Pasteurellaceae isolated from bighorn sheep (Ovis canadensis) from Idaho, Oregon, and Wyoming.

OBJECTIVE: To elucidate the species and biovariants of Pasteurellaceae isolated from clinically normal bighorn sheep (Ovis canadensis) or bighorn sheep with evidence of respiratory disease. SAMPLE: 675 Pasteurellaceae isolates from 290 free-ranging bighorn sheep in Idaho, Oregon and Wyoming. PROCEDURES: Nasal and oropharyngeal swab specimens were inoculated onto selective and nonselective blood agar media. Representatives of each colony type were classified via a biovariant scheme. The association of respective ß-hemolytic isolates with respiratory disease was evaluated via χ(2) analyses. RESULTS: Bacterial isolates belonged to 4 species: Histophilus somni, Mannheimia haemolytica, Pasteurella multocida, and Bibersteinia (Pasteurella) trehalosi. Within the latter 3 species, 112 subspecies, biotypes, and biovariants were identified. Bibersteinia trehalosi 2 and B trehalosi 2B constituted 345 of 675 (51%) isolates. Most (597/618 [97%]) isolates from adult sheep were from clinically normal animals, whereas most (47/57 [82%]) isolates from lambs were from animals with evidence of respiratory disease. Twenty-two Pasteurellaceae biovariants were isolated from sheep with respiratory disease; 17 of these biovariants were also isolated from clinically normal sheep. The ability of isolates to cause ß-hemolysis on blood agar was associated with respiratory disease in adult bighorn sheep (OR, 2.59; 95% confidence interval, 1.10 to 6.07). CONCLUSIONS AND CLINICAL RELEVANCE: Bighorn lambs appeared more susceptible to respiratory disease caused by Pasteurellaceae than did adult sheep. ß-Hemolytic Pasteurellaceae isolates were more likely to be associated with respiratory disease than were non-ß-hemolytic isolates in adult sheep. Identification of Pasteurellaceae with the greatest pathogenic potential will require studies to estimate the risk of disease from specific biovariants.

Detection of Mycoplasma ovipneumoniae and M. arginini in bighorn sheep using enrichment culture coupled with genus- and species-specific polymerase chain reaction.

Mycoplasma species are of interest as possible primary pathogens in the pneumonia complex of bighorn sheep (Ovis canadensis). Previous investigations have not commonly detected low frequencies of Mycoplasma spp. from free-ranging bighorn sheep, possibly due to the fastidious and slow growth of these organisms. We developed a culture protocol that employed an average initial 3-day enrichment culture in liquid Hayflick broth in a CO(2)-enhanced atmosphere. The broth was plated to solid Hayflick medium and the cultures observed for growth for up to 30 days. Polymerase chain reaction (PCR) was performed on DNA isolated from the enrichment broth and on isolates obtained from culture using Mycoplasma genus-specific PCR assays and species-specific PCR assays for M. arginini and M. ovipneumoniae. Some cultures that grew on Hayflick plates were picked as single colonies but were mixed because two organisms may grow together and appear as a single colony. Culture and PCR tests produced similar results for M. arginini, but for M. ovipneumoniae, culture alone was less accurate than PCR. Use of genus-specific primers also may allow detection of other species in samples negative for M. arginini and M. ovipneumoniae. Two methods of transport from field to laboratory (Port-a-Cul™ tubes, cryoprotectant in liquid N(2) and Fisher Transport System) gave similar results under our study conditions.

Domestic sheep (Ovis aries) Pasteurellaceae isolates from diagnostic submissions to the Caine Veterinary Teaching Center (1990-2004).

A retrospective study of Pasteurellaceae isolated from domestic sheep (Ovis aries) was conducted. The aim was to identify Pasteurellaceae present in animals that were clinically healthy and others with evidence of respiratory disease. The bacteria had been isolated from samples submitted to the University of Idaho Caine Veterinary Teaching Center as part of disease diagnostic testing. The 844 isolates identified mainly three species of Pasteurellaceae: Mannheimia haemolytica, Pasteurella multocida, and Pasteurella (Bibersteinia) trehalosi. A total of 114 biovariants were identified among these three species. Individual biovariants were identified 1-180 times. Two of those (M. haemolytica 1 and P. (B.) trehalosi 2) constituted 36% of the isolates, and were the only biovariants sufficiently numerous to account for >7% of the total isolates. Samples were primarily submitted from sheep with signs of respiratory disease. Eighty percent of biovariants were identified most often in animals with signs of respiratory disease, but 26% of biovariants were isolated from both sheep with respiratory disease and apparently healthy sheep. P. multocida constituted 4.7% of isolates, and were exclusively associated with animals with respiratory disease. The ability of isolates to produce beta-hemolysis on culture media was not associated with animals with respiratory disease (odds ratio 0.77, 95% CI 0.50-1.19). The inference of this study is limited due to the retrospective study design. However, it is the first study that provides an extensive baseline list of biovariants associated with respiratory disease in domestic sheep.

Variation in Pasteurella (Bibersteinia) and Mannheimia spp. following transport and antibiotic treatment in free-ranging and captive Rocky Mountain bighorn sheep (Ovis canadensis canadensis).

Morbidity and mortality associated with respiratory disease following capture and translocation of bighorn sheep (Ovis canadensis canadensis) is a significant concern, particularly when establishing new or augmenting existing bighorn populations. Administration of prophylactic antibiotics at the time of capture is often done to minimize the risk of respiratory disease, but the efficacy of this practice is unknown. The effects of oxytetracycline and florfenicol on the Pasteurella (Bibersteinia) and Mannheimia spp. isolated from samples collected from the oropharynx at the time of capture and 3 or 42 day later were evaluated in two groups of bighorn sheep. The most evident change in the isolation rates or types of Pasteurella (Bibersteinia) spp., Mannheimia spp., or both was an increase of beta-hemolytic strains isolated from bighorn sheep 3 day following oxytetracycline treatment. Both groups of bighorn sheep carried Pasteurella (Bibersteinia) trehalosi identified as the same biovariants, but they did not share biovariants of Mannheimia spp. No animals had signs of respiratory disease. Isolates representative of all biovariants present in cultures from the two bighorn sheep groups were sensitive to in vitro tests to both oxytetracycline and florfenicol and the majority were also sensitive to seven other antibiotics tested. The administration of neither oxytetracycline nor florfenicol eliminated Pasteurella (Bibersteinia) or Mannheimia from the oropharyngeal mucosa. Resistance to either antibiotic used in these animals was not noted. Although the prophylactic benefits of these drugs in preventing disease are uncertain, therapeutic levels of antibiotics in lung tissue during times of stress may reduce the risk of disease. Representative sampling of the oropharyngeal microflora of bighorn sheep source and recipient populations prior to being intermingled should be considered as one of the tools to minimize exposure of naive populations to potentially pathogenic bacteria.

Microorganisms associated with a pneumonic epizootic in Rocky Mountain bighorn sheep (Ovis canadensis canadensis).

A comprehensive study of a pneumonic epizootic was initiated when the first signs of disease were noted in a metapopulation of bighorn sheep inhabiting Hells Canyon, bordering Idaho, Oregon, and Washington. A total of 92 bighorn sheep were tested for etiologic agents during the following 6-mo study period. The study population included bighorn sheep believed to be the subpopulation in which disease was first noted, and these sheep were translocated to a holding facility in an effort to contain the disease (group A1, n = 72); bighorn sheep in other subpopulations (group A2) with evidence of clinical disease were captured, sampled, given antibiotics, and released (n = 8) and those that were found dead were necropsied (n = 12). Samples, including oropharyngeal and nasal swabs, and lung and liver tissue were collected from the bighorn sheep identified above. Tissue was collected at necropsy from 60 group A1 bighorn sheep that died following translocation, and samples were cultured for bacteria and viruses. Blood samples were tested for antibodies against known respiratory viruses, and histopathology was conducted on tissue samples. The major cause of death in both group A1 and group A2 bighorn sheep was a rapidly developing fibrinous bronchopneumonia. Multiple biovariants of Pasteurella were isolated from oropharyngeal and nasal samples from both groups, and Mycoplasma ovipneumonia was isolated from five group A1 oropharyngeal samples. Organisms isolated from lung tissue included Pasteurella multocida multocida a and Pasteurella trehalosi, both of which differentiated into multiple strains by restriction enzyme analysis, and parainfluenza-3 virus (PI-3). Paired serum samples revealed > fourfold increases in titers against PI-3 and bovine respiratory syncytial viruses. It was concluded that this epizootic resulted from a complex of factors including multiple potential respiratory pathogens, none of which were identified as a primary pathogen, and possible stress factors.

Haemophilus somnus (Histophilus somni) in bighorn sheep.

Respiratory disease and poor lamb recruitment have been identified as limiting factors for bighorn-sheep populations. Haemophilus somnus (recently reclassified as Histophilus somni) is associated with respiratory disease in American bison, domestic sheep, and cattle. It is also harbored in their reproductive tracts and has been associated with reproductive failure in domestic sheep and cattle. Therefore, reproductive tract and lung samples from bighorn sheep were evaluated for the presence of this organism. Organisms identified as H. somnus were isolated from 6 of 62 vaginal but none of 12 preputial swab samples. Antigen specific to H. somnus was detected by immunohistochemical study in 4 of 12 formalin-fixed lung tissue samples of bighorn sheep that died with evidence of pneumonia. Notably, H. somnus was found in alveolar debris in areas of inflammation. The 6 vaginal isolates and 2 H. somnus isolates previously cultured from pneumonic lungs of bighorn sheep were compared with 3 representative isolates from domestic sheep and 2 from cattle. The profiles of major outer membrane proteins and antigens for all of the isolates were predominantly similar, although differences that may be associated with the host-parasite relationship and virulence were detected. The DNA restriction fragment length profiles of the bighorn-sheep isolates had similarities not shared with the other isolates, suggesting distinct phylogenetic lines. All of the isolates had similar antimicrobial profiles, but the isolates from the bighorn sheep produced less pigment than those from the domestic livestock, and growth of the former was not enhanced by CO2. Wildlife biologists and diagnosticians should be aware of the potential of these organisms to cause disease in bighorn sheep and of growth characteristics that may hinder laboratory detection.

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.

Characterization of Pasteurella spp isolated from healthy domestic pack goats and evaluation of the effects of a commercial Pasteurella vaccine.

OBJECTIVE: To characterize Pasteurella spp isolated from healthy pack goats and evaluate the effects of administration of a commercial Pasteurella vaccine. ANIMALS: 45 goats. PROCEDURE: Pharyngeal swab specimens and blood samples were collected on day 0 before vaccination with a Pasteurella (Mannheimia) haemolytica serotype A1 bacterin. Samples were also collected from 17 goats on days 21 and 35. Isolated Pasteurella spp were assigned to biovariant groups on the basis of results of biochemical utilization tests and serotyped. Serum antibody titers were determined. RESULTS: Multiple strains of Pasteurella spp were isolated from swab specimens and assigned to 30 nonhemolytic and 14 beta-hemolytic biovariant groups. The most common biovariant isolated was nonhemolytic P trehalosi belonging to group 2. This strain was isolated from 41 goats. Nonhemolytic P haemolytica strains were isolated from 31 goats, whereas beta-hemolytic strains of P trehalosi and P haemolytica were isolated from 8 and 35 goats, respectively. Vaccination with the A1 serotype did not affect the proportion of goats from which we isolated each biovariant group or the number of biovariant groups isolated. CONCLUSIONS AND CLINICAL RELEVANCE: Multiple strains of P haemolytica and P trehalosi belonging to nonhemolytic and beta-hemolytic biovariant groups were isolated from the pharynx of healthy domestic pack goats. Because hemolytic activity correlates with leukotoxin production, beta-hemolytic strains may have a greater potential to cause disease in naive populations of wild ruminants. However, vaccination with an A1 serotype bacterin did not decrease the proportion of culture-positive goats.