Vol. 26: 179–188, 2014 ENDANGERED SPECIES RESEARCH Published online December 10 doi: 10.3354/esr00626 Endang Species Res OPENPEN ACCESSCCESS Non-invasive sampling reveals fine-scale genetic structure in black bear Ursus americanus populations from northeastern Mexico Fernando Montiel-Reyes1, Jesús E. Maldonado2,3, Melina Del Real-Monroy1, Norberto Martínez-Méndez4, Jorge Ortega1,* 1Laboratorio de Bioconservación y Manejo, Posgrado en Ciencias Quimicobiológicas, Departamento de Zoología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Prolongación de Carpio y Plan de Ayala s/n, Col. Sto. Tomas, 11340, México, DF 2Center for Conservation and Evolutionary Genetics, Smithsonian Conservation Biology Institute, 3001 Connecticut Avenue NW, Washington, DC 20008, USA 3Department of Vertebrate Zoology, National Museum of Natural History, Smithsonian Institution, MRC 108, Washington, DC 20013, USA 4Laboratorio de Bioconservación y Manejo, Posgrado en Biociencias, Departamento de Zoología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Prolongación de Carpio y Plan de Ayala s/n, Col. Sto. Tomas, 11340, México, DF ABSTRACT: The black bear Ursus americanus is one of the largest terrestrial carnivores that has a wide distribution in northeastern Mexico, the only country in which black bears are listed as endangered. We used 10 nuclear microsatellite loci to evaluate black bear genetic variability in 6 disjointed populations from northeastern Mexico. Non-invasive genetic techniques were applied to fecal and hair samples. Using a panel of 10 polymorphic microsatellites we identified 64 indi- viduals. Black bears showed low to moderate levels of genetic diversity in all sampled populations (mean ± SD expected heterozygosity, He = 0.63 ± 0.628). Pairwise comparisons between all 6 pop- ulations (φST = 0.315, p < 0.05) detected significant genetic differentiation between the western Coahuila and the northeastern Nuevo León regions, suggesting that black bears have low levels of gene flow between these 2 regions. Microsatellites revealed significant structure within the complex of disjoint areas in the region. The inbreeding coefficient was also significant (φIS = 0.143). The largest proportion of genetic variation (82.7%) was found between individuals within populations. Distance and anthropogenic activities may serve to limit gene exchange among pop- ulations which form at least 3 distinct genetic clusters; these may respond differently to environ- mental changes and should be considered distinct management units. KEY WORDS: Black bears · Genetic structure · Microsatellite · Non-invasive sampling · Mexico INTRODUCTION vided indications of the possible effects of natural and artificial dispersal on subpopulations of a partic- Factors influencing the genetic structure of species ular species (Friesen et al. 2006, Small et al. 2006, are important for understanding how populations McDevitt et al. 2013). Other studies have shown the change over time and for predicting how such importance of habitat fragmentation and hetero- changes affect evolutionary processes. Several stud- geneity and the effects of genetic drift of subpopula- ies on the genetic structure of vertebrates have pro- tions as a consequence of restricted dispersal (Bergl © The authors 2014. Open Access under Creative Commons by *Corresponding author: [email protected] Attribution Licence. Use, distribution and reproduction are un - restricted. Authors and original publication must be credited. Publisher: Inter-Research · www.int-res.com 180 Endang Species Res 26: 179–188, 2014 & Vigilant 2007, Heller et al. 2010). A number of differentiation and higher levels of gene flow with analyses have shown low to medium values of populations of western Texas (USA) than with popu- genetic diversity among populations of large mam- lations from the southeastern highlands of Coahuila mals resulting from low densities throughout the spe- and Nuevo León. We predicted that black bears from cies’ respective ranges and despite the high dispersal western Coahuila are more likely to disperse long rates of some individuals (Lawson Handley & Perrin distances along high-elevation woodland habitats in 2007, Archie et al. 2008). the same mountain range than to cross shorter geo- Black bears Ursus americanus are present in all graphic distances through inhospitable desert habi- Canadian provinces, in almost all of the USA except tat between the eastern and western Coahuila high- for the arid areas of the southwest, and also in the lands. We also evaluated the levels of genetic Sierra Madre Occidental and Sierra Madre Oriental diversity and subdivision between highly impacted of Mexico (Larivière 2001). In Mexico, black bears populations in the western highlands of Coahuila primarily inhabit sparsely populated forest areas and Nuevo León compared to the less impacted pop- above 2000 m in the northeast and northwest ranges ulations of northeastern Coahuila. of both Sierras Madre. Mexico is the only country where black bears are listed as endangered (Cebal- los et al. 2002) because they have been severely MATERIALS AND METHODS affected by adverse factors such as habitat loss due to anthropomorphic activities, climate change, poach- Study area ing, and illegal trade (Sierra Corona et al. 2005). Black bear populations inhabiting these high-eleva- We sampled an area that encompassed most of the tion woodlands are often separated by the more known current distributional range of the black bear inhospitable desert and grassland habitats in the Ursus americanus in northeastern Mexico (Fig. 1). lowlands (Varas-Nelson 2010). However, as the The 6 sampling sites were located in the southeastern available habitat on mountain tops has become highlands of the states of Coahuila and Nuevo León, smaller and more fragmented, black bears are occa- and also in the northeastern part of Coahuila (Sierra sionally forced to attempt to move through the desert del Burro and Sierra del Carmen). Overall, the habitat lowland to reach woodlands in neighboring moun- in the desert areas is characterized by abundant xero- tains areas (LeCount & Yarchin 1990). These lowland areas have also been highly impacted by anthro- pogenic disturbances and barriers due to industrial and agricultural development, which has affected the ability of individuals to disperse to nearby moun- tains (Varas-Nelson 2010). The current status of the black bear population in the southeastern highlands of Coahuila and Nuevo León, Mexico, is poorly understood due to the scarcity of records in scientific collections and the lack of reliable data from obser- vations of individual animals. On the other hand, the black bear populations in western Coahuila appear to be healthy and stable as a result of conservation management in the protected areas and well-pre- served suitable habitats (Doan-Crider & Hellgren 1996). Here we used non-invasive sampling coupled with molecular neutral markers to assess the genetic diversity and genetic structure of black bear popula- tions in northeastern Mexico. Our particular objec- tives were to assess the levels of genetic diversity of Fig. 1. Historical distribution of Ursus americanus in North black bear populations and compare our results with America. Inset shows sampled sites in northeastern Mexico for black bear populations in the states of Coahuila and nearby populations in the southeastern USA. We Nuevo León: 1, Real de Minas Viejas/Villaldama; 2, Sierra tested the hypothesis that black bear populations de Picachos; 3, Linares; 4, Santa Catarina; 5, Sierra del from western Coahuila have lower levels of genetic Burro; 6, Sierra del Carmen Montiel-Reyes et al.: Fine genetic structure of Mexican black bears 181 phytic vegetation and rocky outcrops in the arid re- levels of genetic diversity and structure. Between gions, while in the mountains, 6 different associations 2008 and 2012, we surveyed transects throughout the of pine and oak woods are recognized (Jiménez- 6 different locations, walking a maximum distance of Guzmán & Zúñiga-Ramos 1991). The highland areas 10 km, searching for fecal material and placing 10 are mainly comprised of oaks Quercus hypoleucoides baited barbed wire hair traps at intervals of 1 km and Q. muhlenbergii, pines Pinus strobiformis and P. (Woods et al. 1999). Transects were surveyed during arizonica, and cedar Juniperus deppeana and are dry field seasons (February to May) in the early characterized by steep peaks with deep canyons, with morning to avoid negative effects of the weather on an average elevational gradient of 1750 m. The arid the feces (Hájková et al. 2006). In the dry season of land ranges in altitude from 1000 to 1500 m, and has each of the 5 years of the study period we checked for the floristic characteristics of the Chihuahuan Desert, scats and set hair snags. Fresh fecal samples were i.e. it mainly supports Larrea tridentata, Agave placed in Falcon tubes with silica gel and transported lechuguilla, and Parthenium incanum (Muldavin to the laboratory for analysis. Black bears are the 2002). These areas are considered unsuitable habitat only large carnivore present in the area and there- for black bear populations (Onorato et al. 2007). fore we did not need to identify fecal samples to spe- The northeastern part of Coahuila consists princi- cies using a genetic marker because visual identifica- pally of an elevated limestone plateau that is an tion and size of feces were conclusive for species extension of the northern Chisos Mountains