Abstract
The major goal of this study was to investigate whether hair could be collected by hair snares attached to scent posts from free-ranging Florida panthers (Puma concolor coryi) to determine species identity using DNA analyses of the hair samples. The five phases of the study are summarized below. For Phase 1, methods were developed and tested on captive animals with the intention of transitioning the effective methods to the field. In Component 1 of Phase 1, scent preferences of captive Western cougars (Puma concolor stanleyana) were established in the Naples Zoo (Naples, Florida) where the cougars' reactions to various scents could be monitored closely. Four of26 tested scents attracted captive cougars in an initial pilot study. In 30 experimental trials, Bag Balm and Beaver Lure with Catnip were identified as the two most attractive scents stimulating rubbing response rates of93% and 67% respectively. In Component 2 of Phase 1, hair snares effective for collecting hair for DNA analysis were developed using captive Western cougars as test subjects in the Naples Zoo where behavioral interactions to snares associated with scent stations could be observed closely. Three types of hair snares were tested; a circular wire cup brush, a three-ring curry comb and a rectangle wire brush. The circular wire cup, most effective at capturing and retaining cougar hairs, collected 600 hairs, 75 of them with follicles attached. For Phase 2, scent stations (i.e., posts equipped with scent attractants and circular wire cup snares) based on prototypes developed at the Naples Zoo, were field tested on free-ranging Florida panthers in the U.S. Fish and Wildlife Service's Florida Panther National Wildlife Refuge (FPNWR) (Collier County Florida). A total of 72 hairs were collected from 12 scent stations in two separate trials. Scent stations were monitored with trip-motion 35 mm cameras. A total of 10 photographs documented free-ranging Florida panthers in the vicinity of scent stations and only one photograph recorded a panther directly interacting with a scent station. For Phase 3, hairs collected were analyzed in the lab. In Component 1 of Phase 3, hairs collected in hair snares in the FPNWR were visually examined for species identification. Twelve of 72 hairs were identified as black bear (Ursus americanus) and the species identity of the remaining 60 hairs was ambiguous. In Component 2 of Phase 3, hair collected at both the Naples Zoo and the FPNWR were analyzed to determine whether species identity could be confirmed using DNA analyses. A 215bp fragment of the 12S ribosomal DNA (rDNA) was amplified in 47 of 137 (34%) captive Western cougar hairs and in 20 of 60 hairs collected in the FPNWR. Sequences were obtained from four of the amplification products from captive Western cougar hairs, and as expected, all were identified as Puma concolor. Sequence data were successfully obtained from 20 hairs collected in the FPNWR; 17 hairs were identified as Puma concolor, one hair was identified as Ursus americanus, one hair was identified as Homo sapiens likely due to researcher contamination, and species identity in one hair was inconclusive due to insufficient DNA for identification. The 17 hairs identified as Puma concolor were independent sampling events collected from seven different scent stations on nine different collection dates. Based on the results of this study, scent stations equipped with the appropriate scents and hair snares can be used to collect hair with sufficient DNA from free-ranging and highly secretive Florida panthers under field conditions to obtain species identifications using DNA analyses. These are the first steps towards successfully amplifying nuclear DNA (e.g., as microsatellite DNA) and genetic identification of individual free-ranging Florida panthers. DNA was successfully extracted and fragments up to 215bp were amplified from hairs collected under field conditions in Florida. Thus, with some slight modifications, this inexpensive method may be used as a valuable addition or even an alternative to the invasive methods that wildlife managers currently undertake to collect genetic information from free-ranging panthers in Florida as well as other felid species from around the world.