|dc.description.abstract||The principal aim of this study was to improve the laboratory diagnosis of inflammation in farmed red deer, primarily through the investigation of blood cells and inflammatory proteins in a variety of inflammatory conditions.
The objectives were:
1) To establish solidly based reference values for haematological parameters and certain acute phase proteins, including fibrinogen, that may be useful in detecting and assessing infection in red deer.
2) To investigate various factors that might affect the results for these parameters, such information being essential for correct interpretation of test results.
3) To investigate aspects of the acute phase response in red deer.
4) To investigate changes in the above parameters in deer in a selection of infections.
5) To explore the observed responses in terms of possible functions of the components by reference to the literature.
Despite a lower content of myeloperoxidase in deer leucocytes, automated leucocyte differential equipment (Technicon H6000/C) was found to be suitable for use in investigating deer infections for the following reasons. Highly accurate results for the major inflammatory indicators were readily obtained. These included neutrophils, eosinophils, lymphocytes and platelets. In addition, red cell parameters and basophil counts were available from the same technology. Drawbacks included no indication of the number of juvenile neutrophils, counteracted to some degree by the enumeration of 'high peroxidase' cells, and the inclusion of monocytes with the lymphocytes.
Plasma viscosity was used as a screening test for plasma protein changes, fibrinogen and haptoglobin measurements being used to monitor specific acute phase proteins. Fibrinogen was selected as a commonly used inflammatory indicator in veterinary practice, while haptoglobin showed promise of being a particularly sensitive indicator in deer because of very low values in health yet rising to high values in infections.
Reference values were prepared according to documented techniques for a wide range of haematological parameters, supplementing existing information, and providing new information with respect to leucocytes, red cell parameters and inflammatory proteins. A particular problem of assessing inflammatory changes in stags, because of their aggressive behaviour from mid-summer to early winter, was noted.
The effect of some management procedures on these values was tested. The sedative drug xylazine (xylidino-5,6-dihydro-4-H-1,3-thiazine-hydrochloride) was shown to have marked effects, including a substantial reduction in circulating red cell mass, lymphocyte, basophil and platelet counts and small, though statistically significant, falls in fibrinogen and plasma viscosity. These changes, detected using automated equipment aside from the inflammatory proteins, were also detected using standard manual methods. By the use of splenectomised deer on which the effects of xylazine were tested before and after splenectomy, it was shown that the effects of xylazine sedation on red cell parameters and platelet count were considerably reduced in the absence of the spleen, whereas the effect on falls in basophil, lymphocyte and neutrophil counts were unchanged. Controls did not show the the changes seen in xylazine treated deer, except in the case of neutrophils, which also showed similar changes after injection of ACTH. The effect on basophils was very rapid, and maximal within 2 minutes of intramuscular injection of xylazine, whereas lymphocytes and platelets showed a slightly later nadir. In vitro studies indicated that basophils may be degranulated directly by xylazine, and it was suggested that these effects could be a factor in the occasional deaths of stags during velvet removal with the aid of xylazine.
It was shown that major surgery evoked an acute phase response, with sharp rises in plasma fibrinogen, serum haptoglobin and neutrophil count, eosinopaenia, basopaenia, and lymphopaenia. In most respects the pattern of the response was similar to that reported for other species, though the reduction in basophils, which persisted for several days, was unusual. As the return of both eosinophil and basophil values to pre-operation levels coincided with the return of acute phase protein levels, it suggests these cell types were involved in tissue repair.
In ACTH injection and sedation experiments, deer whose cortisol levels increased showed a neutrophil leucocytosis, lymphopaenia and eosinopaenia. Basophil count, haptoglobin, platelet count and red cell parameters showed no change attributable to handling. Studies on deer with lungworm and Strongylid infestation, as well as those with raised eosinophil counts, produced no evidence that parasite infestation was accompanied by an increase in acute phase proteins.
Haematological changes and acute phase proteins in infection were studied in a series of experiments in which groups of deer were artificially exposed to Yersinia pseudotuberculosis by the oral or subcutaneous route, and in naturally occurring yersiniosis, malignant catarrhal fever, occult abscess, and bovine tuberculosis.
In experimental infection with Yersinia, a neutrophil leucocytosis occurred which had subsided prior to the development of clinical signs. There was a thrombocytopaenia, probably due to endotoxin, and lymphopaenia, basopaenia and eosinopaenia in the early stages, with a selective loss of large platelets from the circulation. After several days, a thrombocytosis developed with an increase in large platelets, consistent with activation of megakaryocytes by IL-6. Comparison of animals with clinical signs that survived with those that did not suggests that a red cell mass indicator such as packed cell volume, and fibrinogen, neutrophil, lymphocyte and basophil counts are potential prognostic indicators of survival. Acute phase proteins were raised in animals before any clinical signs developed, and in deer which never showed clinical signs, but had Yersinia recovered from faeces.
Deer with severe yersiniosis showed lower weight gain than less severely affected animals, although the difference was not statistically significant.
Haptoglobin levels rose substantially, but with a dip around the 5th day after exposure when live organisms were used, indicating consumption exceeded production at the time organisms were most actively proliferating. Retrospective analysis of haptoglobin levels on a group of deer in which a wild-type sub-clinical Yersinia infection had occurred demonstrated that haptoglobin rose only in the animals that contracted yersiniosis.
Malignant catarrhal fever is generally a peracute fatal infection, fibrinogen and haptoglobin were above the upper limit of the reference range in all cases, while plasma viscosity was also abnormally high in all but one case. Lymphopaenia was present in all cases, and an increase in crenated red cells, confirmed by scanning electron microscopy, was a constant finding also. It is suggested that ischaemic necrosis due to haemorheological disturbance (high viscosity, capillary endothelial cell thickening, rapid changes in circulating red cell mass) and the lower oxygen carrying capacity of crenated red cells, could contribute to the rapidly fatal nature of malignant catarrhal fever.
In a 'chronic' case of malignant catarrhal fever, disseminated intravascular coagulation developed, reducing the platelets and fibrinogen by consumption. For some days, the only signs of abnormality in the laboratory tests were an increased haptoglobin level and lymphopaenia, though red cell mass indicators became low in the last 2 weeks of an illness lasting 1 month from the initial diagnosis, while fibrinogen and plasma viscosity were slightly low or low normal.
In bovine tuberculosis, the absolute neutrophil count, plasma viscosity and fibrinogen level were found to increase with increasing lesion severity. The basophil count also tended to be higher in lesion animals, but, unlike the previously mentioned tests, only rarely exceeded the upper limit of the reference range. An acute phase response was found to follow application of the standard bovine tuberculin skin test to deer in tuberculous herds, though not in healthy herds, or in animals reactive to avian PPD but not bovine PPD by the standard tuberculin skin test. This acute phase reaction was shown by increased plasma fibrinogen, plasma viscosity and serum haptoglobin, and infiltration of the skin test site by neutrophils at 72 hours after tuberculin injection. It was demonstrated a second time in those animals exhibiting the reaction, and was associated with circulating antibody to bovine PPD and skin test volume increase. The eosinophil count also fell in animals showing significant increases in acute phase proteins. Lesions could not be demonstrated in several deer from tuberculous herds that showed a post-skin test acute phase reaction, while deer in the same herd with the most severe lesions all showed this.
Approximately 80% of all hinds subsequently found to have bovine tuberculous lesions at post mortem had at least one of plasma viscosity, fibrinogen, neutrophil absolute number or neutrophil percentage outside the reference range for healthy deer, while 97% of the most severely affected hinds had at least one abnormal result. The most consistently raised parameter for all tuberculous deer was the plasma viscosity (raised in 57% of cases) while the neutrophil absolute number was the most commonly raised in the most severe cases (83%). Fibrinogen was only moderately successful as a lesion indicator (48%) unless tested 72 hours after PPD injection as for a standard tuberculin skin test, when the specificity of a raised fibrinogen level became 84%. There was some evidence that tuberculous hinds are less likely to become pregnant than non-tuberculous ones in the same mob.||