Bighorn sheep × domestic sheep hybrids survive Mannheimia haemolytica challenge in the absence of vaccination.

Bighorn sheep (BHS, Ovis canadensis) are much more susceptible than domestic sheep (DS, Ovis aries) to pneumonia caused by leukotoxin (Lkt)-producing members of the Family Pasteurellaceae, particularly Mannheimia haemolytica and Bibersteinia trehalosi. Leukotoxin is widely accepted as the critical virulence factor of these bacteria since Lkt-negative mutants do not cause death of BHS. Typically, DS carry Lkt-positive M. haemolytica and/or B. trehalosi as commensal bacteria in their nasopharynx. In contrast, most BHS do not carry Lkt-positive M. haemolytica or B. trehalosi, or carry Lkt-negative strains in their nasopharynx. In previous studies, we demonstrated that unimmunized DS resist M. haemolytica challenge while BHS succumb to it. We hypothesized that Lkt-neutralizing antibodies, induced by Lkt-positive M. haemolytica and/or B. trehalosi innately carried by DS in their nasopharynx, render them less susceptible to infection by these bacteria. In this study we developed BHS×DS F1 hybrids by artificial insemination of domestic ewes with BHS semen. F1 hybrids were fertile, and produced F2 hybrids and back-crosses. The F1, F2, and back-crosses were raised together with domestic ewes. All these animals acquired Lkt-positive M. haemolytica and/or B. trehalosi, and developed high titers of Lkt-neutralizing antibodies in the absence of vaccination. Furthermore, all of these animals resisted challenge with lethal dose of M. haemolytica. These results suggest that lack of previous exposure to Lkt is at least partially responsible for fatal pneumonia in BHS when they acquire Lkt-positive M. haemolytica and/or B. trehalosi from DS when the two species commingle.

Mortality in captive elk from salmonellosis.

Salmonella typhimurium DT104 infections of captive elk (Cervus elaphus nelsoni) calves resulted in mortality in eight of 13 affected calves. Salmonellosis in these elk calves was characterized by diarrhea, fever, lethargy, inappetence and depression, and death usually ensued within 72 hr of initial clinical signs. Affected calves did not respond to antibiotic and fluid therapy. The source of the bacteria likely was one or more of the calves when they were captured in the wild at less than 5 days of age. In our captive holding facility, the disease spread quickly and was difficult to control. Phage typing, pulsed field gel electrophoresis, antibiotic sensitivity testing, and plasmid profiles determined that this Salmonella sp. strain was the epidemic strain common to cattle, sheep and humans.

Susceptibility of elk to lungworms from cattle.

Two studies were conducted to determine the infectivity of the lungworm, (Dictyocaulus viviparus) of cattle origin, in Rocky Mountain elk (Cervus elaphus nelsoni) or wapiti. In the first study, each of three 9-mo-old elk was administered 3,000 D. viviparus larvae from cattle using a nasogastric tube. In the second study, four 16-mo-old elk were each inoculated with 2,000 D. viviparus from cattle using a nasogastric tube. Elk were observed daily for signs of respiratory disease, and fecal samples were collected during the studies and evaluated for lungworm larvae using a modified Baermann technique. One elk was euthanatized during the patent period for recovery of adult lungworms, and three elk were euthanatized after larvae were no longer detected in feces. Lungworm larvae were not detected before inoculation in any of the 16-mo-old elk, but were detected 22 days after inoculation in one elk, 23 days after inoculation in two elk and 24 days after inoculation in all four elk. The prepatent period of this cattle isolate of D. viviparus in elk is therefore 22 to 24 days. The precise prepatent period was not determined in the three 9-mo-old elk, but larvae were detected in all three elk 25 days after inoculation. Numbers of larvae ranged from 1/ to 101/g feces with peak larval detection occurring 32 to 50 days after inoculation. Elk shed larvae from 22 to 83 days after inoculation, and patent periods of the parasite ranged from 24 to 62 days. Clinical signs of respiratory disease, with the exception of mild coughing after exercise, were not observed during the infections. Results from this experiment indicated that D. viviparus larvae of cattle origin can mature in elk and larvae can be passed in large numbers in feces, but this cattle isolate of D. viviparus was not highly pathogenic in elk.

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.

Seroprevalence of Toxoplasma gondii in Rocky Mountain bighorn sheep (Ovis canadensis).

Serum samples from 697 Rocky Mountain bighorn sheep (Ovis canadensis) from North America were examined for antibodies to Toxoplasma gondii by the modified agglutination test incorporating mercaptoethanol and formalin-fixed tachyzoites. Antibodies to T. gondii were found in 25 of 697 (3.6%) sheep in titers of 1:25 (8 sheep), 1:50 (4 sheep), 1:100 (7 sheep), 1:200 (1 sheep), 1:400 (1 sheep), 1:800 (1 sheep), and 1:1,600 (3 sheep). This is the first record of T. gondii exposure in bighorn sheep.

Search for the trematode Prouterina wescotti in black bears in Oregon.

Forty-six heads of free-ranging black bears (Ursus americanus) collected in May and June of 1995 and 1996 from Oregon (USA) were examined for the trematode Prouterina wescotti and other parasites. Only Sarcocystis sp. was detected in two adult male bears. Prouterina wescotti has been found only once previously in one black bear from Idaho (USA); its normal definitive host remains unknown.

Contact transmission of psoroptic mange from bighorn to Stone sheep.

Psoroptes sp. mites and tan crusty lesions were detected in the ears of a captive Stone sheep (Ovis dalli stonei) in April 1996, after contact with infected captive Rocky Mountain bighorn sheep (Ovis canadensis canadensis). Psoroptes spp. have not been detected in free-ranging ungulates in Canada or Alaska (USA). This is the first report of Psoroptes sp. in Stone sheep, indicating their susceptibility to infection.

Occurrence of puma lentivirus infection in cougars from Washington.

Puma lentivirus (PLV) antibodies were detected in 13 (25%) of 52 serum samples obtained from cougars (Felis concolor) collected by hunters. The serum samples were collected from November 1993 through January 1994 from four specific regions throughout the state of Washington (USA), and included the Olympic Mountains, the Cascade Mountains, the Blue Mountains, and the Selkirk Mountains. More (38%) seropositive cougar samples originated from the Cascade Mountains than from any other site. The overall seroprevalence for PLV infection in Washington cougars was higher than previously reported for cougars sampled in Oregon and Idaho (USA), but lower than in cougars sampled in Arizona, Colorado, and California (USA).

Experimental contact of bighorn sheep (Ovis canadensis) with horses and cattle, and comparison of neutrophil sensitivity to Pasteurella haemolytica cytotoxins.

Peripheral blood neutrophils from horses, cattle, and Rocky Mountain bighorn sheep (Ovis canadensis canadensis) were evaluated for susceptibility to cytotoxin-dependent lysis of different biotypes and serotypes of Pasteurella haemolytica of domestic sheep, cattle, bighorn sheep, or mountain goat (Oreamnos americana) origin utilizing a cytotoxicity assay which measures the degree of bacteria cytotoxin-killing of neutrophils. All isolates of P. haemolytica (biotypes A and T) were noncytotoxic to horse neutrophils. Thirteen of 18 R haemolytica biotype A isolates were cytotoxic (> 50% neutrophil death in vitro) to bighorn sheep neutrophils, and four of 10 P. haemolytica biotype A isolates were cytotoxic to neutrophils of cattle; P. haemolytica biotype T (= Pasteurella trehelosi) isolates were noncytotoxic to neutrophils of bighorn sheep and cattle. When six bighorn sheep were pastured with three horses, only P. haemolytica biotype T isolates were recovered from the bighorn sheep throughout the study; Pasteurella spp. organisms were not isolated from the three horses. At initiation of a study where five bighorn sheep were pastured with three cattle, P. haemolytica biotype A, serotype 1, 2 was isolated from all three cattle, and only P. haemolytica biotype T isolates were recovered from the bighorn sheep. One bighorn sheep died in each of the horse and cattle copasturing experiments. Pasteurella haemolytica was not isolated from the bighorn sheep which died in the horse copasturing experiment. A noncytotoxic P. haemolytica biotype A, serotype 2 was isolated at necropsy from the bighorn which died in the cattle contact experiment. Based on these experiments, we believe bighorn sheep and horse association would not be detrimental to bighorns due to P. haemolytica induced pneumonia.

Susceptibility of Dall sheep (Ovis dalli dalli) to pneumonia caused by Pasteurella haemolytica.

We evaluated susceptibility of Dall sheep (Ovis dalli dalli) to bacterial pneumonia induced by two strains of Pasteurella haemolytica of domestic sheep origin by evaluating the sensitivity of blood neutrophils of eight Dall sheep to lysis by cytotoxins of P. haemolytica, and by intratracheal inoculation of three Dall sheep, two bighorn sheep (Ovis canadensis), and two domestic sheep with 3.7 x 10(6) or 2.5 x 10(7) colony forming units of P. haemolytica. Neutrophils from the Dall sheep were more sensitive to lysis by cytotoxins from supernatants of a P. haemolytica, biotype A, serotype 2 (A2), of domestic sheep origin, than were neutrophils from six bighorn sheep. This cytotoxic bacterium was the same isolate that was used for intratracheal inoculation of two Dall sheep and two domestic sheep. Inoculation of this cytotoxic P. haemolytica A2 resulted in fatal fibrinopurulent pleuropneumonia in the first Dall sheep within 24 hr of inoculation, and pneumonic lesions in the second Dall sheep before it was euthanized 52 hr after inoculation. This strain of P. haemolytica A2 did not cause respiratory disease when inoculated into two domestic sheep. A noncytotoxic strain of P. haemolytica; biotype T, serotype 3,4,10 of domestic sheep origin did not result in pneumonia in the third Dall sheep or two bighorn sheep. Prior to inoculation, P. haemolytica, biotype T isolates were obtained from all three Dall sheep, but none of these isolates was cytotoxic. At necropsy, cytotoxic P. haemolytica A2 was isolated from lungs and other tissues of the two pneumonic Dall sheep. Based on these results, we conclude that Dall sheep appear to be at least as sensitive as bighorn sheep to pneumonia caused by P. haemolytica A2 of domestic sheep origin. Because in vitro and in vivo results appear closely correlated in this and other studies, we believe with additional evaluation and standardization, neutrophil cytotoxicity tests may serve as a substitute for live animal challenges in future studies of pathogenic P. haemolytica in wild sheep.

Mule deer (Odocoileus hemionus) and elk (Cervus elaphus) as experimental definitive hosts for Fascioloides magna.

In August 1992, six mule deer (Odocoileus hemionus hemionus) fawns and four elk (Cervus elaphus) calves (n = 2) or yearlings (n = 2) each were inoculated orally with 50, 250, or 2,000 metacercariae of the liver fluke Fascioloides magna to evaluate their potential to serve as definitive hosts. Animals were maintained for up to 403 days. Three mule deer each inoculated with 50 metacercariae survived the infection and shed eggs in feces; thus mule deer can function as definitive hosts for F. magna. The other three mule deer inoculated with 50 (n = 1) or 250 (n = 2) metacercariae died from fluke infection on days 91, 150, and 162 days postinoculation, respectively, and only immature F. magna were recovered. One elk calf inoculated with 2,000 metacercariae died from fluke infection 44 days after inoculation. The remaining three elk, each inoculated with 250 metacercariae, survived infection, and two of the three shed eggs in feces. The third elk contained only one immature F. magna at necropsy. The prepatent period in mule deer and elk was approximately 6 to 7 months.

Susceptibility of bighorn sheep (Ovis canadensis) to experimentally-induced Fascioloides magna infections.

Three captive Rocky Mountain bighorn sheep (Ovis canadensis canadensis), consisting of a female lamb, a yearling ram, and a 2 1/2-yr-old castrated ram were inoculated orally with 50 (n = 1) or 100 (n = 2) metacercariae of Fascioloides magna in November 1991. All three sheep died from fluke infection on post-inoculation days 104, 140, and 197, respectively. Numbers of F. magna recovered were 3 (3%), 18 (36%), and 21 (21%). All dukes were immature and were recovered from liver (n = 36), lungs (n = 2), or peritoneal spaces (n = 4). Two white-tailed deer (Odocoileus virginianus), each were inoculated orally with 100 metacercariae at the same time as the bighorn sheep. Eggs of F. magna were detected in the feces of the deer on postinoculation days 199 and 211, respectively. Both deer remained healthy for the year-long experiment. Thus, bighorn sheep are susceptible to infection with F. magna and are likely to die within approximately 6 months of exposure.

Prouterina wescotti n. gen., n. sp. (Trematoda: Prouterinidae N. Fam.) from the brain, lungs, and nasal sinuses of a black bear (Ursus americanus) from Idaho.

Prouterina wescotti gen. n. and sp. n. (Trematoda: Prouterinidae N. Fam.) is described from a free-ranging black bear (Ursus americanus) which died in May 1995 in northern Idaho (USA). Adult digenetic trematodes were detected in brain, lungs, and nasal sinuses, and were likely responsible for the emaciated condition, copious nasal discharge, neurological signs, and death of the bear. Mature trematodes recovered from the bear were conical with small spines on the tegument. The anterior end was broad and tapered gradually toward the posterior. Mean (+/-SE) size of the mature trematodes was 3.67 (+/-0.08) by 2.14 (+/-0.04) mm (n = 80). Eggs are operculated, gold, and 68.2 (+/-0.42) by 41.4 (+/-0.41) microns (n = 75). Suckers are well developed and located in the anterior half of the body, with the genital pore just posterior to the ventral sucker. Testes are tandem and the ovary is lateral and slightly anterior to the anterior testes just posterior to the ventral sucker. The uterus is predominantly anterior to the ventral sucker and is the most distinctive feature of the trematode.

Attempted protection of bighorn sheep (Ovis canadensis) from pneumonia using a nonlethal cytotoxic strain of Pasteurella haemolytica, biotype A, serotype.

Between February and April, 1994, we tested the hypothesis that bighorn sheep (Ovis canadensis canadensis) inoculated with a cytotoxic isolate of Pasteurella haemolytica biotype A, serotype 11 (A11) could withstand challenge inoculation with a cytotoxic strain of P. haemolytica A2 of domestic sheep origin known to cause lethal pneumonia in bighorn sheep. On experimental day O, two bighorn sheep were inoculated intratracheally with 6 x 10(9) colony forming units (cfu) of a cytotoxic strain of P. haemolytica A11 (group 1); two bighorn sheep were inoculated intratracheally with 6 x 10(9) cfu of a noncytotoxic P. haemolytica A11 (group 2), and two control bighorn sheep were inoculated intratracheally with a similar volume of brain heart infusion (BHI) broth (group 3). After inoculation, all bighorn sheep remained healthy. On experimental day 16, group 1 bighorn sheep each were given the same intratracheal inoculation as on day O, and groups 2 and 3 bighorn sheep each were inoculated with BHI broth at the same volume as group 1. All bighorn sheep remained healthy following inoculations. On experimental day 42, bighorn sheep in groups 1 and 3 each were challenged with an intratracheal inoculation of 6 x 10(9) cfu of P. haemolytica A2 of domestic sheep origin known to be lethal in bighorn sheep. Group 2 sheep each were inoculated intratracheally with BHI broth at the same volume as groups 1 and 3. The four bighorn sheep in groups 1 and 3 that received the challenge inoculation died from acute bronchopneumonia within 72 hours after challenge inoculation, and cytotoxic P. haemolytica A2 was isolated from the four dead bighorn sheep. Both cytotoxic or noncytotoxic strains of P. haemolytica A11 were not lethal and did not cause pneumonia in the experimentally inoculated bighorn sheep. However, previous inoculation with cytotoxic P. haemolytica A11 did not protect the bighorn sheep against later experimental challenge inoculation with a known lethal strain of cytotoxic P. haemolytica A2 under the conditions defined in these experiments.

Experimental infections of Sarcocystis spp. in Rocky Mountain elk (Cervus elaphus) calves.

Four 4-mo-old elk calves (Cervus elaphus) obtained from northeastern Oregon (USA) each were inoculated orally with 250,000 sporocysts of Sarcocystis spp., including S. sybillensis and S. wapiti. Three similar elk calves of comparable ages and weights served as uninoculated controls maintained with the inoculated elk during the experimental period between September and December 1993. Body weights were evaluated at 0 and 90 days postinoculation (PI); packed cell volumes of whole blood were evaluated at 0, 30, and 60 days PI, and numbers of sarcocysts in histologic sections from 11 selected tissues were evaluated at 90 days PI. Significant differences in blood packed cell volumes were not detected between groups (P > 0.05). Except for weight gain, elk remained healthy. Mean (+/- SE) weight gain of inoculated elk (27.1 +/- 1.6 kg) was significantly (P < 0.05) less than that of controls (40.2 +/- 4.9 kg). Mean (+/- SE) number of sarcocysts in tissues of inoculated (114.4 +/- 25.7 cm2) and controls (4.5 +/- 1.4 cm2) differed significantly (P < 0.05). Heart, esophagus and skeletal muscle contained the most sarcocysts. No sarcocysts were detected in brain, spinal cord, or testicles. Histologically, mononuclear myositis and myocarditis, with numerous intralesional sarcocysts were seen. Less severe, but widespread inflammation occurred in brain, spinal cord, and optic nerve. Mortality and anemia were not seen, but weight gain depression was detected in the inoculated elk over the 90 day experimental period.

Susceptibility of phagocytes from elk, deer, bighorn sheep, and domestic sheep to Pasteurella haemolytica cytotoxins.

Alveolar macrophages and peripheral blood neutrophils from elk (Cervus elaphus), bighorn sheep (Ovis canadensis canadensis), and domestic sheep were exposed to culture supernatants from Pasteurella haemolytica isolated from bighorn sheep and domestic sheep. In a second experiment, peripheral blood neutrophils from mule deer (Odocoileus hemionus), elk, and bighorn sheep were exposed to culture supernatants from P. haemolytica isolated from elk, bighorn sheep and domestic sheep. Alveolar macrophages from elk, bighorn sheep and domestic sheep were resistant to killing by P. haemolytica supernatants from bighorn sheep and domestic sheep; susceptibility of neutrophils to cell death, as measured by release of lactate dehydrogenase, differed significantly (P < 0.05) between the four species tested. Bighorn sheep and domestic sheep neutrophils were susceptible to cytotoxin damage by the P. haemolytica isolates used; bighorn sheep neutrophils were four- to eight-fold more susceptible to cytotoxin damage than domestic sheep neutrophils. Neutrophils from deer and elk were resistant to killing by P. haemolytica cytotoxins from any species tested.

Comparative leukotoxicities of Pasteurella haemolytica isolates from domestic sheep and free-ranging bighorn sheep (Ovis canadensis).

Twenty-eight isolates of Pasteurella haemolytica from domestic sheep (n = 14 isolates) and bighorn sheep (n = 14 isolates) were evaluated for leucotoxicity against peripheral blood neutrophils of bighorn sheep by adding bacterial culture supernatants to bighorn sheep neutrophils in vitro. Leukotoxic isolates of P. haemolytica, defined as causing > 50% neutrophil death as measured by release of lactate dehydrogenase into culture supernatants, were identified from eight of 14 domestic sheep isolates and from 0 of 14 bighorn sheep isolates. The in vitro assay of isolates of P. haemolytica may provide a valid predictive measure of strain virulence of P. haemolytica, and of potential pneumonic episodes in bighorn sheep populations.

Experimental infections of Eimeria wapiti and E. zuernii-like oocysts in Rocky Mountain elk (Cervus elaphus) calves.

Four Rocky Mountain elk (Cervus elaphus) 10 to 14 wk of age each were inoculated orally with a mixture of 50,000 sporulated oocysts of an Eimeria zuernii-like apicomplexan (70%) and E. wapiti (30%). Maximum numbers of oocysts per gram of feces (OPG) in each elk ranged from 985 to 15,517, but all calves remained healthy and clinical signs of coccidiosis were not observed. The prepatent period for E. zuernii was 8 days and the patent period approximately 37 days, with a maximum mean (+/- SE) recovery of 6,643 (+/- 3,756) OPG on post-inoculation day 8. The prepatent period for E. wapiti was 10 to 12 days and the patent period approximately 8 days, with a maximum mean recovery of 4,408 (+/- 2,308) OPG on postinoculation day 12. Based on these data, infections of E. zuernii and E. wapiti at the numbers given were not pathogenic in healthy elk calves.

Bot fly larvae (Cephenemyia jellisoni) as a cause of neurologic signs in an elk.

In June 1993, a yearling female elk (Cervus elaphus) near John Day, Oregon (USA) was observed twice over a four week period with signs of neurologic disease including weakness, walking in circles with an uncoordinated gait and disorientation. The elk was shot, and the head and neck were examined grossly for parasites and lesions. Thirty-five second and third instar larvae of Cephenemyia jellisoni were recovered from an encapsulated space in the nasopharyngeal area dorsal to the soft palate. Larvae protruded into the caudodorsal end of the ventral nasal meatus, obliterating the opening of the left eustachian tube. Larvae were not recovered from their normal location in the retropharyngeal recesses. Thus the effects of several Cephenemyia jellisoni larvae in an aberrant location mimicked signs observed in meningeal worm infections.