University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln USGS Staff -- Published Research US Geological Survey 2006 Phylogeography and Spatial Genetic Structure of the Southern Torrent Salamander: Implications for Conservation and Management Mark P. Miller Utah State University Susan M. Haig U.S. Geological Survey, [email protected] R. Steven Wagner Central Washington University, [email protected] Follow this and additional works at: https://digitalcommons.unl.edu/usgsstaffpub Miller, Mark P.; Haig, Susan M.; and Wagner, R. Steven, "Phylogeography and Spatial Genetic Structure of the Southern Torrent Salamander: Implications for Conservation and Management" (2006). USGS Staff -- Published Research. 681. https://digitalcommons.unl.edu/usgsstaffpub/681 This Article is brought to you for free and open access by the US Geological Survey at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in USGS Staff -- Published Research by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. Journal of Heredity 2006:97(6):561–570 ª The American Genetic Association. 2006. All rights reserved. doi:10.1093/jhered/esl038 For permissions, please email: [email protected]. Advance Access publication November 29, 2006 Phylogeography and Spatial Genetic Structure of the Southern Torrent Salamander: Implications for Conservation and Management MARK P. MILLER,SUSAN M. HAIG, AND R. STEVEN WAGNER Downloaded from From the Department of Biology, 5305 Old Main Hill, Utah State University, Logan, UT 84322-5305 (Miller); United States Geological Survey Forest and Rangeland Ecosystem Science Center, 3200 Southwest Jefferson Way, Corvallis, OR 97331 (Haig and Wagner); and Genetics Program, Oregon State University, Corvallis, OR 97331 (Wagner). R. Steven Wagner is now at the Department of Biological Science, Central Washington University, Ellensburg, WA 98926. http://jhered.oxfordjournals.org/ Address correspondence to R. S. Wagner at the address above, or e-mail: [email protected]. Abstract The Southern torrent salamander (Rhyacotriton variegatus) was recently found not warranted for listing under the US Endan- gered Species Act due to lack of information regarding population fragmentation and gene flow. Found in small-order streams associated with late-successional coniferous forests of the US Pacific Northwest, threats to their persistence include distur- at University of Nebraka-Lincoln Libraries on January 10, 2013 bance related to timber harvest activities. We conducted a study of genetic diversity throughout this species’ range to 1) iden- tify major phylogenetic lineages and phylogeographic barriers and 2) elucidate regional patterns of population genetic and spatial phylogeographic structure. Cytochrome b sequence variation was examined for 189 individuals from 72 localities. We identified 3 major lineages corresponding to nonoverlapping geographic regions: a northern California clade, a central Oregon clade, and a northern Oregon clade. The Yaquina River may be a phylogeographic barrier between the northern Oregon and central Oregon clades, whereas the Smith River in northern California appears to correspond to the discontinuity between the central Oregon and northern California clades. Spatial analyses of genetic variation within regions encompassing major clades indicated that the extent of genetic structure is comparable among regions. We discuss our results in the context of con- servation efforts for Southern torrent salamanders. Genetic analyses of natural populations can become more and population demography (e.g., Jarne 1995; Miller et al. than theoretical investigations of biological, geographic, 2002; Mahoney 2004). In the case of endangered taxa, gen- and historical processes when threatened or endangered spe- eration of such information can assist with understanding the cies are involved as they provide unique opportunities to apply basic biology and life history of species that facilitates formu- the wealth of knowledge accumulated over the last century for lation of species conservation strategies. the purposes of addressing contemporary conservation issues. The Southern torrent salamander (Rhyacotriton variegatus) For example, pure phylogeographic analyses of molecular was recently found not warranted for listing under the US genetic variation can provide valuable insights about specific Endangered Species Act after the US Fish and Wildlife Service factors that contribute to patterns of genetic diversity and concluded there was a ‘‘lack of information [about whether] divergence across a species’ range (Avise 2000). Such explo- the species is threatened by low gene flow and low genetic di- rations may identify geographic features that produce deep versity across its range’’ (Federal Register 60: 33785). This spe- patterns of genetic divergence and may indicate timing of his- cies is widely but patchily distributed throughout the Pacific torically important events (e.g., Haig et al. 2004; Ripplinger Coast mountain range of the US Pacific Northwest, extending and Wagner 2004; Miller et al. 2005). From a conservation from Tillamook County, OR, south to Mendicino County, CA. perspective, these analyses can prove invaluable for defining Although they are limited primarily to the Pacific Coast moun- conservation units for species management purposes (Moritz tain range, their distribution also extends eastward into the 1994a, 1994b). Likewise, genetic analyses can also provide Central Cascade Range of Oregon (Figure 1; Leonard et al. information about dispersal abilities, reproductive strategies, 1993; Wagner et al. 2006). Mostly found in small streams 561 Journal of Heredity 2006:97(6) atures (Brattstrom 1963; Nussbaum and Tait 1977; Nussbaum et al. 1983). Subsequently, removal of the forest canopy may increase mean stream temperatures and stream sedimentation, leadingtoextirpationoflocalpopulations(BuryandCorn1988b; Corn and Bury 1989; Welsh 1990; Welsh and Lind 1992). As a consequence, recolonization after extirpation is thought to be low, due to these ecological factors and their apparent limited dispersal abilities (Nussbaum and Tait 1977; Nijhuis and Kaplan 1998). In this paper, we examine patterns of phylogeographic and population genetic structure in the Southern torrent sal- amander cytochrome b gene at both regional and local spatial scales. We compare our results to previously conducted allo- zyme studies (Good et al. 1987; Good and Wake 1992) and Downloaded from discuss our results with respect to conservation issues related to the Southern torrent salamander. Methods http://jhered.oxfordjournals.org/ DNA Isolation and Sequencing Southern torrent salamanders were sampled from 72 localities (1–3 individuals per location, 189 in total) throughout their known range (Figure 1 and Table 1). Also included in sequence analyses were limited numbers of Cascade torrent, Columbia torrent, and Olympic torrent salamanders (Rhyacotriton cascadae, Rhyacotriton kezeri,andRhyacotriton olympicus, respectively) as out- groups (Table 1 and Figure 1). Sample tissue was taken by non- at University of Nebraka-Lincoln Libraries on January 10, 2013 lethal tail clipping (;1 cm) from hand-captured adults and was stored immediately in a cryogenic tube containing buffer so- lution (100 mM Tris–HC1 pH 8.0, 100 mM ethylenediamine- tetraacetic acid pH 8.0, 10 mM NaCl, 0.5% sodium dodecyl sulfate) until transferred to an ultracold freezer (À80 °C). A modified phenol–chloroform extraction procedure was used to isolate DNA (Sambrook et al. 1989) with the final extracted aqueous layer purified in a microcon-50 filter (Millipore, Billerica, MA). An ;850-bp fragment of the cy- tochrome b gene was amplified for each of the 189 individ- uals in the data set using the following primers designed for vertebrates: MVZ15 (5#-GAACTAATGGCCCACACWW- TACGNAA-3#) and MVZ16 (5#-AAATAGGAAATATCA- TTCTGGTTTAAT-3#) (Kocher et al. 1989). Each polymerase chain reaction was carried out with 100 ng DNA in a 50-ll volume using 0.5 units of Taq Gold (Perkin Elmer, Wellesley, MA), 100 lM each deoxynucleoside triphosphate, 2 mM MgCl, Figure 1. Map of collection locations for Southern torrent and 1 lM of each primer. Thermal cycling was performed using salamanders and out-groups in the western United States the following parameters: an initial denaturation of 10 min at of America. Variation in symbols for Southern torrent 93 °C, followed by 40 cycles of denaturation for 1 min at 93 °C, salamanders indicates the 3 major phylogenetic lineages annealing for 1 min at 52 °C, and extension at 72 °C for 2 min. identified from cytb sequence analyses (Figures 2 and 3). A final extension at 72 °C for 10 min completed the reaction. Reaction products were run on 1% agarose gels, and amplified and headwaters associated with late-successional forests, they cytochrome b fragments were extracted from gel slices using are impacted by timber harvest and related disturbance activ- an ultra-free-mc 0.45 filter (Millipore). The supernatant was ities (Bury and Corn 1988a; Corn and Bury 1989; Diller and then transferred to a microcon-50 filter (Millipore) and washed Wallace1996).Juvenilesarerestrictedtocold,clear,fast-flowing twice with 400 ll distilled deionized water. Sequencing was streams, and adults are rarely found more than a few meters performed on an Applied Biosystems (373A) sequencer. Se- from stream banks. Both adults and juveniles appear sensitive quencing primers included MVZ15, MVZ16, and cytb2(5#-