Preliminary observations on the experimental transmission of scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation

To determine the transmissibility of scrapie to Rocky Mountain elk (Cervus elaphus nelsoni), six elk calves were inoculated intracerebrally with brain suspension from sheep naturally affected with scrapie. One elk developed a brain abscess and was euthanatized at 7 weeks postinoculation (PI), and two others died at 6 and 15 months PI because of physical injuries. At 25 and 35 months PI, two other elk died after brief terminal neurologic episodes. Necropsy of these revealed moderate weight loss but no other gross lesions. Microscopically, characteristic lesions of spongiform encephalopathy were seen throughout the brains and the spinal cords, and in both cases these tissues were positive for PrP(res) by immunohistochemistry. Brains of both animals were positive for PrP(res) by western blot and for scrapie-associated fibrils (SAFs) by negative stain electron microscopy. PrP(res) and SAFs were not detected in the three elk that died or were euthanatized because of coincidental causes. Over 3.5 years after initiation of this experiment, the one remaining inoculated elk and two uninoculated (control) elk are alive and apparently healthy. These preliminary findings demonstrate that 1) sheep scrapie agent can be transmitted to elk by intracerebral inoculation; 2) the infection can result in severe, widely distributed spongiform change and accumulations of PrP(res) in the central nervous system (CNS); and 3) based on the examination of a limited number of CNS sections from two cases, this condition cannot be distinguished from chronic wasting disease with currently available diagnostic techniques.     

Transmission of sheep scrapie to elk (Cervus elaphus nelsoni) by intracerebral inoculation: final outcome of the experiment.

This is a final report of an experimental transmission of sheep scrapie agent by intracerebral inoculation to Rocky Mountain elk (Cervus elaphus nelsoni). It documents results obtained in experimental (n = 6) and control (n = 2) elk. During the first 2 years postinoculation (PI), 3 animals died or were euthanized because of infection or injuries other than spongiform encephalopathy (SE). In years 3 and 4 PI, 3 other inoculated elk died after brief terminal neurological episodes. Necropsy of these animals revealed moderate weight loss but no other gross lesions. Microscopically, characteristic lesions of SE were seen throughout the brain and spinal cord, and the tissue was positive for proteinase K-resistant prion protein (PrPres) by immunohistochemistry (IHC) and by Western blot. Scrapie-associated fibrils (SAF) were observed by negative-stain electron microscopy in the brain of elk with neurologic signs. PrPres and SAF were not detected in the 3 inoculated elk necropsied during the first 2 years or in the 2 control animals. Retrospective analysis of the gene-encoding cervid PrP revealed a polymorphism at codon 132. The elk with SE were either homozygous (MM) or heterozygous (LM). These findings confirm that intracerebral inoculation of sheep scrapie agent results in SE with accumulations of PrPres in the central nervous system of elk. Based on morphologic and IHC findings, the experimentally induced SE cannot be distinguished from chronic wasting disease of elk with currently available diagnostic techniques.     

Detection of PrP(CWD) in postmortem rectal lymphoid tissues in Rocky Mountain elk (Cervus elaphus nelsoni) infected with chronic wasting disease.

Preclinical diagnostic tests for transmissible spongiform encephalopathies have been described for mule deer (Odocoileus hemionus), using biopsy tissues of palatine tonsil, and for sheep, using lymphoid tissues from palatine tonsil, third eyelid, and rectal mucosa. The utility of examining the rectal mucosal lymphoid tissues to detect chronic wasting disease (CWD) was investigated in Rocky Mountain elk (Cervus elaphus nelsoni), a species for which there is not a live-animal diagnostic test. Postmortem rectal mucosal sections were examined from 308 elk from two privately owned herds that were depopulated. The results of the postmortem rectal mucosal sections were compared to immunohistochemical staining of the brainstem, retropharyngeal lymph nodes, and palatine tonsil. Seven elk were found positive using the brainstem (dorsal motor nucleus of the vagus nerve), retropharyngeal lymph nodes, and palatine tonsil. Six of these elk were also found positive using postmortem rectal mucosal sections. The remaining 301 elk in which CWD-associated abnormal isoform of the prion protein (PrP(CWD)) was not detected in the brainstem and cranial lymphoid tissues were also found to be free of PrP(CWD) when postmortem rectal mucosal sections were examined. The use of rectal mucosal lymphoid tissues may be suitable for a live-animal diagnostic test as part of an integrated management strategy to limit CWD in elk.     

Evaluation of vaginal implant transmitters in elk (Cervus elaphus nelsoni).

The effects of vaginal implant transmitters for tissue damage after 11 wk in 13 captive adult elk (Cervus elaphus nelsoni) and subsequent reproductive performance in 38 free-ranging elk were evaluated. Vaginal implant transmitters are designed to be shed at parturition and are used to locate birth sites of wild ungulates; however, potential adverse effects of these transmitters on tissues associated with the vaginal walls or subsequent reproductive performance have not been assessed. Vaginal implant transmitters consist of a transmitter encased in inert acrylic with an antenna trailing out the distal end and wings at the proximal end to hold the transmitter in place. Using a laparoscope on sedated captive elk, necrosis or measurable differences in tissue trauma between designs with wing spans of 80 versus 150 mm over an 11-wk trial were not observed. After the captive elk trial, vaginal implant transmitters with 80-mm wings were placed into 38 pregnant wild elk, and 31 live births were documented. Fates of seven calves were not determined, because their transmitters were not shed at the birth site. We recaptured 36 of these cow elk again in fall 2003 or spring 2004, and 32 were pregnant. This study was unable to document any short- or long-term effects of vaginal implant transmitters on reproductive performance of cow elk in captive and free-range environments.     

Variable patterns of distribution of PrP(CWD) in the obex and cranial lymphoid tissues of Rocky Mountain elk (Cervus elaphus nelsoni) with subclinical chronic wasting disease

Sections of medulla oblongata, taken at the level of the obex, palatine tonsil and medial retropharyngeal lymph node from 10,269 captive Rocky Mountain elk (Cervus elaphus nelsoni), were examined by immunohistochemical staining with monoclonal antibody for the prion protein associated with the transmissible spongiform encephalopathy of cervids, chronic wasting disease (PrP(CWD)). The protein was detected in 226 of them. On the basis of the anatomical location of the deposits in the brainstem of 183 elk, four distinct patterns of distribution of PrP(CWD) within the parasympathetic region of the dorsal motor nucleus of the vagus nerve and the adjacent nuclei were observed. Mild gross lesions of chronic wasting disease (serous atrophy of fat) were observed in only three elk, all with spongiform degeneration; the other elk were considered to be in the preclinical stage of the disease. In contrast with the relatively predictable distribution of prion protein (PrP) in the brain and cranial nodes of sheep and mule deer, the distribution of PrP(CWD) in the brain and nodes of the elk was more variable and unrelated to their PrP genotype. One hundred and fifty-five of the 226 positive elk had deposits of PrP(CWD) in the brainstem and lymphoid tissues, 43 had deposits only in the lymphoid tissue and 28 had deposits only in the brainstem.     

Experimental transmission of chronic wasting disease (CWD) of elk (Cervus elaphus nelsoni), white-tailed deer (Odocoileus virginianus), and mule deer (Odocoileus hemionus hemionus) to white-tailed deer by intracerebral route

To compare clinical and pathologic findings of chronic wasting disease (CWD) in a natural host, 3 groups (n = 5) of white-tailed deer (WTD) fawns were intracerebrally inoculated with a CWD prion of WTD, mule deer, or elk origin. Three other uninoculated fawns served as controls. Approximately 10 months postinoculation (MPI), 1 deer from each of the 3 inoculated groups was necropsied and their tissues were examined for lesions of spongiform encephalopathy (SE) and for the presence of abnormal prion protein (PrP(d)) by immunohistochemistry (IHC) and Western blot (WB). The remaining deer were allowed to live until they developed clinical signs of the disease which began approximately 18 MPI. By 26 MPI, all deer were euthanatized on humane grounds. Obvious differences in clinical signs or the incubation periods were not observed between the 3 groups of deer given CWD. In 1 of 3 nonclinical deer euthanatized at 10 MPI, minimal microscopic lesions of SE were seen in the central nervous system (CNS) tissues, and PrP(d) was observed by IHC in tissues of all 3 deer. In the clinical deer, CNS lesions of SE and PrP(d) accumulations were more severe and extensive. It is concluded that the 3 sources of CWD prion did not induce significant differences in time to clinical disease or qualitative differences in signs or lesions in WTD. However, this observation does not imply that these CWD agents would necessarily behave similarly in other recipient species.     

Experimental oral transmission of chronic wasting disease to red deer (Cervus elaphus elaphus): Early detection and late stage distribution of protease-resistant prion protein

Chronic wasting disease (CWD), an important emerging prion disease of cervids, is readily transmitted by intracerebral or oral inoculation from deer-to-deer and elk-to-elk, suggesting the latter is a natural route of exposure. Studies of host range susceptibility to oral infection, particularly of those species found in habitats where CWD currently exists are imperative. This report describes the experimental transmission of CWD to red deer following oral inoculation with infectious CWD material of elk origin. At 18 to 20 months post-inoculation, mild to moderate neurological signs and weight loss were observed and animals were euthanized and tested using 3 conventional immunological assays. The data indicate that red deer are susceptible to oral challenge and that tissues currently used for CWD diagnosis show strong abnormal prion (PrP^CWD) accumulation. Widespread peripheral PrP^CWD deposition involves lymphoreticular tissues, endocrine tissues, and cardiac muscle and suggests a potential source of prion infectivity, a means of horizontal transmission and carrier state.     

Pharmacokinetics of florfenicol in North American elk (Cervus elaphus)

Florfenicol pharmacokinetics after administration of a single subcutaneous (s.c.) dose of 40 mg/kg of body weight in adult elk (Cervus elaphus) was investigated. Serum florfenicol concentrations were determined by a sensitive high-performance liquid chromatographic method with limit of quantification of 0.03 microg/mL. Florfenicol pharmacokinetic parameters in elk were estimated using a noncompartmental approach. After a single s.c. injection, florfenicol concentrations remained above 1 microg/mL for approximately 36 h and above 0.5 microg/mL for approximately 72 h. Following s.c. injection, florfenicol was absorbed rapidly with a mean maximum concentration (C(max)) of 3.7 microg/mL achieved at 4.2 h (T(max)). The C(max) value in elk is similar to values reported in cattle at the same dose, suggesting that the 40 mg/kg s.c. dose achieves therapeutic concentrations in elk. A mean elimination half-life (t(1/2)) of 44 h is shorter than that reported in cattle. The more rapid elimination half-life in elk suggests that elk may require a multiple dose regimen for therapeutic success with s.c. Nuflor. We recommend s.c. Nuflor be administered subcutaneously to elk every 24 h at a dose level of 40 mg/kg.     

Field validation and assessment of an enzyme-linked immunosorbent assay for detecting chronic wasting disease in mule deer (Odocoileus hemionus), white-tailed deer (Odocoileus virginianus), and Rocky Mountain elk (Cervus elaphus nelsoni).

Tissue samples (n = 25,050 total) from 23,256 mule deer (Odocoileus hemionus), Rocky Mountain elk (Cervus elaphus nelsoni), and white-tailed deer (Odocoileus virginianus) collected statewide in Colorado were examined for chronic wasting disease (CWD) using an enzyme-linked immunosorbent assay developed by Bio-Rad Laboratories, Inc. (brELISA), in a 2-phase study. In the validation phase of this study, a total of 4,175 retropharyngeal lymph nodes (RLN) or obex (OB) tissue samples were examined independently by brELISA and immunohistochemistry (IHC). There were 137 IHC-positive samples and 4,038 IHC-negative samples. Optical density (OD) values from brELISA were classified as "not detected" or "suspect" based on recommended cutoff values during the validation phase. Using IHC-positive cases as known CWD-infected individuals and assuming IHC-negative cases as uninfected, the relative sensitivity of brELISA depending on species ranged from 98.3% to 100% for RLN samples and 92.1% to 93.3% for OB samples; the relative specificity of brELISA depending on species ranged from 99.9% to 100% for RLN samples and was 100% for OB samples. Overall agreement between brELISA and IHC was > or = 97.6% in RLN samples and > or = 95.7% in OB samples of all species where values could be calculated; moreover, mean brELISA OD values were > or = 46X higher in IHC-positive samples than in IHC-negative samples. Discrepancies were observed only in early-stage cases of CWD. Based on the validation phase data, only RLN samples were collected for the field application phase of this study and only samples with brELISA OD values > 0.1 were examined by IHC. Among 20,875 RLN samples screened with brELISA during this second testing phase, 155 of 8,877 mule deer, 33 of 11,731 elk, and 9 of 267 white-tailed deer samples (197 total) had OD values > 0.1 and were further evaluated by IHC to confirm evidence of CWD infection. Of cases flagged for IHC follow-up, 143 of 155 mule deer, 29 of 33 elk, and all 9 white-tailed deer were confirmed positive. Mean (+/- SE) OD values for IHC-positive cases detected during the field application phase were comparable with those measured in RLN tissues during the validation phase. Based on these data, brELISA was determined to be an excellent rapid test for screening large numbers of samples in surveys designed to detect CWD infections in deer and elk populations.     

Effects of supplemental feeding on gastrointestinal parasite infection in elk (Cervus elaphus): preliminary observations

The effects of management practices on the spread and impact of parasites and infectious diseases in wildlife and domestic animals are of increasing concern worldwide, particularly in cases where management of wild species can influence disease spill-over into domestic animals. In the Greater Yellowstone Ecosystem, USA, winter supplemental feeding of Rocky Mountain elk (Cervus elaphus) may enhance parasite and disease transmission by aggregating elk on feedgrounds. In this study, we tested the effect of supplemental feeding on gastrointestinal parasite infection in elk by comparing fecal egg/oocyst counts of fed and unfed elk. We collected fecal samples from fed and unfed elk at feedground and control sites from January to April 2006, and screened all samples for parasites. Six different parasite types were identified, and 48.7% of samples were infected with at least one parasite. Gastrointestinal (GI) nematodes (Nematoda: Strongylida), Trichuris spp., and coccidia were the most common parasites observed. For all three of these parasites, fecal egg/oocyst counts increased from January to April. Supplementally fed elk had significantly higher GI nematode egg counts than unfed elk in January and February, but significantly lower counts in April. These patterns suggest that supplemental feeding may both increase exposure and decrease susceptibility of elk to GI nematodes, resulting in differences in temporal patterns of egg shedding between fed and unfed elk.     

Mortality in Pennsylvania captive elk (Cervus elaphus): 1998-2006.

The necropsy records of 65 elk ranging from 1 day to 13(1/2) years of age were reviewed to determine the primary cause of death of each animal. The animal carcasses were submitted from 22 separate Pennsylvania elk farms over an approximately 8-year period. The most common causes of mortality in animals in which the cause of death was determined were gastrointestinal parasitism (21 cases), pneumonia (7 cases), enterocolitis (5 cases), malnutrition (5 cases), and rumenitis/acidosis (5 cases). The cause of death was undetermined in 4 of the 65 elk. Nematode organisms (20 cases) were the primary parasites identified in cases of mortality owing to gastrointestinal parasitism. Arcanobacterium pyogenes (3 cases), Escherichia coli (3 cases), and Streptococcus sp. (2 cases) were the most commonly isolated bacteria from the lung in the cases of pneumonia. Fungal organisms, consistent with Aspergillus sp. were present histologically within the pulmonary lesions in 5 cases. Bacterial agents associated with enterocolitis included Clostridium perfringens (2 cases), E. coli (1 case), Salmonella Newport (1 case) and Salmonella Thompson (1 case). The majority (56.3%) of the death loss in elk of known ages occurred in animals 1 year of age or less, with 71.4% of the gastrointestinal parasitism, 60.0% of the malnutrition, 60.0% of the enterocolitis, and 57.1% of the pneumonia cases occurring in animals within this age range. In conclusion, gastrointestinal parasitism, pneumonia, malnutrition, enterocolitis and rumenitis/acidosis were considered the most common causes of death in captive elk in this study.     

An estimation of reproductive performance of farmed elk (Cervus elaphus) in North America

North American farmed elk (Cervus elaphus) producers were surveyed to determine period specific rates of calving, abortion, and dystocia, and to examine the effect of reproduction-related mortality on the productivity of farmed elk. The overall calving rate was 86.3% and the calving rate for adult cows was found to be significantly higher than that for heifers. Abortions were seen on 7.6% of farms and the overall abortion rate was approximately 0.6%. Dystocia was experienced in 5.9% of births. Neonatal mortality, summer mortality, weaning mortality, and postweaning mortality rates were calculated and compared. Failure to produce a live neonate was responsible for 64% of the total reproductive loss and calf mortalities from birth to 1 year of age accounted for 36%. Failure to produce a live neonate had a more significant effect on productivity than did mortalities from birth to 1 year of age, suggesting that reproduction has a greater economic effect on elk farms than neonatal mortality.     

Evidence of Parelaphostrongylus tenuis infections in free-ranging elk (Cervus elaphus) in southern Ontario

The antemortem detection of a Parelaphostrongylus tenuis infection in a free-ranging wild elk (Cervus elaphus) in southern Ontario is documented. Postmortems on other free-ranging elk that died during 2000-2005 indicated that 59% (17/29) were infected with P. tenuis, based on presence of lesions in the brain.     

Preliminary observations on lungworms in farmed and feral red deer (Cervus elaphus) in Denmark

Deer farming is on its increase in Denmark. Based on experiences from other countries, lungworm infection is expected to cause severe problems in farmed deer. Therefore, in the present paper, which is the first one to deal with pathogens in Danish deer farms, the faecal larval excretion in red deer is examined.     

A retrospective study of the causes of morbidity and mortality in farmed elk (Cervus elaphus)

A survey of North American farmed elk (Cervus elaphus) producers was performed to determine the causes of sickness and mortality in farmed elk and to estimate mortality rates. Records over a 10-year period from 8 North American veterinary diagnostic pathology laboratories were also examined and summarized. The primary diagnosis for each record was used to classify diseases into categories such as parasitic, infectious, toxicological, and neoplastic. Nonspecific trauma was the most frequently reported known cause of mortality in both sexes and all age classes by elk producers. Ranked on perceived economic importance, producers cited trauma, chronic wasting disease, calf scours, dystocia, pneumonia, winter tick, tuberculosis, and grain overload. One-year mortality rates for adults and yearlings were 2.6% and 2.7%, respectively. Mortality rates for male and female adult animals were 2.4% and 2.7%, respectively. In general, the major findings of the survey matched reported causes for mortality provided by elk producers.     

A case of chronic wasting disease in an elk imported to Korea from Canada

A seven-year-old male elk (Cervus elaphus nelsoni) was euthanized and necropsied after having a 3-week history of body weight loss, emaciation, excessive salivation, teeth grinding, fever, anorexia, and respiratory distress. The elk was imported into Korea from Canada on March 9, 1997. Gross pathologic findings were restricted to a diffuse fibrinous pneumonia. Microscopic lesions included mild neuronal vacuolation and spongiform change in the neuropil of selected brain stem nuclei and generalized astrocytosis. Immunohistochemistry for protease-resistant prion protein (PrP(res)) was positive in all brain sections but more pronounced in the section of the obex of the medulla. And the PrP(res) was also detected by western immunoblotting in the brain and spinal cord. All the remaining elk and deer that had been in contact with this elk were destroyed and negative for chronic wasting disease (CWD). To our knowledge, this is the first case of CWD occurring outside of the U.S.A. and Canada.     

Preliminary findings on the experimental transmission of chronic wasting disease agent of mule deer to cattle.

To determine the transmissibility of chronic wasting disease (CWD) to cattle and to provide information about clinical course, lesions, and suitability of currently used diagnostic procedures for detection of CWD in cattle, 13 calves were inoculated intracerebrally with brain suspension from mule deer naturally affected with CWD. Between 24 and 27 months postinoculation, 3 animals became recumbent and were euthanized. Gross necropsies revealed emaciation in 2 animals and a large pulmonary abscess in the third. Brains were examined for protease-resistant prion protein (PrP(res)) by immunohistochemistry and Western blotting and for scrapie-associated fibrils (SAFs) by negative-stain electron microscopy. Microscopic lesions in the brain were subtle in 2 animals and absent in the third case. However, all 3 animals were positive for PrP(res) by immunohistochemistry and Western blot, and SAFs were detected in 2 of the animals. An uninoculated control animal euthanized during the same period did not have PrP(res) in its brain. These are preliminary observations from a currently in-progress experiment. Three years after the CWD challenge, the 10 remaining inoculated cattle are alive and apparently healthy. These preliminary findings demonstrate that diagnostic techniques currently used for bovine spongiform encephalopathy (BSE) surveillance would also detect CWD in cattle should it occur naturally.     

Gross and microscopic lesions of naturally occurring tuberculosis in a captive herd of wapiti (Cervus elaphus nelsoni) in Colorado.

A Mycobacterium bovis-infected herd of captive wapiti (Cervus elaphus nelsoni) in Colorado was depopulated after lesions of bovine tuberculosis were confirmed in 8 of 10 tuberculin skin test reactors. Of the 43 animals > 1 year of age, 26 had gross lesions suggestive of tuberculosis, 24 had microscopic lesions of tuberculosis, and 23 had acid-fast bacilli associated with the lesions. Lungs and retropharyngeal lymph nodes were the most frequently affected sites. Most lesions grossly and microscopically resembled tuberculosis in cattle; however, some lesions resembled abscesses or ovine caseous lymphadenitis lesions. Special stains and immunohistochemical techniques labeled few to numerous mycobacteria associated with the lesions.     

Experimental transmission of chronic wasting disease (CWD) from elk and white-tailed deer to fallow deer by intracerebral route: final report

Final observations on experimental transmission of chronic wasting disease (CWD) from elk (Cervus elaphus nelsoni) and white-tailed deer (Odocoileus virginianus) to fallow deer (Dama dama) are reported herein. During the 5-year study, 13 fawns were inoculated intracerebrally with CWD-infected brain material from white-tailed deer (n = 7; Group A) or elk (n = 6; Group B), and 3 other fawns were kept as uninoculated controls (Group C). As described previously, 3 CWD-inoculated deer were euthanized at 7.6 mo post-inoculation (MPI). None revealed presence of abnormal prion protein (PrP(d)) in their tissues. At 24 (Group A) and 26 (Group B) MPI, 2 deer were necropsied. Both animals had a small focal accumulation of PrP(d) in their midbrains. Between 29 and 37 MPI, 3 other deer (all from Group A) were euthanized. The 5 remaining deer became sick and were euthanized between 51 and 60 MPI (1 from Group A and 4 from Group B). Microscopic lesions of spongiform encephalopathy (SE) were observed in only these 5 animals; however, PrP(d) was detected in tissues of the central nervous system by immunohistochemistry, Western blot, and by commercial rapid test in all animals that survived beyond 24 MPI. This study demonstrates that intracerebrally inoculated fallow deer not only amplify CWD prions, but also develop lesions of spongiform encephalopathy.