Genetic resistance to experimental infection with Mycobacterium bovis in red deer (Cervus elaphus)

Tuberculosis (Tb) caused by Mycobacterium bovis is a worldwide threat to livestock and humans. One control strategy is to breed livestock that are more resistant to Mycobacterium bovis. In a 3-year heritability study 6 farmed red deer stags were selected from 39 on the basis of their differing responses to experimental challenge via the tonsillar sac with approximately 500 CFU of M. bovis. Two stags remained uninfected, two were moderately affected, and two developed serious spreading Tb. Seventy offspring, bred from these six stags by artificial insemination using stored semen, were similarly challenged with M. bovis. The offspring showed patterns of response to M. bovis challenge similar to those of their sires, providing evidence for a strong genetic basis to resistance to Tb, with an estimated heritability of 0.48 (standard error, 0.096; P < 0. 01). This is the first time the heritability of Tb resistance in domestic livestock has been measured. The breeding of selection lines of resistant and susceptible deer will provide an ideal model to study the mechanisms of Tb resistance in a ruminant and could provide an additional strategy for reducing the number and severity of outbreaks of Tb in farmed deer herds. Laboratory studies to identify genetic and immunological markers for resistance to Tb are under way. Preliminary studies showed no associations between NRAMP or DRB genes and resistance to Tb in deer. Patterns of immune responses seen in resistant animals suggest that both innate and acquired pathways of immunity are necessary to produce the resistant phenotype.     

Vocal anatomy, tongue protrusion behaviour and the acoustics of rutting roars in free-ranging Iberian red deer stags (Cervus elaphus hispanicus)

Roaring in rutting Iberian red deer stags Cervus elaphus hispanicus is unusual compared to other subspecies of red deer, which radiated from the Iberian refugium after the last glacial maximum. In all red deer stags, the larynx occupies a permanent low mid‐neck resting position and is momentarily retracted almost down to the rostral end of the sternum during the production of rutting calls. Simultaneous with the retraction of the larynx, male Iberian red deer pronouncedly protrude the tongue during most of their rutting roars. This poses a mechanical challenge for the vocal tract (vt) and for the hyoid apparatus, as tongue and larynx are strongly pulled in opposite directions. This study (i) examines the vocal anatomy and the acoustics of the rutting roars in free‐ranging male C. e. hispanicus; (ii) establishes a potential mechanism of simultaneous tongue protrusion and larynx retraction by applying a two‐dimensional model based on graphic reconstructions in single video frames of unrestrained animals; and (iii) advances a hypothesis of evaporative cooling by tongue protrusion in the males of a subspecies of red deer constrained to perform all of the exhausting rutting activities, including acoustic display, in a hot and arid season.     

Systemic mycosis in Scottish red deer (Cervus elaphus)

The pathological features of 10 cases of systemic infection of young red deer by fungi morphologically identified as phycomycetes are described. The most frequently affected organs were the kidneys, followed by liver and central nervous system. The fungi stimulated an acute pyogenic inflammatory reaction and a phagocytic giant cell response. Vasculitis with thrombosis and spread of fungi to the surrounding tissue was a common finding. Culture was attempted in 4 cases; one yielded Absidia, 2 Aspergillus and the other an unidentified fungal growth. Analysis of the histories suggests that, in nine of the ten cases, stressful husbandry may have been a predisposing factor.     

Malignant catarrhal fever-like disease in Barbary red deer (Cervus elaphus barbarus) naturally infected with a virus resembling alcelaphine herpesvirus 2

Eight Barbary red deer (Cervus elaphus barbarus) developed clinical signs suggestive of malignant catarrhal fever (MCF) over a 28-day period. These animals were housed outdoors with four other species of ruminants. Affected red deer had lethargy, ocular signs, and nasal discharge and were euthanatized within 48 h. Lesions included ulcers of the muzzle, lips, and oral cavity associated with infiltrates of neutrophils and lymphocytes. Serologically, six of seven red deer tested during the outbreak were positive by competitive enzyme-linked immunosorbent assay for antibodies to a shared MCF virus antigen. PCR using oligonucleotide primers designed for a conserved protein of alcelaphine herpesviruses 1 (AlHV-1) and 2 (AlHV-2) and for conserved regions of a herpesvirus DNA polymerase gene was positive for tissues from all eight clinically affected animals and negative for eight out of eight red deer without clinical signs of MCF. DNA sequencing of PCR amplicons from the diseased red deer indicated that they were infected with a novel herpesvirus closely related to AlHV-2; immunohistochemistry using polyclonal anti-AlHV-2 serum and in situ hybridization demonstrated the presence of virus within salivary glands adjacent to oral lesions of affected animals. A survey of other ruminants near the outbreak subsequently showed that normal Jackson's hartebeest (Alcelaphus buselaphus jacksoni) that were cohoused with the diseased red deer were infected with the same virus and were shedding the virus in nasal excretions. These findings suggest that a herpesvirus closely related to AlHV-2 caused the MCF-like disease epizootic in Barbary red deer and that the virus may have originated from Jackson's hartebeest.     

Seasonal dynamics of the fecal excretion of Elaphostrongylus cervi (Nematoda, Metastrongyloidea) first-stage larvae in Iberian red deer (Cervus elaphus hispanicus) from southern Spain

Elaphostrongylus cervi (Nematoda, Metastrongyloidea) is an extrapulmonary lungworm of red deer (Cervus elaphus) whose first-stage larvae (L1) require terrestrial gastropods as intermediate hosts. The seasonal pattern of fecal excretion of E. cervi L1 in Iberian red deer (Cervus elaphus hispanicus) was monitored during three annual periods (June 2000–May 2003) on a hunting estate from south-central Spain. The lowest rates of mean intensity of fecal L1 were found in summer, whereas no seasonal variation was found for prevalence. Monthly intensity of excretion was positively associated with early rainfall (the next month) rather than with rainfall of the same month. This seasonal rhythm of E. cervi L1 discharge may be the result of parasite adaptation to the seasonal Mediterranean climate and habitat constraints to improve the chance of parasite transmission. We have standardized the sampling period to compare E. cervi L1 infection rates of Iberian red deer populations in Mediterranean Spain, a natural limit of both the parasite and deer historical ranges, for potential use in the assessment of management strategies.     

Differential immune responses of red Deer (Cervus elaphus) following experimental challenge with Mycobacterium avium subsp. paratuberculosis

Immune responses of red deer (Cervus elaphus) that presented with different levels of paucibacillary pathology were profiled to detail immune changes during the progression of Johne's disease. Immune responses were monitored using an immunoglobulin G1 (IgG1) antibody enzyme-linked immunosorbent assay (ELISA), a gamma interferon (IFN-gamma) ELISA, and flow cytometry. Animals in the study were divided into outcome groups postmortem according to disease severity. All animals mounted IgG1 antibody and IFN-gamma responses to both the vaccination and experimental challenges. The Mycobacterium avium subsp. paratuberculosis-specific IgG1 antibody responses in the challenged group showed marked differences between infected and severely diseased animals. Slightly higher IFN-gamma responses were seen in infected animals compared with severely diseased animals. No significant changes were seen in the phenotype of lymphocyte populations investigated. Vaccination with killed M. avium subsp. paratuberculosis in mineral oil adjuvant reduced the level of severe disease; however, it obscured immunological differences between the infected and severely diseased groups. This suggests protection is not exclusively mediated via the presence of a type 1 response and, furthermore, the presence of a type 2 response is compatible with protection. These profiles provide information on the different immune processes in Johne's disease progression.     

Presumptive adenoviral bronchiolitis in a red deer (Cervus elaphus)

A case of fatal presumptive adenoviral bronchiolitis in a 1-week-old red deer (Cervus elaphus) fawn is described. Retrospective diagnosis of the adenoviral infection was made by electron microscopy on lung tissue that had been embedded in paraffin wax.     

Bone turnover associated with antler growth in red deer (Cervus elaphus)

Although it is known that skeletal bone depletion occurs during antler growth in deer, it is not clear whether repletion of the skeleton takes place before or after completion of antler development. This study attempted to correlate repeated scanning electron microscopic measures of ilium and rib bone porosity from six approximately 2‐monthly biopsy samples (using back‐scattered imaging) and biochemical markers of bone turnover (serum hydroxyproline and osteocalcin concentrations) taken for 11 months with antler growth in six red deer stags. No changes were detected in ilium samples but changes in porosity of rib bones and an elevation of the biochemical markers indicated that skeletal depletion occurred during the antler growth period. However, the decrease in rib bone porosity and decline in markers of bone turnover took place before completion of antler growth, indicating that a considerable amount of skeletal repletion could have occurred whilst antlers were also undergoing bone accretion. This latter finding extends the current view of antler growth being accompanied by a form of reversible osteoporosis in the skeleton by showing that there is a period when the antlers and skeleton are both undergoing net bone formation. Anat Rec 256:14–19, 1999. © 1999 Wiley‐Liss, Inc.     

Histological examination of antler regeneration in red deer (Cervus elaphus)

Annual antler renewal presents the only case of epimorphic regeneration (de novo formation of a lost appendage distal to the level of amputation) in mammals. Epimorphic regeneration is also referred to as a blastema-based process, as blastema formation at an initial stage is the prerequisite for this type of regeneration. Therefore, antler regeneration has been claimed to take place through initial blastema formation. However, this claim has never been confirmed experimentally. The present study set out to describe systematically the progression of antler regeneration in order to make a direct histological comparison with blastema formation. The results showed that wound healing over a pedicle stump was achieved by ingrowth of full-thickness pedicle skin and resulted in formation of a scar. The growth centers for the antler main beam and brow tine were formed independently at the posterior and anterior corners of the pedicle stump, respectively. The hyperplastic perichondrium surmounting each growth center was directly formed in situ by a single type of tissue: the thickening distal pedicle periosteum, which is the derivative of initial antlerogenic periosteum. Therefore, the cells residing in the pedicle periosteum can be called antler stem cells. Antler stem cells formed each growth center by initially forming bone through intramembranous ossification, then osseocartilage through transitional ossification, and finally cartilage through endochondral ossification. There was an overlap between the establishment of antler growth centers and the completion of wound healing over the pedicle stump. Overall, our results demonstrate that antler regeneration is achieved through general wound healing- and stem cell-based process, rather than through initial blastema formation. Pedicle periosteal cells directly give rise to antlers. Histogenesis of antler regeneration may recapitulate the process of initial antler generation.     

Anaplastic oligodendroglioma with meningeal infiltration in a free-ranging red deer (Cervus elaphus)

This report describes an anaplastic oligodendroglioma in a red deer, extending from the cerebellum and anterior medulla along the dorsal part of the brain stem to the posterior cerebral hemispheres and infiltrating the meninges. Immunohistologically, the tumour cells were labelled for S-100 but were negative for glial fibrillary acidic protein, vimentin and synaptophysin. This would appear to be the first report of an oligodendroglioma in a deer.     

Enterocytozoon bieneusi in sika deer (Cervus nippon) and red deer (Cervus elaphus): deer specificity and zoonotic potential of ITS genotypes

As the most common cause of the human microsporidiosis, Enterocytozoon bieneusi has been found in a wide variety of animal hosts. Deers are the ruminant mammals living in a variety of biomes, and the distribution of deer species differ by geography. To understand the prevalence of natural infection of E. bieneusi in deer and to assess their epidemiological role in the transmission of microsporidiosis caused by E. bieneusi, 91 fecal specimens were collected from 86 sika deers and five red deers in the northeast of China. By PCR and sequencing of the internal transcribed spacer (ITS) region of the ribosomal RNA (rRNA) gene of E. bieneusi, an average infection rate of 31.9 % (29/91) was observed in deer, with 32.6 % (28/86) for sika deer, and 20 % (1/5) for red deer. Six ITS genotypes were identified: one known genotype BEB6 (n?=?20) and five novel genotypes HLJD-I to HLJD-IV (one each) and HLJD-V (n?=?5). A phylogenetic analysis based on a neighbor-joining tree of the ITS gene sequences of E. bieneusi indicated that genotypes HLJD-II and HLJD-III fell into group 1 of zoonotic potential, while the other genotypes (BEB6, HLJD-I, HLJD-IV, HLJD-V) were clustered into so-called bovine-specific group 2. This is the first report of E. bieneusi in deer in China. The observation of genotype BEB6 in humans previously and in deer here and also the findings of the two novel genotypes (HLJD-II to HLJ-III) belonging to potential zoonotic group 1 suggested the possibility of deer in the transmission of E. bieneusi to humans.     

Apoptosis in lymph node granulomas from white-tailed deer (Odocoileus virginianus) experimentally infected with Mycobacterium bovis

Apoptosis is a morphologically and biochemically distinct mechanism of cell death seen in many physiological conditions as well as in various infectious diseases. To examine apoptosis in tuberculous white-tailed deer, 32 deer were each given an intra-tonsillar injection of 300 colony-forming units of Mycobacterium bovis. Medial retropharyngeal lymph nodes were collected at 15, 28, 42, 56, 89, 180, 262 and 328 days after inoculation. Microscopical sections of lymph nodes were labelled for apoptotic cells by the terminal deoxynucleotidyl transferase nick end labelling (TUNEL) method. TUNEL, and other morphological changes within developing granulomas, were analysed and quantified by computerized image analysis. TUNEL within granulomas was greatest 28 days after inoculation and had declined to negligible levels by 328 days. Granuloma enlargement was due primarily to an increase in size of the caseo-necrotic core of the granuloma and not to increased inflammatory cellular infiltrate. These findings suggested that cell death within M. bovis -induced granulomas in white-tailed deer was due mainly to mechanisms other than apoptosis.     

Bone turnover associated with antler growth in red deer (Cervus elaphus).

Although it is known that skeletal bone depletion occurs during antler growth in deer, it is not clear whether repletion of the skeleton takes place before or after completion of antler development. This study attempted to correlate repeated scanning electron microscopic measures of ilium and rib bone porosity from six approximately 2-monthly biopsy samples (using back-scattered imaging) and biochemical markers of bone turnover (serum hydroxyproline and osteocalcin concentrations) taken for 11 months with antler growth in six red deer stags. No changes were detected in ilium samples but changes in porosity of rib bones and an elevation of the biochemical markers indicated that skeletal depletion occurred during the antler growth period. However, the decrease in rib bone porosity and decline in markers of bone turnover took place before completion of antler growth, indicating that a considerable amount of skeletal repletion could have occurred whilst antlers were also undergoing bone accretion. This latter finding extends the current view of antler growth being accompanied by a form of reversible osteoporosis in the skeleton by showing that there is a period when the antlers and skeleton are both undergoing net bone formation.     

Haematological reference values for farmed red deer (Cervus elaphus) in New Zealand

An automatic haematology analyser (Technicon H6000/C) providing red cell, white cell and platelet parameters was calibrated for the blood of red deer Cervus elaphus. Haematology reference values for these parameters, plasma viscosity and plasma fibrinogen are presented for defined groups of female weaner red deer (n = 123), adult red deer hinds (n = 86) and weaner male red deer (n = 37), the latter being bled using physical restraint and also using xylazine sedation. Age, sex, sedation and season-related differences were found. Reference values are also presented for serum haptoglobin in adult hinds and levels were similar to those seen in cattle. Seasonal analysis of the hinds was checked on a further group of 336 hinds, showing higher acute phase proteins in spring and summer which were possibly related to an increase in eosinophil count starting in spring and continuing through into autumn, and slightly increased neutrophil counts in the summer. Circulating red cell mass parameters were higher in the winter in the hinds, whereas the reverse was found in male weaner deer. There was some evidence of mild iron deficiency in late pregnancy and during lactation in the females. MCV appeared to increase with age, and basophil count was higher in the male deer than in females. No evidence for a neutrophil leucocytosis during pregnancy was found. Plasma viscosity was essentially the same for adult hinds and male weaners, but slightly lower in female weaners, the male group showing a slightly higher upper limit for fibrinogen. Plasma viscosity exhibited seasonal variation.     

Iron storage disease in red deer (Cervus elaphus elaphus) is not associated with mutations in the HFE gene

Iron storage diseases are rare conditions of dysregulated iron metabolism in man and animals. A genetic basis has been confirmed only for human haemochromatosis. Iron storage disease was diagnosed in six related, 2-year-old male red deer of the same herd. These animals presented with weight loss and rough hair coats. Haematological examination was unremarkable. At necropsy examination, gross lesions were restricted to cachexia. Microscopical examination revealed severe, diffuse hepatocellular necrosis and iron accumulation in hepatocytes, Kupffer cells, cardiac myocytes and renal tubular cells in all affected animals. Four animals also had moderate bridging fibrosis in the liver. Hepatic iron concentrations were increased (1108-2275 mg/kg wet weight; reference range 100-200 mg/kg). Drinking water in rusty iron tubs in the deer park contained eight times more iron than the accepted level for human drinking water. To test for a possible genetic basis of increased iron uptake and storage in red deer, the cervid haemochromatosis gene (HFE) was identified. Sequence comparisons between the six diseased animals and three healthy free-ranging unrelated animals failed to identify differences in the HFE sequences. Furthermore, the disease was not associated with common amino acid substitutions reported in human patients with haemochromatosis, including C282Y and H63D. Polymorphisms in other non-HFE genes involved in iron metabolism may have led to a higher sensitivity to iron and this, together with the high iron content of the drinking water, may have been the cause of the observed iron storage in these red deer.     

Correlation of cytokine gene expression with pathology in white-tailed deer (Odocoileus virginianus) infected with Mycobacterium bovis

Mycobacterium bovis-infected white-tailed deer (WTD) in northeast Michigan are a reservoir of mycobacteria that pose a threat to both domestic animals and humans. Relatively little work has been done to characterize the immune response of WTD to M. bovis infection; however, an understanding of the immune response to infection and pathogenesis may be critical to the development of an effective vaccine. Immunological responses to infection were characterized by monitoring cytokine gene expression in M. bovis-infected and uninfected deer. Peripheral blood leukocytes (PBL) from infected WTD expressed more gamma interferon (IFN-gamma), interleukin-12p40 (IL-12p40), granulocyte-monocyte colony-stimulating factor, and IL-4 mRNA than did PBL from uninfected deer; however, differences were not detected in expression of IL-10 and transforming growth factor-beta mRNA. Infected animals could be divided into two groups based on pathology. Lesions were confined primarily to the lymph nodes of the head in animals with less severe pathology. Animals with more severe pathology had lesions in the lung and associated lymph nodes as well as the lymph nodes of the head. More robust IFN-gamma mRNA expression correlated with pathology early in infection. These findings indicate that IFN-gamma expression likely plays a role in both protection and pathogenesis.     

Influence of social status on ovarian function in farmed red deer (Cervus elaphus)

This study examines the effects of behavioural and environmental factors on ovarian function in red deer hinds. Patterns of postovulatory luteal progesterone secretion were investigated in groups of farmed red deer hinds following oestrus induced by progesterone administration and withdrawal. Hinds were held under conditions leading to low (Experiment 1, held in paddocks: 0.29 per animal/h) or high (Experiment 2, held in pens: 1.11 per animal/h) interaction rates, and progesterone was measured in jugular venous blood samples obtained daily for 14 days after ovulation. Plasma progesterone levels tended to differ with respect to dominance status in both experiments (p < 0.1). Progesterone levels were substantially lower following prolonged hot dry weather (mean 0.67 +/- SEM 0.03 ng/mL) than in a year of relatively typical climatic conditions (mean 1.63 +/- 0.04 ng/mL; p < 0.0005). Progesterone levels were not related to the body weight of the hinds, and were not affected by housing conditions leading to different interaction rates. We conclude that although there is a tendency for dominance status to affect ovarian function, as observed before, this relationship is obscured in farmed red deer where all animals in the herd enjoy a higher plane of nutrition and movements of individuals between groups confuse dominance relationships.     

Epidemiology and risk factors analysis of elaphostrongylosis in red deer (Cervus elaphus) from Spain

We studied the distribution and faecal shedding pattern of the first-stage larvae (L1) of Elaphostrongylus cervi (Nematoda: Protostrongylidae) in the red deer (Cervus elaphus) across Spain, where excretion was widespread. We evaluated the effects of individual, population and environmental factors on E. cervi L1 counts in 18 free-ranging red deer populations in South Central Spain. In this area, prevalence was 71.42±2.14% (n=448) and mean intensity (n=320) was 74.50±10.35. Aggregation of deer at water-holes was positively associated with E. cervi L1 prevalence, possibly due to spatial and temporal odds of infected gastropods, red deer and infective E. cervi L1 larvae being encountered. Prevalence increased with age, and there was also a trend towards males having higher intensities than females. A slightly decreasing age–intensity profile was identified for females, which may suggest a role of acquired immunity.