Walter M. Boyce

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.

Genetic structure of mountain lion (Puma concolor) populations in California

Analysis of 12 microsatellite loci from431 mountain lions (Puma concolor)revealed distinct genetic subdivision that wasassociated with geographic barriers andisolation by distance in California. Levels ofgenetic variation differed among geographicregions, and mountain lions that inhabitedcoastal areas exhibited less heterozygositythan those sampled inland. The San FranciscoBay and Sacramento-San Joaquin River Delta, theCentral Valley, and the Los Angeles Basinappeared to be substantial barriers to geneflow, and allele frequencies of populationsseparated by those features differedsubstantially. A partial barrier to gene flowappeared to exist along the crest of the SierraNevada. Estimated gene flow was high amongmountain lions inhabiting the Modoc Plateau,the western Sierra Nevada, and northern sectionof the eastern Sierra Nevada. SouthernCalifornia mountain lion populations mayfunction as a metapopulation; however, humandevelopments threaten to eliminate habitat andmovement corridors. While north-south geneflow along the western Sierra Nevada wasestimated to be very high, projected loss andfragmentation of foothill habitat may reducegene flow and subdivide populations. Preservation of existing movement corridorsamong regions could prevent population declinesand loss of genetic variation. This studyshows that mountain lion management andconservation efforts should be individualizedaccording to region and incorporatelandscape-level considerations to protecthabitat connectivity.

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.

Diffuse Unilateral Subacute Neuroretinitis: Morphometric, Serologic, and Epidemiologic Support for Baylisascaris as a Causative Agent

PURPOSE Several nematodes have been postulated as etiologic agents in diffuse unilateral subacute neuroretinitis (DUSN), but the cause of this condition remains uncertain. The authors report the first case of DUSN from the western United States (northern California), along with morphometric, serologic, and epidemiologic evidence supporting Baylisascaris procyonis as its cause. METHODS One patient was examined and evaluated for disc edema and transient obscurations of vision. A diagnosis of DUSN was confirmed when a motile nematode was identified within the substance of the patients retina. Morphometric analysis of the nematode was done from projected fundus photographs. Serologic evidence of Baylisascaris infection was suggested by Western blot analysis. A necropsy was done on 12 raccoons from the area. They were examined for evidence of Baylisascaris infection, previously believed to be nonendemic in the region. RESULTS The intraocular nematode measured 1727 x 67 microns, most consistent with Baylisascaris. The patient had considerable exposure to raccoons, and was seropositive for B. procyonis infection on Western blot analysis. Necropsy evaluation showed B. procyonis infection in 8 of 12 raccoons examined from the area. CONCLUSIONS The morphometric, serologic, and epidemiologic findings in this case provide evidence that the raccoon ascarid, B. procyonis, is a cause of the large nematode variant of DUSN.

Three Pathogens in Sympatric Populations of Pumas, Bobcats, and Domestic Cats: Implications for Infectious Disease Transmission

Anthropogenic landscape change can lead to increased opportunities for pathogen transmission between domestic and non-domestic animals. Pumas, bobcats, and domestic cats are sympatric in many areas of North America and share many of the same pathogens, some of which are zoonotic. We analyzed bobcat, puma, and feral domestic cat samples collected from targeted geographic areas. We examined exposure to three pathogens that are taxonomically diverse (bacterial, protozoal, viral), that incorporate multiple transmission strategies (vector-borne, environmental exposure/ingestion, and direct contact), and that vary in species-specificity. Bartonella spp., Feline Immunodeficiency Virus (FIV), and Toxoplasma gondii IgG were detected in all three species with mean respective prevalence as follows: puma 16%, 41% and 75%; bobcat 31%, 22% and 43%; domestic cat 45%, 10% and 1%. Bartonella spp. were highly prevalent among domestic cats in Southern California compared to other cohort groups. Feline Immunodeficiency Virus exposure was primarily associated with species and age, and was not influenced by geographic location. Pumas were more likely to be infected with FIV than bobcats, with domestic cats having the lowest infection rate. Toxoplasma gondii seroprevalence was high in both pumas and bobcats across all sites; in contrast, few domestic cats were seropositive, despite the fact that feral, free ranging domestic cats were targeted in this study. Interestingly, a directly transmitted species-specific disease (FIV) was not associated with geographic location, while exposure to indirectly transmitted diseases – vector-borne for Bartonella spp. and ingestion of oocysts via infected prey or environmental exposure for T. gondii – varied significantly by site. Pathogens transmitted by direct contact may be more dependent upon individual behaviors and intra-specific encounters. Future studies will integrate host density, as well as landscape features, to better understand the mechanisms driving disease exposure and to predict zones of cross-species pathogen transmission among wild and domestic felids.

Range expansion, population sizes, and management of wild pigs in California

The introduction and spread of nonnative organisms to new regious can disrupt ecosystems by causing declines or extinctions of native species. Widely mtroduced wild pigs (Sus scrofa ) have contributed to declines and extinctions of numerons species on oceanic islands and can have pronounced negative ecological effects on mainland areas when population densities are high. Although range expansion by introduced wild pigs has ceased in many regions of the United States, it has increased significantly since the 1950s in California. Our analyses of data from annual hunter survevs and mapped locations of hunter-killed wild pigs shows that the distribution of wild pigs increased from around 10 coastal connties in the early 1960s to parts of 49 of Californias 58 counties by 1996. An index to density based on locations of hunter-killed wild pigs plotted in a Geographic Information System (GIS) indicated that within the 79,550 km 2 (25%) of the total land area of the state now occupied by wild pigs populations are most abundant in the central and north-coast regions. By stratifying each county into 1 of 3 relative abundance classes and assigning density values based on research at multiple sites, we estimated tbere were around 133.106 (range = 106,485-159,727) wild pigs in California in 1996. The recent increase in the range of wild pigs in California was facilitated by a combination of multiple hunting-related introductions, deliberate releases of domestic pigs, and, possibly, increased forage availability associated with agricultural development. Natural range expansion also has occured, however, and the dynamics of the spread of wild pigs in California appear typical of invasions observed among other organisms. Forage and water availability are important factors influencing the dist ribution and abundance of wild pigs in California. and predation may be important to an unknown extent. Up to 40% of Californias wild pigs are removed from the population annually hich may control populations in some areas but not others. Wild pigs may be cansing ecological damage in some coastal regions where population densities are very high, and detailed studies are needed in those areas to help mitigate potential problems with this increasingly widespread mammal in California.

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 ≤ 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.

Genetic variation and population structure in desert bighorn sheep: implications for conservation

Bighorn sheep populations experienced a drastic reduction in both distribution and abundance until the advent of modern wildlife management, where improving viability of extant populations and translocating animals into historical habitat range have been the most important management policies. The fact that subspecies relationships among bighorn are ambiguous,together with the importance of selecting appropriate source stock and the expense of translocation projects, makes an understanding of subspecies relationships and genetic variation, within and between populations, important for the management and conservation of this species. In this study, genetic variation in 279 bighorn sheep from 13 study sites in Arizona, California, New Mexico and Alberta, Canada were examined by analyzing ten microsatellite loci to determine interpopulation differentiation and relationships between closely related taxa. All populations contained a substantial amount of genetic variation. Genetic differences between populations were large and roughly proportional to geographic distance. The significance of this to desert subspecies relationships and management is discussed.

Estimating sizes of wild pig populations in the north and central coast regions of California

We developed a mark-sighting approach with automatic camera systems and assessed the size and density of wild pig (Sus scrofa) populations in the North and Central Coast regions of California in 1994 and 1995. Eighty and 149 wild pigs were captured and tagged at 4 and 6 study sites in 1994 and 1995, respectively. Of those animals, 202 (88%) were subsequently sighted among over 3,000 photographs of wild pigs obtained from 54 camera stations. Sighting rates for both tagged and nontagged wild pigs were similar between years and averaged 2.8 ± 0.5 and 2.8 ± 0.4 wild pigs/camera-station night, respectively. Sighting rates for all wild pigs during the study averaged 5.7 ± 0.8 wild pigs/camera-station night. Mark-sighting data analyzed with program NOREMARK provided estimates of population size, and narrow 95% confidence intervals (95% CI) for most research sites suggested we obtained sufficient sightings for reliable estimates of size of populations. It also proved possible to identify nontagged wild pigs in photographs for determining minimum population sizes, and the 95% CI from mark-sighting estimates encompassed the minimum population sizes for 8 of 10 study sites. An important factor influencing the reliability of population estimates was the proportion of tagged animals resighted. At one site where few tagged animals were sighted, 95% CIs were large and did not encompass the minimum population estimate. Mean population densities ranged from 0.7 to 3.8 wild pigs/ km 2 , comparable to previous research on wild pigs in the regions we studied. Densities increased from 1994 to 1995, potentially related to higher rainfall and increased forage availability in 1995. Also, densities were lower in areas with relatively high hunting pressure, suggesting that sport hunting may be effective at reducing wild pigs numbers in some areas.

Avian influenza viruses in wild birds: A moving target

The long-standing evolutionary and ecological relationships between wild birds and influenza A viruses has created a broad pool of viral genetic diversity and a reservoir of potentially transmissible viruses. An understanding of these relationships can help us identify and modify critical control points to reduce transmission of avian influenza viruses into animal and human populations.