A bighorn sheep die-off in southern Colorado involving a Pasteurellaceae strain that may have originated from syntopic cattle

We investigated a pasteurellosis epizootic in free-ranging bighorn sheep (Ovis canadensis) wherein a Pasteurellaceae strain carried by syntopic cattle (Bos taurus) under severe winter conditions appeared to contribute to pneumonia in affected bighorns. Twenty-one moribund or dead bighorn sheep were found on the "Fossil Ridge" herd's winter range, Colorado, USA, between 13 December 2007 and 29 February 2008. Eight carcasses examined showed gross or microscopic evidence of acute to subacute fibrinous bronchopneumonia. All eight carcasses yielded at least one β-hemolytic Mannheimia haemolytica biogroup 1(±(G)) strain, and seven also yielded a β-hemolytic Bibersteinia trehalosi biogroup 4 (CDS) strain; evidence of Pasteurella multocida, Mycoplasma ovipneumoniae, and parainfluenza 3 and bovine respiratory syncytial viruses was also detected. Isolates of β-hemolytic Manneimia haemolytica biogroup 1(G) from a bighorn carcass and a syntopic cow showed 99.5% similarity in genetic fingerprints; B. trehalosi biogroup 4(CDS) isolates were ≥94.9% similar to an isolate from a nearby bighorn herd. Field and laboratory observations suggested that pneumonia in affected bighorns may have been caused by a combination of pathogens including two pathogenic Pasteurellaceae strains--one likely of cattle origin and one likely of bighorn origin--with infections in some cases perhaps exacerbated by other respiratory pathogens and severe weather conditions. Our and others' findings suggest that intimate interactions between wild sheep and cattle should be discouraged as part of a comprehensive approach to health management and conservation of North American wild sheep species.     

Chlamydial-caused infectious keratoconjunctivitis in bighorn sheep of Yellowstone National Park.

An epizootic of infectious keratoconjunctivitis occurred in bighorn sheep (Ovis canadensis) in Yellowstone National Park during the winter of 1981-82. The causative organism was identified as Chlamydia sp. Mortality related to the epizootic was approximately 60% of an estimated 500 bighorn sheep in the northern range population. The infection probably affected all sex and age classes, but field surveys of live animals and mortality suggested that mature rams died disproportionately. Limited field observations the following winter on individuals having both normal and cloudy-appearing eyes suggested that half of the bighorns then present on the core units of winter range had contracted the disease and survived. By 1988, there were about 300 bighorn sheep in the population.     

Population response of reintroduced bighorn sheep after observed commingling with domestic sheep

Bighorn sheep (Ovis canadensis) often die from respiratory disease after commingling with domestic sheep. From 2000 to 2009, we observed commingling between domestic and reintroduced bighorn sheep in 3 populations in UT, USA. We investigated how commingling affected survival of radio-collared female bighorns that were released initially (founder) and those that were subsequently released (augmented). We predicted that the proportion of young surviving to their first winter and population growth would be lower after observed commingling with domestic sheep. We observed groups of bighorns year-round on 2,712 occasions and commingling between domestic sheep and bighorns in 6 instances. On Mount Timpanogos, survival rates were best modeled as constant for females (n?=?57) before and after observed commingling with domestic sheep. Survival rates of female bighorns, however, decreased significantly in Rock Canyon (n?=?21) and on Mount Nebo (n?=?22) for founder, but not augmented bighorns after observed commingling with domestic sheep. Also, the proportion of young surviving to their first winter was almost 3 times lower and population growth was reduced for bighorns after observed commingling with domestic sheep in Rock Canyon and on Mount Nebo. Commingling between domestic and bighorn sheep reduced population parameters in 2 of 3 bighorn populations we studied; however, on Mount Timpanogos, interactions between those 2 species were not fatal for radio-collared female bighorns. Wildlife biologists should manage for spatial separation of these 2 species and consider the location of hobby farms and trailing operations of domestic sheep near release sites for bighorns.     

Spontaneous pasteurellosis in captive Rocky Mountain bighorn sheep (Ovis canadensis canadensis): clinical, laboratory, and epizootiological observations.

We observed clinical signs, compared adrenal responses, and performed diagnostic tests on 12 captive Rocky Mountain bighorn sheep (Ovis canadensis canadensis) during a spontaneous outbreak of pasteurellosis. Cortisol in urine and feces was measured for bighorns sampled three times between 20 October and 1 November 1986. By 6 November, four of these had developed pneumonia, four showed only mild rhinitis, and four remained clinically normal. Bighorns that ultimately developed pneumonia showed elevated mean urinary (P = 0.003) and fecal (P = 0.046) cortisol excretion over the 12-day sampling period. Twenty-four hour mean urine cortisol: creatinine ratios ranged from 10 to 57 ng/mg dry matter for affected and 5 to 22 ng/mg for healthy individuals; 24 hr mean fecal cortisol concentrations ranged from 7.2 to 20 ng/g dry matter for affected and 3.6 to 9.1 ng/g dry matter for healthy individuals. Elevated cortisol excretion preceded clinical pneumonia in affected bighorns by less than or equal to 16 days. Beta-hemolytic Pasteurella haemolytica biotype T, serotype 3 or 4, was isolated from nasal and pharyngeal swabs from all eight bighorns with pneumonia or mild rhinitis. We detected no evidence of parainfluenza 3, bovine respiratory syncytial virus, or Chlamydia psittaci using fluorescent antibody and/or serologic tests. Although elevated cortisol excretion was associated with pneumonia, we also believe age, reproductive physiology, and/or prior recovery from clinical pasteurellosis may have influenced individual susceptibility to pneumonia during this epizootic.     

Effects of delivery method on serological responses of bighorn sheep to a multivalent Pasteurella haemolytica supernatant vaccine

The safety and efficacy of a remotely delivered multivalent Pasteurella haemolytica supernatant vaccine (serotypes A2 and T10) were examined in captive Rocky, Mountain bighorn sheep (Ovis canadensis canadensis). Twenty bighorn sheep were grouped according to baseline leukotoxin neutralizing antibody titers (< or =2 or >2 log2(-1)) and vaccination history (previously vaccinated or unvaccinated). Within these groups, animals were randomly assigned to one of two delivery treatments: hand injection (control) or biobullet implantation. All bighorns received a single dose from the same lot of vaccine (n = 10/treatment); four additional animals were injected intramuscularly with 0.9% saline as unvaccinated sentinels. Mild, transient lameness one day after hand injection or biobullet implantation was the only adverse effect. Serum neutralizing antibody titers to P. haemolytica leukotoxin differed between delivery treatments (P = 0.009) and among baseline titer/vaccination history groups (P = 0.013). Neutralizing titers were higher among hand-injected bighorns. Although neutralizing titers were lower among implanted bighorns than hand-injected controls at 1 wk (P = 0.002) and 2 wk (P = 0.021) after vaccination, seroconversion rates in response to implantation (6/10) and hand injection (9/10) did not differ (P = 0.303). Agglutinating antibody titers to T10 were high and did not vary over time or between delivery treatments. Agglutinating antibody titers to A2 in the hand-injected controls were not different (P > or = 0.07) than those in bighorns vaccinated with biobullet implantation. These data demonstrate that although hand injection elicits higher absolute titers, biobullet implantation may also stimulate effective antibody responses to P. haemolytica supernatant vaccine. Further evaluation of biobullet vaccination against pneumonic pasteurellosis in free-ranging populations of wild bighorn sheep is warranted.     

Characterization of Pasteurella multocida associated with pneumonia in bighorn sheep

Pasteurella multocida is a highly diverse group of bacteria recognized as important pathogens. Although P. multocida is not ordinarily associated with disease in Rocky Mountain bighorn sheep (Ovis canadensis canadensis), numerous isolates were cultured in high numbers from free-ranging bighorn sheep in the Hells Canyon area of Idaho, Washington, and Oregon (USA) during the winter of 1995-96. Animals captured in Hells Canyon and held in captivity, and their offspring, also harbored P. multocida. Biochemical utilization tests on 90 isolates identified three subspecies: P. multocida multocida a (n = 54); P. multocida multocida b (n = 13); and P. multocida gallicida (n = 15); and a non-speciated biotype, U6 (n = 8). Genomic DNA digestion with restriction endonuclease Hha I separated the isolates into 62 unique restriction fragment length polymorphism profiles. Capsular type A was predominant (72% of isolates). Only one isolate type, which may have been transmitted from a feral goat, was capsular type D, possessed the structural gene, toxA, for dermonecrotoxin detected by polymerase chain reaction, and produced toxin as determined by monoclonal antibody immunoblot. In conclusion, bighorn sheep in this study carried diverse types of generally non-toxigenic P. multocida associated with epizootic pneumonia.     

Toxoplasmic encephalitis in a free-ranging Rocky Mountain bighorn sheep from Washington

A 4-mo-old free-ranging Rocky Mountain bighorn sheep (Ovis canadensis canadensis) from the Hells Canyon area (Washington, USA) was diagnosed with encephalitis associated with Toxoplasma gondii infection. The sheep had concurrent pneumonic pasteurellosis and resided in a geographic area with endemic Pasteurella-associated pneumonia and mortality in bighorn sheep. The brain had multifocal necrotizing and nonsuppurative encephalitis with intralesional protozoa. The protozoa were identified as T. gondii by immunohistochemistry. To our knowledge, this is the first report of T. gondii infection in a Rocky Mountain bighorn sheep.     

Experimental contact transmission of Pasteurella haemolytica from clinically normal domestic sheep causing pneumonia in Rocky Mountain bighorn sheep

Two Rocky Mountain bighorn lambs (Ovis canadensis canadensis) were held in captivity for 120 days before being housed with two domestic sheep. The lambs were clinically normal and had no Pasteurella spp. on nasal swab cultures. The domestic sheep were known to carry Pasteurella haemolytica biotype A in the nasal passages. After being in close contact for 19 days. P. haemolytica biotype A was cultured from nasal swabs of one of the bighorn lambs. By 26 days, both bighorn sheep developed coughs, were anorectic and became lethargic and nasal swabs yielded P. haemolytica biotype T, serotype 10. Twenty-nine days after contact, the lambs were necropsied and found to have extensive fibrinous bronchopneumonia. From affected tissues pure cultures of beta-hemolytic P. haemolytica biotype T, serotype 10 were grown. Both domestic sheep remained clinically normal and had no gross or microscopic lesions, but they carried the same P. haemolytica serotype in their tonsils. Behavioural observations gave no indication of stress in the bighorn lambs.     

Bighorn Sheep Abundance Following a Suspected Pneumonia Epidemic in Rocky Mountain National Park

Abstract: Anecdotal evidence of a pneumonia epizootic among bighorn sheep (Ovis canadensis canadensis) in Rocky Mountain National Park (RMNP), Colorado, USA, during the mid-1990s prompted park officials to examine the current condition of the herds. Here we present a mark—resight study design to estimate population abundance that, in many circumstances, is a reliable and cost-effective alternative to traditional mark—recapture or to indices of population abundance. We captured 59 adult females and radiocollared them via helicopter net-gunning during winter 2002–2003. From ground resighting surveys conducted May—September, we estimated the total RMNP bighorn population at 389.9 (SE = 34.9, CI = 327.2–464.6) in 2003 and 366.4 (SE = 34.7, CI = 304.4–441.0) in 2004. Previous abundance estimates suggest a park-wide decline has occurred between the late 1980s and the suspected pneumonia epidemic of the mid-1990s. Although the 2 years of data from our study are not enough to predict whether the herds are capable of recovering to previous levels, they provide park officials the tools necessary to make the most informed decisions for future monitoring and management of this fragile species.     

Immunologic responses of domestic and bighorn sheep to a multivalent Pasteurella haemolytica vaccine

The efficacy of a Pasteurella haemolytica vaccine (serotypes A1, A2, and T10) to induce humoral antibodies and alter colonization of the upper respiratory tract by related P. haemolytica spp. strains was evaluated in 10 bighorn (Ovis canadensis canadensis) and 10 domestic (Ovis aries) sheep. Sheep of each species were divided into five pairs based on age and history of respiratory disease. One sheep in each pair was vaccinated twice 2 wk apart with 2 ml of vaccine (VAC group) and the remaining animals (NV group) were injected with 2 ml of sterile saline. Mild, transient lameness was the only observed adverse effect. Blood sera from the sheep were tested for agglutinating antibodies against whole cells of A1, A2, and T10 and for leukotoxin neutralizing antibodies. Antibody titers were expressed as the reciprocal log2 of the highest reactive dilutions. Domestic sheep > 1-yr-old and two bighorn sheep with a history of A1 infection had higher titers throughout the study against A1 cells than domestic sheep < 1-yr-old and bighorns without a history of A1 infection. Both domestic and bighorn sheep had log2 titers of 8 to 12 against A2 cells and 6 to 12 against T10 cells during this time. Bighorn sheep in the VAC group had 2 to 32 fold titer increases for A1 cells by 2 wk post-vaccination (PV) compared to 0 to 2 fold increases in VAC domestic sheep. Two to 16 and 0 to 8 fold increases in antibodies titers to A2 and T10 cells, respectively, were detected in sera of both VAC groups. Sera of bighorn sheep with a history of respiratory disease and all domestic sheep had log2 leukotoxin neutralizing antibody titers of 4 to 14 in contrast to < or = 2 in sera of bighorn sheep without a history of respiratory disease. Neutralizing antibody titers of two bighorns without a history of respiratory disease in the VAC group increased from log2 0 to 5 in one and from 0 to 9 in the other 2 wk PV. Antibody increases in these animals were no longer evident at 16 wk PV while titers of animals with histories of disease remained relatively stable. The types and numbers of Pasteurella spp. isolated from nasal and pharyngeal swabs varied throughout the study without conclusive evidence of suppression of colonization. Although the animals were not experimentally challenged to determine the efficacy of the vaccine, one VAC and one NV bighorn sheep died following introduction of an A2 P. haemolytica strain when leukotoxin neutralizing antibodies had returned to pre-vaccination levels. This vaccine appeared to be safe for use in bighorn sheep and stimulated moderate but transient increases in antibody levels which should provide some protection against naturally occurring disease. A vaccine which would induce production of high and maintained antibodies against multiple strains of P. haemolytica would be valuable for use in bighorn sheep maintained in captivity or when captured for relocation.     

Comparison of pulmonary defense mechanisms in Rocky Mountain bighorn (Ovis canadensis canadensis) and domestic sheep

Alveolar macrophages were obtained from Rocky Mountain bighorn sheep (Ovis canadensis canadensis) and domestic sheep for the purpose of comparing pulmonary host defense mechanisms in the two species. Specific variables studied included (1) characterization of the cell types present in the lung, (2) alveolar macrophage phagocytic and bactericidal functions, (3) measurement of protein levels in lavage fluid, and (4) measurement of cortisol levels in lavage fluid. While phagocytic cell populations were similar between bighorn and domestic sheep, a significantly higher percentage of lymphocytes were present in bighorns than domestics (20% in bighorn versus 6% in domestic sheep). Significant differences were not observed in the phagocytic or bactericidal functions of macrophages between the two species. Significant differences were not observed in either lavage fluid protein levels or in cortisol levels.     

Hemorrhagic disease in bighorn sheep in Arizona

Two bighorn sheep from Arizona (USA) were submitted for necropsy. One was a Rocky Mountain bighorn (Ovis canadensis canadensis) and the other was a desert bighorn (Ovis canadensis mexicana). Both had lesions consistent with those of hemorrhagic disease (HD). Epizootic hemorrhagic disease virus (EHDV) type-2 and bluetongue virus (BTV) type-17, respectively, were isolated from the sheep tissues. To our knowledge, HD caused by either EHDV or BTV infection has not been documented previously in Arizona bighorn sheep.     

Isolation of Pasteurella haemolytica from tonsillar biopsies of Rocky Mountain bighorn sheep

Isolations of Pasteurella haemolytica were compared from tonsillar biopsies versus nasal passages for 29 free-ranging Rocky Mountain bighorn sheep (Ovis canadensis canadensis) from central Idaho. Overall, P. haemolytica was isolated from 11 (38%) of 29 sheep. Two (18%) of the 11 positive samples were from only nasal passages compared to eight (73%) from tonsillar biopsies. Pasteurella haemolytica biotype T was isolated from tonsils of nine sheep and from nasal biopsies. Pasteurella haemolytica biotype T was isolated from tonsils of nine sheep and from nasal passages of only one sheep. Two sheep were positive for P. haemolytica biotype A from nasal passages. Culturing tonsillar biopsies as compared to nasal swab samples was a more reliable technique in detecting P. haemolytica, especially biotype T, in bighorn sheep.     

Pathologic changes and microorganisms found in bighorn sheep during a stress-related die-off

An all-age die-off of Rocky Mountain bighorn sheep (Ovis c. canadensis Shaw) occurred from late October 1980 through March 1981 in Waterton Canyon, Colorado, with a loss of 75 to 85% of the sheep. The cause of death was a subacute to chronic bronchopneumonia and the primary etiologic agents isolated from the respiratory system were a Pasteurella sp., P. multocida, Corynebacterium pyogenes, and Protostrongylus stilesi Dikmans, 1931. The underlying predisposing factors that initiated this die-off were believed to be related to multiple chronic environmental stressors associated with the building of a dam which included human contact, vehicular traffic, atmospheric dust, noise and harassment. The die-off was succeeded by a 100% lamb mortality the following summer and a 67% lamb mortality the next two summers. The pneumonia found in these lambs was similar to that found in adult sheep during the previous die-off, except that mature lungworms were absent.     

Using ribosomal RNA gene restriction patterns in distinguishing isolates of Pasteurella haemolytica from bighorn sheep (Ovis canadensis).

Pasteurella haemolytica isolates (n = 31) from two isolated captive herds of Rocky Mountain bighorn sheep (Ovis canadensis canadensis) were characterized and compared phenotypically (biotype, serotype, hemolytic activity) and by a genomic fingerprinting method known as ribotyping. Seven to nine distinct phenotypes were observed. Depending on the method used for serotyping, one to three phenotypes were common to both herds. Eighteen isolates, recovered from both herds, were non-hemolytic, biotype T, indirect hemagglutination assay serotype 4. Ribotyping, a method for highlighting genetically conserved deoxyribonucleic acid restriction site heterogeneity with a 32P-labelled Escherichia coli ribosomal ribonucleic acid probe, produced six to eight distinct ribotype pattern groups within the 31 P. haemolytica isolates, depending on the restriction enzyme used. In contrast to phenotypes, ribotypes appeared unique to each herd, and ribotyping helped to further differentiate some isolates of the same biotype and serotype. In addition, ribotyping provided an alternative means for evaluating relationships between isolates differing in hemolytic activity but which were otherwise phenotypically identical. We propose that ribotyping may be a useful adjunct to other bacterial characterization methods in studying the epizootiology of pasteurellosis in bighorn sheep.     

Evaluation of management treatments intended to increase lamb recruitment in a bighorn sheep herd

We administered a suite of treatments to a herd of Rocky Mountain bighorn sheep (Ovis canadensis canadensis) that was experiencing poor lamb recruitment and showing signs of respiratory disease. Despite 3 yr of treatment with various combinations of anthelmentics, antibiotics, vaccines, and hyperimmune serum products, recruitment was not improved.     

Evaluation of ewe vaccination as a tool for increasing bighorn lamb survival following pasteurellosis epizootics

We conducted field and laboratory experiments to evaluate whether treating pregnant bighorn ewes with a combination of an experimental Pasteurella trehalosi and Mannheimia haemolytica (formerly P. haemolytica) vaccine and a commercially-available bovine P. multocida and M. haemolytica vaccine would increase lamb survival following a pneumonia epidemic. Three free-ranging bighorn herds affected by pasteurellosis outbreaks between November 1995 and June 1996 were included in the field experiment. Post-epidemic lamb survival was low in all three herds in 1996, with November lamb:ewe ratios of < or = 8:100. In March 1997, thirty-six ewes (12/herd) were captured and radiocollared. Half of the ewes captured in each herd were randomly selected to receive both vaccines; the other half were injected with 0.9% saline solution as controls. Lambs born to radiocollared ewes were observed two or more times per week and were considered to have survived if they were alive in October 1997, about 6 mo after birth. Lamb survival differed among herds (range 22% to 100%), and survival of lambs born to vaccinated ewes was lower (P = 0.08) than survival of lambs born to unvaccinated ewes. Bronchopneumonia (pasteurellosis) was the dominant cause of mortality among lambs examined. We concurrently evaluated vaccine effects on survival of lambs born to seven captive ewes removed from the wild during the 1995-96 epidemic. Antibody titers were high in captive ewes prior to vaccination, and vaccines failed to enhance antibody titers in treated captive ewes. None of the captive-born lambs survived. These data suggest that, using existing technology, vaccinating bighorn ewes following pneumonia epidemics has little chance of increasing neonatal survival and population recovery.     

Survival of bighorn sheep (Ovis canadensis) commingled with domestic sheep (Ovis aries) in the absence of Mycoplasma ovipneumoniae

To test the hypothesis that Mycoplasma ovipneumoniae is an important agent of the bighorn sheep (Ovis canadensis) pneumonia that has previously inevitably followed experimental commingling with domestic sheep (Ovis aries), we commingled M. ovipneumoniae-free domestic and bighorn sheep (n=4 each). One bighorn sheep died with acute pneumonia 90 days after commingling, but the other three remained healthy for >100 days. This unprecedented survival rate is significantly different (P=0.002) from that of previous bighorn-domestic sheep contact studies but similar to (P>0.05) bighorn sheep survival following commingling with other ungulates. The absence of epizootic respiratory disease in this experiment supports the hypothesized role of M. ovipneumoniae as a key pathogen of epizootic pneumonia in bighorn sheep commingled with domestic sheep.     

Response of vaccinated and unvaccinated bighorn sheep (Ovis canadensis canadensis) to experimental respiratory syncytial virus challenge

Five Rocky Mountain bighorn sheep (Ovis canadensis canadensis), approximately 5 mo old and without detectable antibody titers to respiratory syncytial virus (RSV), were assigned to two groups to study the effects of RSV challenge inoculation in vaccinated (n = 3) and unvaccinated (n = 2) bighorns. The three lambs vaccinated with a modified live bovine RSV vaccine developed a detectable antibody response to the vaccine. Vaccinated and unvaccinated lambs challenged with an ovine isolate of RSV developed increased levels of neutralizing antibody, but clinical signs of disease were not observed. Neutralizing antibody titers to RSV remained higher (2-4-fold) in vaccinated lambs over time when compared to unvaccinated lambs.     

Sharing of Pasteurella spp. between free-ranging bighorn sheep and feral goats

Pasteurella spp. were isolated from feral goats and free-ranging bighorn sheep (Ovis canadensis canadensis) in the Hells Canyon National Recreation Area bordering Idaho, Oregon, and Washington (USA). Biovariant 1 Pasteurella haemolytica organisms were isolated from one goat and one of two bighorn sheep found in close association. Both isolates produced leukotoxin and had identical electrophoretic patterns of DNA fragments following cutting with restriction endonuclease HaeIII. Similarly Pasteurella multocida multocida a isolates cultured from the goat and one of the bighorn sheep had D type capsules, serotype 4 somatic antigens, produced dermonecrotoxin and had identical HaeIII electrophoretic profiles. A biovariant U(beta) P.haemolytica strain isolated from two other feral goats, not known to have been closely associated with bighorn sheep, did not produce leukotoxin but had biochemical utilization and HaeIII electrophoretic profiles identical to those of isolates from bighorn sheep. It was concluded that identical Pasteurella strains were shared by the goats and bighorn sheep. Although the direction of transmission could not be established, evidence suggests transmission of strains from goats to bighorn sheep. Goats may serve as a reservoir of Pasteurella strains that may be virulent in bighorn sheep; therefore, goats in bighorn sheep habitat should be managed to prevent contact with bighorn sheep. Bighorn sheep which have nose-to-nose contact with goats should be removed from the habitat.