Frequent transmission of immunodeficiency viruses among bobcats and pumas.

With the exception of human immunodeficiency virus (HIV), which emerged in humans after cross-species transmissions of simian immunodeficiency viruses from nonhuman primates, immunodeficiency viruses of the family Lentiviridae represent species-specific viruses that rarely cross species barriers to infect new hosts. Among the Felidae, numerous immunodeficiency-like lentiviruses have been documented, but only a few cross-species transmissions have been recorded, and these have not been perpetuated in the recipient species. Lentivirus seroprevalence was determined for 79 bobcats (Lynx rufus) and 31 pumas (Puma concolor) from well-defined populations in Southern California. Partial genomic sequences were subsequently obtained from 18 and 12 seropositive bobcats and pumas, respectively. Genotypes were analyzed for phylogenic relatedness and genotypic composition among the study set and archived feline lentivirus sequences. This investigation of feline immunodeficiency virus infection in bobcats and pumas of Southern California provides evidence that cross-species infection has occurred frequently among these animals. The data suggest that transmission has occurred in multiple locations and are most consistent with the spread of the virus from bobcats to pumas. Although the ultimate causes remain unknown, these transmission events may occur as a result of puma predation on bobcats, a situation similar to that which fostered transmission of HIV to humans, and likely represent the emergence of a lentivirus with relaxed barriers to cross-species transmission. This unusual observation provides a valuable opportunity to evaluate the ecological, behavioral, and molecular conditions that favor repeated transmissions and persistence of lentivirus between species.

Ants as first intermediate hosts of Mesocestoides on San Miguel Island, USA.

This study tested the hypotheses that ants (Formicidae) function as a first intermediate host of Mesocestoides (Cestoda: Mesocestoididae) and that deer mice (Peromyscus maniculatus) develop metacestode infections after ingesting cysticercoid or procercoid-infected ants. Field studies were conducted at an island fox (Urocyon littoralis littoralis) breeding facility located on San Miguel Island, California Channel Islands National Park, USA, where > 40% of captive foxes were infected with adult Mesocestoides. Eight percent (8%) of deer mice at the fox pen site were infected with Mesocestoides metacestodes while none were infected at a distant site where foxes were absent (campground), thereby indicating the potential localized presence of a first intermediate host. To test whether ants from San Miguel Island contained Mesocestoides DNA, a polymerase chain reaction (PCR)-based diagnostic assay was developed using nested primers that could detect a single hexacanth larva within pooled samples of ten ants. Ants (Lasius niger and Tapinoma sessile) collected near the fox breeding facility were tested using the nested-PCR assay. Seven of 223 pooled samples of L. niger (3.1%) and 2 of 84 pooled samples of T. sessile (2.4%) tested positive for Mesocestoides DNA, while none of the ants were positive at the campground site. Positive samples were sequenced and found to match DNA sequences from Mesocestoides obtained from island fox and deer mice. Finally, to determine whether ants function as a first intermediate host for Mesocestoides, colony-raised deer mice (n = 47) were fed L. niger (n = 3860) or T. sessile (n = 339) collected from the San Miguel Island fox breeding facility. No mouse became infected with Mesocestoides metacestodes after ingesting ants. While both L. niger and T. sessile from SMI were positive for Mesocestoides DNA, they were not infective to deer mice in the laboratory.

Is the decline of desert bighorn sheep from infectious disease the result of low MHC variation?

Bighorn sheep populations have greatly declined in numbers and distribution since European settlement, primarily because of high susceptibility to infectious diseases transmitted to them from domestic livestock. It has been suggested that low variation at major histocompatibility complex (MHC) genes, the most important genetic aspect of the vertebrate immune system, may result in high susceptibility to infectious disease. Therefore, we examined genetic polymorphism at a MHC gene (Ovca-DRB) in a large sample, both numerically and geographically, of bighorn sheep. Strikingly, there were 21 different alleles that showed extensive nucleotide and amino acid sequence divergence. In other words, low MHC variation does not appear to be the basis of the high disease susceptibility and decline in bighorn sheep. On the other hand, analysis of the pattern of the MHC polymorphism suggested that nonsynonymous substitutions predominated, especially at amino acids in the antigen-binding site. The average overall heterozygosity for the 16 amino acid positions that are part of the antigen binding site is 0.389 whereas that for the 67 amino acid positions not involved with antigen binding is 0.076. These findings imply that the diversity present in this gene is functionally significant and is, or has been, maintained by balancing selection. To examine the evolution of DRB alleles in related species, a phylogenetic analysis including other published ruminant (Bovidae and Cervidae) species, was carried out. An intermixture of sequences from bighorn sheep, domestic sheep, goats, cattle, bison, and musk ox was observed supporting trans-species polymorphism for these species. To reconcile the species and gene trees for the 104 sequences examined, 95 'deep coalescent' events were necessary, illustrating the importance of balancing selection maintaining variation over speciation events.

Prevalence of bovine tuberculosis in African buffalo at Kruger National Park.

Bovine tuberculosis (BTB) was first detected in Kruger National Park (KNP) in a single African buffalo (Syncerus caffer) in 1990. In 1991/1992, 2,071 African buffalo were examined for BTB as part of a culling program that removed animals from all known herds in KNP. The prevalence of BTB in 1991/1992 was estimated to be 0%, 4.4% (+/-0.6%), and 27.1% (+/-1.4%), in the north, central, and south zones of KNP, respectively. In 1998, a stratified, two-stage cluster sampling method was used to estimate that the prevalence of BTB was 1.5% (+/-2.5%), 16% (+/-5.3%), and 38.2% (+/-6.3%), in the north, central, and south zones, respectively. This represented a significant increase in prevalence (P < or = 0.05) in the south and central zones, but not in the north zone. Continued monitoring of BTB in KNP is important for understanding disease transmission risks, potential population effects, and the efficacy of disease management strategies. The methodology and sample sizes used in 1998 are appropriate for future BTB monitoring in KNP.

Epidermoptid mange in Laysan albatross fledglings in Hawaii.

Mange caused by the epidermoptid mite Myialges nudus (Acari: Epidermoptidae) is described in 31 dead fledgling Laysan albatrosses (Phoebastria immutabilis) from Midway Atoll (Hawaii, USA) sampled from 18 June to 10 July 1990 and from 21 June to 22 July 1991. This is the first record for this parasite from this host. Mites were collected from the skin; were located primarily in the stratum corneum; and were associated with mild to severe granulomatous inflammation, hyperkeratosis, dermal edema, ballooning degeneration of keratinocytes, neovascularization, and subdermal fibrosis. The severity of inflammation in some birds suggested that dermatitis due to M. nudus could be a significant cause of morbidity, or even mortality, in these birds.

Phylogeny and host specificity of Psoroptic mange mites (Acarina: Psoroptidae) as indicated by ITS sequence data.

We used molecular phylogenetic techniques to study the systematic relationships and host specificity of Psoroptes mange mites, which are pests of numerous domestic and wild ungulates. Phylogenetic analysis of DNA sequence data from the internal transcribed spacer region 1 (ITS1) of nuclear ribosomal DNA indicated that populations of Psoroptes are not host specific. Furthermore, the currently used taxonomy of Psoroptes is not concordant with the phylogeny derived from ITS1. During the course of the study, we discovered apparent paralogous ITS sequences within individual mites as a result of varying polymerase chain reaction reaction conditions. This finding concords with other studies of ITS and suggests a cautious approach when interpreting data from ITS sequences. Host DNA contamination was also found to be a significant problem in data collection, and we report on the development of methods to overcome the problems of contamination in parasitic mites.

Evaluation of changes in hematologic and clinical biochemical values after exposure to petroleum products in mink (Mustela vison) as a model for assessment of sea otters (Enhydra lutris).

OBJECTIVE: To determine the effects of petroleum exposure on hematologic and clinical biochemical results of mink and to identify variables that may be useful for making management decisions involving sea otters (Enhydra lutris) that have been exposed to oil in their environment. ANIMALS: 122 American mink (Mustela vison). PROCEDURES: Mink were exposed once to a slick of oil (Alaskan North Slope crude oil or bunker C fuel oil) on seawater or via low-level contamination of their daily rations. RESULTS: In the acute phase of exposure, petroleum directly affected RBC, WBC, neutrophil, and lymphocyte counts, fibrinogen, sodium, calcium, creatinine, total protein, and cholesterol concentrations, and alanine transaminase, creatine kinase, alkaline phosphatase, and gamma-glutamyltransferase activities. Aspartate transaminase, alkaline phosphatase, gamma-glutamyltransferase, and lactate dehydrogenase activities and cholesterol concentration also varied as a result of chronic low-level contamination of feed. CONCLUSIONS AND CLINICAL RELEVANCE: Our results are in agreement with reports that attribute increased alanine transaminase and alkaline phosphatase activities and decreased total protein concentration to petroleum exposure in sea otters during an oil spill. Sodium, calcium, creatinine, cholesterol, and lactate dehydrogenase may be valuable variables to assess for guidance during initial treatment of sea otters exposed to oil spills as well as for predicting which petroleum-exposed sea otters will reproduce following an oil spill. Measurement of these variables should aid wildlife professionals in making decisions regarding treatment of sea otters after oil spills.

Bartonella spp. isolated from wild and domestic ruminants in North America.

Bartonella species were isolated from 49% of 128 cattle from California and Oklahoma, 90% of 42 mule deer from California, and 15% of 100 elk from California and Oregon. Isolates from all 63 cattle, 14 deer, and 1 elk had the same polymerase chain reaction/restriction fragment length polymorphism profiles. Our findings indicate potential for inter- and intraspecies transmission among ruminants, as well as risk that these Bartonella spp. could act as zoonotic agents.

Biochemical and hematologic reference intervals for free-ranging desert bighorn sheep.

Over 200 clinically normal desert bighorn sheep (Ovis canadensis) from multiple geographic areas were sampled utilizing a uniform protocol. The goals of this study were to develop comprehensive reference intervals for hematologic and biochemical analytes using central 90th percentile nonparametric analyses. Adult female sheep had greater erythrocyte mass (hemoglobin and hematocrit) compared with adult male sheep. Young animals < or = 1-yr-old had greater erythrocyte mass (hemoglobin, hematocrit and red blood cell count), higher alkaline phosphatase activity, and lower serum protein and globulin concentrations compared with adult animals. Because of the large sample size, wide geographic range, and uniform sample and handling protocol in this study, these reference intervals should be robust and applicable to other free-ranging desert bighorn sheep populations.

Molecular systematics of Mesocestoides sPP (cestoda: mesocestoididae) from domestic dogs (Canis familiaris) and coyotes (Canis latrans).

The genus Mesocestoides Vaillant, 1863 includes tapeworms of uncertain phylogenetic affinities and with poorly defined life histories. We previously documented 11 cases of peritoneal cestodiasis in dogs (Canis familiaris L.) in western North America caused by metacestodes of Mesocestoides spp. In the current study, DNA sequences were obtained from metacestodes collected from these dogs (n = 10), as well as proglottids from dogs (n = 3) and coyotes (Canis latrans Say, 1823 [n = 2]), and tetrathyridia representing laboratory isolates of M. corti (n = 3), and these data were analyzed phylogenetically. Two nuclear genetic markers, 18S ribosomal DNA and the second internal-transcribed spacer (ITS 2), were sequenced. Phylogenetic analysis of the 18S rDNA data recovered a monophyletic group composed of all samples of Mesocestoides spp., distinct from closely related outgroup taxa (Amurotaenia Akhmerov, 1941 and Tetrabothrius Rudolphi, 1819). Initial analysis of the ITS 2 data resolved 3 clades within Mesocestoides. Two proglottids from dogs formed a basal clade, a second clade was represented by tetrathyridial isolates, and a third clade included all other samples. Interpretation of these data from an apomorphy-based perspective identified 6 evolutionary lineages. We also assessed whether metacestodes from dogs (n = 4) are capable of asexual proliferation in laboratory mice. One tetrathyridial and 2 acephalic isolates from dogs proliferated asexually. Further investigation is warranted to determine which of the lineages represent distinct species and to determine the life history strategies of Mesocestoides spp.

Molecular tracking of mountain lions in the Yosemite valley region in California: genetic analysis using microsatellites and faecal DNA.

Twelve microsatellite loci were characterized in California mountain lions (Puma concolor) and sufficient polymorphism was found to uniquely genotype 62 animals sampled at necropsy. Microsatellite genotypes obtained using mountain lion faecal DNA matched those from muscle for all of 15 individuals examined. DNA from potential prey species and animals whose faeces could be misidentified as mountain lion faeces were reliably distinguished from mountain lions using this microsatellite panel. In a field application of this technique, 32 faecal samples were collected from hiking trails in the Yosemite Valley region where seven mountain lions previously had been captured, sampled, and released. Twelve samples yielded characteristic mountain lion genotypes, three displayed bobcat-type genotypes, and 17 did not amplify. The genotype of one of the 12 mountain lion faecal samples was identical to one of the mountain lions that previously had been captured. Three of the 12 faecal samples yielded identical genotypes, and eight new genotypes were detected in the remaining samples. This analysis provided a minimum estimate of 16 mountain lions (seven identified by capture and nine identified by faecal DNA) living in or travelling through Yosemite Valley from March 1997 to August 1998. Match probabilities (probabilities that identical DNA genotypes would be drawn at random a second time from the population) indicated that the samples with identical genotypes probably came from the same mountain lion. Our results demonstrate that faecal DNA analysis is an effective method for detecting and identifying individual mountain lions.

An epizootic of adenovirus-induced hemorrhagic disease in captive black-tailed deer (Odocoileus hemionus).

Ten fawns and four adult black-tailed deer (Odocoileus hemionus) in a captive herd died as a result of adenovirus-induced hemorrhagic disease. Acute, systemic infections were characterized by hemorrhagic enteropathy, pulmonary edema, and occasional ulceration of the upper alimentary tract. Localized infections were limited to the upper alimentary tract and included stomatitis, pharyngitis, mandibular osteomyelitis, and rumenitis. In deer with acute, systemic infections, a diagnosis was made by histopathology and immunohistochemistry. The serum neutralization test was useful for confirming a diagnosis in animals with prolonged, localized infections. Deer originating from herds with a history of adenovirus infection should not be transferred to other captive herds or released into free-ranging populations because they may serve as carriers of adenovirus.

Dermacentor hunteri (Acari: Ixodidae): an experimental vector of Anaplasma marginale and A. ovis (Rickettsiales: Anaplasmataceae) to calves and sheep.

The experimental vector competence of laboratory-reared Dermacentor hunteri Bishopp for Anaplasma marginale Theiler and Anaplasma ovis Lestoquard was evaluated by delayed transfer of male ticks from infected to susceptible Holstein calves and from infected to susceptible domestic sheep, respectively. After feeding for 4 or 5 d on rickettsemic acquisition hosts, the ticks were held off the host at 26 degrees C, approximately 93% RH, and a photoperiod of 14:10 (L:D) h for 7 or 8 d, then test fed for 5 or 7 d. Additionally, ticks test-fed for 5 d on 2 susceptible calves were removed, held off the host for 7 d, and test-fed for 5 d on a 3rd susceptible calf to test the tick's ability to transmit A. marginale by delayed serial transfer. Tick transmission of A. marginale to 3 test calves and A. ovis to 3 test sheep was demonstrated by blood smear and indirect immunofluorescence serology. These data indicate that males of D. hunteri, a tick commonly found on desert bighorn, Ovis canadensis Shaw, in the southwestern United States and northern Mexico, may be competent natural vectors of these organisms present in desert bighorn populations.

Population subdivision among desert bighorn sheep (Ovis canadensis) ewes revealed by mitochondrial DNA analysis.

We used behavioural observations and mitochondrial DNA (mtDNA) sequence analysis to examine demographic and genetic structure within and among home-range groups of desert bighorn sheep (Ovis canadensis) ewes in the Peninsular Ranges of southern California, USA. We identified substantial genetic variation in the first 515 bp of the mtDNA control region and determined that seven haplotypes were distributed in a nonrandom fashion among these ewe subpopulations. Although a significant (P < 0.01) amount of mtDNA variation (33%) was partitioned among home-range groups, we did not find strong evidence for matrilineal substructuring within these groups. Based on analyses of molecular variance, and comparisons of behavioural associations and distances between centres of activity, we concluded that within a given home-range group, bighorn sheep ewes generally associate with other ewes based on their availability rather than their matrilineal relationships. Our results also supported the conclusion that multiple ewe subpopulations exist within the Peninsular Ranges, and that these subpopulations are the most basic demographic and genetic units.

Dermacentor hunteri (Acari: Ixodidae): seasonal variation in questing adults and on-host juvenile stages, and host associations and feeding behavior of larvae and nymphs.

Dermacentor hunteri Bishopp is the only completely desert adapted tick in the Nearctic realm, and chiefly parasitizes desert bighorn sheep (Ovis canadensis Shaw) as an adult. The remainder of its life history has been unknown. We conducted field investigations in the Sonoran desert of the temporal and spatial variation of adult host-seeking ticks and of the host associations of juvenile ticks. Additionally, the feeding success of juvenile ticks was assessed in the laboratory. Adult ticks were found in significant numbers only in plateau and rocky slopes habitats, chiefly during the period from January to June. Questing adults were not found in July and August, and they were present in small numbers from September through December. Juvenile stages were found only on desert woodrats, Neotoma lepida Thomas (larvae and nymphs), and cactus mice, Peromyscus eremicus Baird (larvae only), in March, May, and early June. In the laboratory; both larvae and nymphs fed on N. lepida, but only larvae fed on P. maniculatus bairdii (Wagner). We concluded that the life history of D. hunteri may be constrained by the co-distribution of desert bighorn, desert woodrats, and perhaps cactus mice; and that adults oversummer either on desert bighorn or sequestered in favorable microclimates off the host.

Diagnostic procedures and treatment of eleven dogs with peritoneal infections caused by Mesocestoides spp.

An 8-year-old spayed Schnauzer with a distended abdomen was examined because of straining to urinate and suspected urinary tract infection. Abdominal radiography revealed a ground-glass appearance, and ultrasonography revealed numerous cystic structures in the peritoneal cavity. Examination of an aspirate of abdominal fluid revealed tissues consistent with metacestodes. Tissues were definitively identified as Mesocestoides spp on the basis of polymerase chain reaction amplification of restriction fragment length polymorphisms. The dog required several courses of treatment with fenbendazole to eliminate the infection. This was 1 of 11 dogs infected with Mesocestoides metacestodes. Treatment involving the use of praziquantel and albendazole were ineffective, but fenbendazole successfully cleared Mesocestoides infections in 5 of 6 dogs.

Evaluation of bluetongue virus diagnostic tests in free-ranging bighorn sheep.

Five bluetongue virus (BTV) diagnostic tests were evaluated for use in free-ranging bighorn sheep. We sampled one bighorn sheep population four times between 1989 and 1995. The tests evaluated included virus isolation (VI), polymerase-chain reaction (PCR), serum neutralization (SN), agar-gel immunodiffusion (AGID), and competitive enzyme-linked immunosorbent assay (c-ELISA). The c-ELISA, AGID and SN tests had high levels of agreement in determining serogroup exposure in bighorn sheep. We used maximum-likelihood algorithms to estimate the parameters of each diagnostic test used. Although the c-ELISA and AGID had high sensitivity and specificity, the SN had perfect specificity but lower apparent sensitivity. Due to the potential of cross-reactions among multiple serotypes, results of the SN must be interpreted with caution when assessing serotype exposure in an area where multiple serotypes are endemic. The PCR assay delineated convalescent antibody titers from more-recent infections, and consequently, was pivotal in distinguishing a different exposure pattern between the bighorn sheep and cattle in an adjacent herd. Based on an increasing seroprevalence (50% to 100%), BTV circulated through this bighorn sheep population between 1989 and 1993. This increase in seroprevalence coincided with a bighorn die-off due to BTV infection in June, 1991. An adjacent cattle herd was sampled in 1995 for comparison. The bighorn sheep and adjacent cattle had different patterns of exposure to BTV between 1994 and 1995. There was no evidence that BTV circulated through the bighorn sheep population from 1994 to 1995. In 1995, seroprevalence to BTV decreased to 72%, none of yearling bighorn was seropositive, and all of the 39 bighorn sheep were PCR-negative. In contrast, all adult cattle were seropositive to BTV by c-ELISA and SN, and 4 of the calves were seropositive; 11 of the 24 cattle were PCR-positive, including all five calves. Overall, the pattern of temporal herd immunity in the bighorn sheep appeared to follow a classic epidemic curve, with the appearance and subsequent disappearance of herd immunity coinciding with the 1991 die-off in this population. As low levels of herd immunity and high proportions of susceptible animals are key factors in the development of epidemics, this population of bighorn sheep may be at increased risk for a BTV epidemic in the future.

DNA sequence variation in Dermacentor hunteri and estimated phylogenies of Dermacentor spp. (Acari: Ixodidae) in the New World.

The genus Dermacentor is represented by 12 species in the New World. We sequenced a 300-bp portion of the mitochondrial 16S ribosomal DNA gene for 28 individual ticks representing 9 of these species and analyzed their phylogenetic relationships. Maximum parsimony, distance (neighbor-joining), and maximum likelihood were all used to resolve tree topologies. Eleven specimens of Dermacentor hunteri Bishopp representing populations across the tick's entire geographic range showed negligible genetic variation, with only single base-pair differences between each of 5 haplotypes. We found high degrees of bootstrap support (66-86%) for monophyly of the genus, but variable support for monophyly of species within the genus. D. hunteri, D. occidentalis Marx, and D. variabilis (Say) each resolved as a monophyletic taxon (79-99% support). D. andersoni Stiles and D. parumapertus Neumann formed a paraphyletic clade (99% support). D. albipictus Packard showed substantial intraspecific variation and warrants further investigation. D. imitans Warburton was distinct from all other Dermacentor spp. on all trees.

Prevalence of and associated risk factors for shedding Cryptosporidium parvum oocysts and Giardia cysts within feral pig populations in California.

Populations of feral pigs (Sus scrofa) may serve as an environmental reservoir of Cryptosporidium parvum oocysts and Giardia sp. cysts for source water. We conducted a cross-sectional study to determine the prevalence of and associated demographic and environmental risk factors for the shedding of C. parvum oocysts and Giardia sp. cysts. Feral pigs were either live-trapped or dispatched from 10 populations located along the coastal mountains of western California, and fecal samples were obtained for immunofluorescence detection of C. parvum oocysts and Giardia sp. cysts. We found that 12 (5.4%) and 17 (7.6%) of 221 feral pigs were shedding C. parvum oocysts and Giardia sp. cysts, respectively. The pig's sex and body condition and the presence of cattle were not associated with the probability of the shedding of C. parvum oocysts. However, younger pigs (< or = 8 months) and pigs from high-density populations (> 2.0 feral pigs/km2) were significantly more likely to shed oocysts compared to older pigs (> 8 months) and pigs from low-density populations (< or = 1.9 feral pigs/km2). In contrast, none of these demographic and environmental variables were associated with the probability of the shedding of Giardia sp. cysts among feral pigs. These results suggest that given the propensity for feral pigs to focus their activity in riparian areas, feral pigs may serve as a source of protozoal contamination for surface water.

Immobilization and physiological parameters associated with chemical restraint of wild pigs with Telazol and xylazine hydrochloride.

We used a combination of Telazol (3.3 mg/kg) and xylazine hydrochloride (1.6 mg/kg to immobilize 144 wild pigs (Sus scrofa) with blow darts. This drug combination was safe and effective for rapidly immobilizing animals ranging in size from 34 to > 170 kg and avoided difficulties associated with hand injections. For 123 single injection immobilizations, mean (+/- SD) induction times and effective handling periods averaged 5 (+/- 2.5) and 52 (+/- 18) min, respectively, and animals generally recovered for release within 120 min of initial injections. Animals that required two injections to immobilize (n = 21) received lower initial doses of Telazol and xylazine hydrochloride than those immobilized with a single injection because of errors in estimating body sizes; we found that there was a threshold dose required to immobilize wild pigs from 2.8 to 3.3 mg/kg Telazol and 1.4 to 1.6 mg/kg xylazine. Although neither age or sex influenced immobilization parameters, animals in good condition required longer to recover than those in poor condition. However, animals immobilized with two injections recovered as rapidly as those immobilized with a single injection. Heart rates and body temperatures declined slightly during the immobilization period, but respiration rates and blood oxygen saturation levels remained stable. In general, single injection immobilizations were preferable because they minimized problems associated with injecting partially immobilized animals. because it was difficult to accurately estimate the sizes of large wild pigs (> or = 90 kg), and because wild pigs that were partially immobilized were difficult to handle, we recommend increasing the drug doses to 4 mg/kg Telazol and 2 mg/kg xylazine hydrochloride when injecting relatively large animals to assure single injection immobilizations. Although recovery periods may be prolonged, higher doses of Telazol and xylazine should be safe based on data from domestic pigs.