Tryptase was detected in all lung, skin and gut mast cells by this antibody, and transcripts for tryptase were detected in all three tissues by RT PCR. Sheep mast cell proteinase-1, detected by a specific monoclonal antibody, was present in all intestinal and gastric mucosal mast cells, but was not found in mast cells of the muscularis, thus defining at least two mast cell phenotypes in the gut. Whereas all dermal and pulmonary mast cells were tryptase positive, only a low proportion in the lung, and almost none in the dermis, were positive for sMCP-1. CONCLUSION: In view of the structural and functional similarities of sheep and human tryptases, and their similarity in tissue distribution in normal sheep, the sheep lung appears to be a good model for in vivo studies relating to human tryptase.

Amino Acid Sequence↗

Treatment and prophylaxis of psoroptic mange (sheep scab) using an ivermectin intraruminal controlled-release bolus for sheep.

In an experiment to determine the therapeutic efficacy of an ivermectin intraruminal controlled-release (CR) bolus, 14 mixed breed sheep of one lot were infested with Psoroptes ovis and subsequently divided into two groups of seven. In one of these groups each sheep received one ivermectin CR bolus appropriate to its weight, the other group remained as an untreated control. All mites were eliminated from the group receiving the bolus while the control group remained infested, the disease progressed, and all but one sheep required treatment for psoroptic mange before the end of the experimental period. A second lot of 14 sheep, free from P. ovis, were divided equally into two groups to determine the prophylactic efficacy of the ivermectin CR bolus. In one group, each sheep was given an ivermectin CR bolus according to body weight and the sheep in the other group received no medication and served as untreated controls. Twenty-one days later two sheep infested with psoroptic mange were introduced into each of the groups. These donor sheep were removed 10 days later. The group treated with the ivermectin CR bolus remained mange-free and did not harbour any mites. All of the sheep in the control group developed psoroptic mange and required treatment to control the infestation at the end of the experimental period. Sheep that received the ivermectin CR bolus had greater mean weight gains than the control groups in these experiments. The ivermectin CR bolus releases a minimum dose of 20 microg ivermectin kg/day for 100 days: this prolonged activity should prove a valuable asset for the treatment and control of psoroptic mange in sheep.

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Development of pneumonia in desert bighorn sheep after exposure to a flock of exotic wild and domestic sheep.

From 1986 to 1989, 5 desert bighorn sheep (3 Ovis canadensis mexicana and 2 O c nelsoni), ranging in age from 2 to 3 years, were exposed to a flock of exotic wild and domestic sheep to potentially achieve naturally acquired pneumonia. Pasteurella multocida was isolated from nasal samples from 4 of 6 sheep randomly sampled from the flock. Bighorn sheep were exposed individually and each exposure period was a trial. Treatment before and after exposure varied and included combinations of alpha interferon, antibiotics, anti-inflammatory drugs, and vaccines. Treatments were chosen on the basis of recommendations of others for treating pneumonia in desert bighorn sheep as well as our own experience in sheep and cattle. Regardless of treatment used, bighorn sheep in trials 1 to 4 developed signs of pneumonia within 10 to 14 days of exposure. Bighorn sheep in trials 1 to 3 died within 11 to 17 days of initial exposure. In trial 4, the bighorn sheep was isolated from the carrier sheep for treatment of pneumonia on day 14 and died on day 30. Pasteurella multocida was isolated from lung tissue in 3 of the 4 bighorn sheep. On the basis of results of trials 1 to 4, a more in depth clinical study was conducted in trial 5. Nasal and blood specimens were collected prior to and during trial 5 for bacteriologic culturing and serologic testing for bovine viral diarrhea virus, infectious bovine rhinotracheitis, parainfluenza-3 virus, and respiratory syncytial virus.(ABSTRACT TRUNCATED AT 250 WORDS)

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Jaagsiekte sheep retrovirus can be detected in the peripheral blood during the pre-clinical period of sheep pulmonary adenomatosis.

Peripheral blood leukocytes (PBLs) and tissue samples from 36 sheep were examined for jaagsiekte sheep retrovirus (JSRV) by hemi-nested PCR. Animals were classified according to the status of sheep pulmonary adenomatosis (SPA), which was confirmed by pathological examination, as follows: (i) sheep with classical SPA (cSPA, n=10), (ii) sheep with atypical SPA (aSPA, n=6), (iii) non-affected sheep from SPA-affected flocks (in-contact, n=10) and (iv) non-affected sheep from SPA-free flocks (control, n=10). JSRV proviral DNA was detected in the PBLs of 10/10 cSPA, 5/6 aSPA, 4/10 in-contact and 0/10 control sheep. Lung tumours and lymphoid organs were also found to be JSRV-positive. The number of positive PCR results was greater for sheep in the cSPA group than for those in the aSPA and in-contact groups. For the first time, it is concluded that JSRV can be detected in naturally infected sheep before the onset of clinical disease and even before the development of discernible tumours.

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