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An Isotopic Technique to Marie:Mid-Sized Vertebrates Non-Lnvaslvelv Journal of Zoology. Print ISSN 0952-8369 An isotopic technique to marie:mid-sized vertebrates non-lnvaslvelv 1,2 ,2 1,2 3 J. N. Pauli , M. Ben-David1 , S. W. Buskirk , J. E. DePue2 & W. P. Smith 1 Prograrn in Ecology, University of Wyorning, Laramie, WY, USA 2 Department of Zoology and Physiology, University of Wyoming, Laramie, WY, USA 3 USDA Forest Service, Pacific Northwest Research Station, Forestry and Range Sciences Laboratory, La Grande, OR, USA Keywords Abstract carbon; deuterium; nitrogen; mesocarnivore; movement; ranging. Although dispersal is an important attribute of animal population ecology, knowledge of dispersal rates or distances for many wide-ranging species is lacking. Correspondence Current methods require capturing and restraining animals, which can be cost- Jonathan N. Pauli, Department of Zoology prohibitive, fail to collect sufficient samples or change animal ranging behaviors. and Physiology, University of Wyoming, Herein, we describe a novel, cost-effective and non-invasive method, using bait 1000 E. University Avenue, Laramie, enriched with stable isotopes to mark the hair of American martens Martes WY82071, USA. americana. Captive martens that consumed isotopically labeled glycine exhibited Email: [email protected] significant and progressive enrichment in the isotopic signature of 13C,lsN and 2H in both whole blood and hair. A distinct mark in hair, >2 standard deviations Editor: Nigel Bennett above natural abundance, occurred within 14 days of the second dose. The rate of isotopic labeling of hair was higher in spring, possibly because labeled amino acids Received 7 November 2008; revised 31 became diluted among the many hairs growing during the autumn. Because hair January 2009; accepted 3 February 2009 and feathers can be collected non-invasively from large geographic areas without capturing animals, this labeling method can be used to mark and study the doi:10.1111/j.1469-7998.2009.00562.x movement and dispersal rates of animals across landscapes efficiently. Introduction bats Lasiurus cinereus can disperse > 2000: km. Similarly, using isotopic signatures of feathers, Hobson & Wassenaar Dispersal is a fundamental aspect of animal life history and (1997) determined the breeding origin of wintering Neotro- ecology; knowledge of it is necessary to develop effective pical migrants. These studies have largely relied on natural conservation strategies (Macdonald & Johnson, 2001). abundances of deuterium 2H) across latitudinal gradients, However, quantifying dispersal rates and distances for which does not exhibit sufficient variation to determine the wide-ranging species is challenging. Traditionally, methods dispersal of animals moving short or intermediate distances such as mark-recapture, radio-telemetry and, more recently, « 100 km; Wunder & Norris, 2008). Previous researchers DNA genotyping have been used to assess movements over have, however, used organic and inorganic compounds, long distances (Nathan et al., 2003). Although generally artificially enriched with stable isotopes, to track the fates accurate, these approaches require large investments of of substances in various ecosystem (Hall & Tank, 2003) and time, money and personnel, limiting the number of study physiological (Boutton, 1991; Hirons, Schell & St. Aubin, animals and duration that each can be monitored. Such 2001) processes. Animal ecologists recently have applied constraints reduce the likelihood of detecting rare, but these enriched isotopes as organismal markers: by dripping important dispersal events (Nathan et al., 2003). These enriched nitrogen 15N) into streams and ponds, Caudill methods also require capture, restraint and often anestheti- (2003) and Macneale, Peckarsky & Likens (2004) marked zation of animals, which can be stressful (Harper & Austad, and subsequently recaptured developing invertebrate larvae 2000) and alter subsequent behaviors (e.g. Clinchy, Krebs & to estimate dispersal distances; similarly, Wanner et at. Jarman, 2001). Therefore, for wide-ranging vertebrates, (2006) used 44Ca to mark and track the dispersal and particularly endangered ones, reliable estimates of dispersal foraging activity of the parasitoid wasp Cotesia glomerata. are lacking, impeding our ability to parameterize popula- Vertebrate ecologists are increasingly using tissues col- tion-based models for conservation (Macdonald & Johnson, lected non-invasively and DNA-based analyses to estimate 2001). dispersal and gene flow (DeYoung & Honeycutt, 2005). Naturally occurring stable isotopes have been used suc- However, this approach can be limited by degradation of cessfully to document long-distance movements of volant samples and related genotyping errors (Mills, 2007), which animals (Rubenstein & Hobson, 2004). For example, Cryan can increase bias of population estimates (Lukacs & Burn- et al. (2004) used stable isotopes in hair to show that hoary ham, 2005). Such analytical errors could be avoided with the Journal of Zoology 278 (2009) 141-148 © 2009 The Authors. Journal compilation © 2009 The Zoological Society of London 141 A non-invasive mark for mid-sized vertebrates J. N. Pauli et al. use of an independent organismal marker. We reasoned Biology Laboratory at the University of Wyoming. Here, that isotopically enriched compounds could also be used captive martens were exposed to ambient changes in photo- to mark free-ranging vertebrates. With artificially enriched period and temperature, minimizing interference with their baits and non-invasive sampling methods, researchers could moult cycle and fur growth. They were fed ad libitum mark animals and subsequently collect hair, feather or other (ExclusiveT cat food, PMI Nutrition's Henderson, CO, tissues to track the movements of many individuals. For USA) and had continuous access to water. Pens were mammals and birds, in which most amino acids incorpo- furnished with PVC tubes, nest boxes, branches and small rated into hair and feathers are derived from recently trees as environmental enrichment. Martens were accli- consumed food (Ayliffe et al., 2004), isotopic enrichment of mated to captivity and the cat food diet for 5 weeks. In the ingested amino acids could rapidly mark keratinous tissues. second week of May (coinciding with the moult; Soutiere & No previous research has tested whether this approach can Steventon, 1981), we administered a unique combination l3 15 2 produce an unambiguous mark in tissues that can later be of enriched (98-99 at. %) isotopes ( C, N and H; ISO- collected non-invasively. TEC™, Miamisburg, OH, USA) to the food of each animal l3 2 15 As part of a project assessing the effects of fragmentation (n = 2 for C and H, and n = 3 for N); animals that did l3 15 2 on the forest-associated mesocarnivore, the American mar- not receive an enriched diet for C, N or H were used as 13 2 ten Martes americana, we required a cost-effective and non- controls for that isotope (n = 3 for Cand H, and n = 2 for 15 invasive method to investigate movement of individuals N). For example, an animal that received a diet enriched in 2 13 15 between habitat patches. By providing bait enriched with H served as a control for C and N. Isotopic labels were different isotopic labels to martens inhabiting isolated forest provided in the form of glycine (Boutton, 1991), an amino fragments and subsequently collecting hair samples non- acid that is incorporated into both blood and hair (Hirons invasively from a large area over multiple sampling periods, et al., 2001). We estimated the isotopic dose required to we intended to estimate dispersal rates and distances. Before produce an isotopic label in the hair and blood of martens, using isotopically labeled baits in the field, we conducted assuming no elemental routing to different tissue types (i.e, feeding trials on captive martens to ascertain that consump- complete mixing; Table 1). Specifically, we calculated the ΕN tion of an isotopically enriched diet would produce a reliable mass of an isotope ( Ag)needed to elicit a significant mark in the hair. Our specific objectives were to: (1) measure increase in the signature of tissue with the equation: the dose-specific assimilation of isotopically labeled amino acids into marten blood and hair; (2) determine minimum ENA = [ENAxNA x A ]- [A xNA] (1) dosages needed to label wild martens; (3) identify the season g g g in which isotopically enriched bait would provide the most reliable mark. where eN A is the isotopic enrichment desired (the proportion of labeled isotope above natural abundance). For example, our desired enrichment was 1.05 times natural abundance Materials and methods for 15N, 1.04 times for l3C and, because roughly 20% of During the last week of March 2006, we captured five hydrogen within hair exchanges with the atmosphere 2 N martens along snowmobile routes and logging roads in the (Cryan et al., 2004), 1.50 times for H. A is the natural Snowy Range Mountains of south-eastern Wyoming abundance of the heavy isotope, and Ag is the mass of the (4104'N 106°9'W). We anesthetized captured martens with element in the species (Table 1). To determine the mass ketamine hydrochloride (l5.0mgkg-1 body weight) and of enriched glycine to feed martens, we multiplied the xylazine hydrochloride (1.6 mg kg-1 body weight; Ben- grams of enriched element to achieve the necessary mark David, Schell & Flynn, 1997) and transported them to by the molecular mass (Mr) and divided by the atomic individual outdoor pens at Red Buttes Environmental mass (ma) of that isotope: Marten body mass was
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