Miocene extensional tectonics explain ancient ... - Wiley Online Library

Journal of Biogeography (J. Biogeogr.) (2015) 42, 1052–1065

ORIGINAL ARTICLE

Miocene extensional tectonics explain ancient patterns of diversification among turret-building tarantulas (Aphonopelma mojave group) in the Mojave and Sonoran deserts Matthew R. Graham1*, Brent E. Hendrixson2, Chris A. Hamilton3 and Jason E. Bond3

1

Department of Biology, Eastern Connecticut State University, Willimantic, CT 06226, USA, 2Department of Biology, Millsaps College, Jackson, MS 39210, USA, 3 Department of Biological Sciences and Auburn University Museum of Natural History, Auburn University, Auburn, AL 36849, USA

ABSTRACT

Aim Phylogeographical studies in the Mojave and Sonoran deserts often find genetic discontinuities that pre-date the Pleistocene. A recent synthesis of phylogeographical data, called the Mojave Assembly Model, provides a hypothesis for the historical assembly of these desert biotas but does not adequately capture the complexity of pre-Pleistocene vicariance events. We tested this model and assessed pre-Pleistocene divergences by exploring the phylogeography of the Aphonopelma mojave group, which is composed of turret-building tarantula species from the Mojave and Sonoran deserts. Location Mojave and Sonoran deserts, south-western USA. Methods We augmented the sampling from a previous study by sequencing mitochondrial DNA (COI) from new material of the A. mojave group. We used phylogenetic and network analyses to identify clades and a molecular clock and lineages-through-time plots (LTT plots) to explore the timing and tempo of diversification. We tested for demographic expansion using neutrality tests and mismatch distributions. Species distribution models (SDMs) were constructed to compare current suitable habitat to that at the Last Glacial Maximum (LGM). Results Phylogenetic, network and molecular-clock analyses identified six major clades that probably diverged during the late Miocene. The rate of diversification appears to have slowed during the Pliocene. Most clades exhibit signals of recent demographic expansion. SDMs predicted that suitable habitat shifted south and to lower elevations during the LGM.

*Correspondence: Matthew R. Graham, Department of Biology, Eastern Connecticut State University, 83 Windham Street, Willimantic, CT 06226, USA. E-mail: [email protected]

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Main conclusions Phylogeographical analyses suggest that the A. mojave group experienced a burst of diversification in the late Miocene, followed by population expansions during the Pleistocene. Six major clades with origins in the late Miocene cannot be adequately explained by the Mojave Assembly Model. We propose the novel hypothesis that Miocene extensional tectonics caused populations to diverge in allopatry by producing low-elevation habitat barriers. Geological models, such as kinematic reconstructions, provide an ideal but underutilized framework for testing biogeographical hypotheses in these deserts and the wider Basin and Range Province. Keywords Aphonopelma joshua, BEAST, biogeography, COI, divergence dating, MAXENT, Mygalomorphae, south-western USA.

http://wileyonlinelibrary.com/journal/jbi doi:10.1111/jbi.12494

ª 2015 John Wiley & Sons Ltd

Turret-building tarantula diversification INTRODUCTION The Mojave and Sonoran deserts have undergone a vibrant biogeographical history shaped by Neogene tectonics and Quaternary climate fluctuations (Axelrod, 1983; Riddle et al., 2008). In response to the changing landscapes and climates, the distributions of arid-adapted organisms that inhabit these regions are thought to have repeatedly fragmented, in turn elevating rates of diversification and endemism. As a result, these deserts now harbour rich biotas and provide ideal systems for studying the evolution of desert ecosystems and their responses to changing climates and landscape deformations. Phylogeographical studies of Mojave and Sonoran desert taxa have undergone a recent surge. Prior to this, broadscale phylogeographical studies of desert organisms commonly discovered ancient pre-Pleistocene divergences, particularly across the Sonoran–Chihuahuan boundary and along the Baja California Peninsula (reviewed in Hafner & Riddle, 2011). In a meta-analysis of phylogeographical studies, Wood et al. (2012) found that pre-Pleistocene divergences also exist across the Mojave and Sonoran deserts, and documented them in seven of the 12 taxa studied. In addition, pre-Pleistocene divergences were discovered in two of four new phylogeographical studies published since the meta-analysis (Table 1). No ecological or biogeographical explanations have yet been proposed to account for why some arid-adapted organisms but not others exhibit this pattern. Phylogeographical analyses of additional codistributed species and species groups may indicate that shared biological or geological explanations for these genetic patterns of divergence exist.

A major development in the biogeography of the Mojave and Sonoran deserts was the synthesis of available phylogeographical data into a model of biotic assembly called the ‘Mojave Assembly Model’. First proposed by Bell et al. (2010) and subsequently refined by Graham et al. (2013a), this model outlines a working hypothesis for the historical assembly of the Mojave Desert biota, including that of the north-western region of the Sonoran Desert. Briefly, the Mojave Assembly Model first describes a series of vicariance events associated with the late Miocene to early Pliocene ‘Bouse Formation’ (reviewed in Mulcahy et al., 2006), late Pliocene orogenesis of the Sierra Nevada and Transverse Ranges (e.g. Jones et al., 2004; Warrick & Mertes, 2009), and uplift of the western Mojave Desert (Cox et al., 2003). The model then predicts that additional diversification occurred as aridadapted populations allopatrically diverged in rain-shadowed basins during the Pliocene, and in desert basins and fragmented arid refugia throughout the Pleistocene, followed by late Pleistocene range expansion. As this model continues to be refined, we argue that the Mojave and Sonoran deserts are becoming one of the best regions in which to study the historical assembly of desert biotas, and a perfect setting for investigating the effects of Neogene tectonics and Pleistocene climate fluctuations on the generation and maintenance of biological diversity [see Fig. 1 in Graham et al. (2013a) for a visual and textual description of the model]. Our current knowledge of the genetic patterns in Mojave and Sonoran terrestrial fauna is largely a product of phylogeographical studies conducted on vertebrates, but genetic patterns are also beginning to arise from studies of codistributed arachnids (Graham et al., 2013a,b; M.R.G., unpubl. data). In a recent taxonomic assessment of the tarantulas of

Table 1 Recent phylogeographical studies of terrestrial fauna that co-occur with Aphonopelma mojave group tarantulas and whether taxa are thought to have undergone pre-Pleistocene divergences (PPD) in the Mojave and Sonoran deserts, south-western USA. Taxon Herpetofauna Anaxyrus punctatus – red-spotted toad Chionactis occipitalis – shovel-nosed snake Crotaphytus bicinctores – collared lizard Crotalus cerastes – sidewinder Lichanura trivirgata – rosy boa Sceloporus magister – desert spiny lizard Uma spp. – fringe-toed lizards Xantusia vigilis – desert night lizard Mammals Ammospermophilus spp. – antelope squirrel Xerospermophilus spp. – ground squirrel (two species) Chaetodipus penicillatus – pocket mouse Thomomys bottae – pocket gopher Arthropods Homalonychus spp. – ground-dwelling spiders (two species) Hadrurus arizonensis – Arizona giant hairy scorpion Paruroctonus becki – Beck desert scorpion

PPD

Source

No Yes Yes Yes No Yes No Yes

Jaeger et al. (2005) Wood et al. (2008a) McGuire et al. (2007) Pece (2004) Wood et al. (2008b) Leache & Mulcahy (2007) Gottscho et al. (2014)* Leavitt et al. (2007)

Yes/No† No No No

Mantooth et al. (2013)* Bell et al. (2010)* Jezkova et al. (2009) Alvarez-Casta~ neda (2010)

Yes No Yes

Crews & Hedin (2006) Graham et al. (2013a)* Graham et al. (2013b)*

*Indicates phylogeographical studies not included in Wood et al. (2012). † Different fossil calibrations produced conflicting divergence date estimates. Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

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M. R. Graham et al.

Figure 1 Map of the south-western USA depicting locations of Aphonopelma mojave group samples used in genetic analyses. Numbers correspond to locality data presented in Appendix S1.

the Aphonopelma mojave group (comprising A. mojave Prentice, 1997; A. joshua Prentice, 1997; and at least one undescribed species), Hendrixson et al. (2013) identified significant genetic structure across the Mojave and Sonoran deserts, suggesting that the group has potential for providing novel phylogeographical insights in these regions. In this contribution, we expanded the genetic dataset provided by Hendrixson et al. (2013) by sequencing mitochondrial DNA (mtDNA) from additional A. mojave group samples collected throughout the Mojave and Sonoran deserts. We then used these data to conduct a phylogeographical appraisal of the A. mojave group. Although we recognize that there are potential shortcomings with using only maternally inherited mtDNA (Edwards & Bensch, 2009), the species groups evaluated here have been strongly supported using hundreds of nuclear loci from a novel anchored phylogenomics probe set designed for spiders (C.A.H., unpubl. data). Furthermore, mtDNA has proven to be a useful tool in phylogeographical studies of other codistributed arachnids (Crews & Hedin, 2006; Hamilton et al., 2011; Graham et al., 2013a,b), to which our mtDNA dataset can be easily compared. We explored the mtDNA data by conducting phylogenetic analyses using Bayesian inference (BI) and maximum likelihood (ML). We then placed the phylogeny in a temporal context using a relaxed molecular clock and a lineagesthrough-time plot (LLT plot) to assess whether the A. mojave group represents another desert taxon that diversified prior to the Pleistocene. We used network and demographic analyses, as well as species distribution modelling, to estimate the demographic histories of clades within the A. mojave group inferred by the phylogenetic analyses, and investigate these histories in the light of the Mojave Assembly Model. We predict that if turret-building tarantulas were influenced by the 1054

events outlined by the Mojave Assembly Model then their phylogeographical patterns should be similar to those observed in codistributed taxa, and species distribution models should suggest a fragmented distribution during the Last Glacial Maximum (LGM). Furthermore, Pleistocene fragmentation is known to cause lineage formation and higher genetic diversity in areas where climates remained suitable (Hewitt, 2000), so the genetic data should geographically corroborate predictions made by the species distribution models. MATERIALS AND METHODS Taxon sampling Using species distribution models and literature records as a guide (Prentice, 1997), we collected tarantulas from 18 new general locations (sites within 5 km of each other were pooled in some analyses) by searching for active burrows. When added to the samples from Hendrixson et al. (2013), our sampling covered a total of 82 general locations across the Mojave and Sonoran deserts (Fig. 1, and see Appendix S1 in Supporting Information). Tarantulas were collected from burrows using a combination of flooding and excavation techniques. Wandering adult males were collected opportunistically. Muscle tissue was collected by removing two to four legs from the right side. These legs were immediately submerged in RNAlater RNA Stabilization Reagent (Qiagen, Valencia, CA, USA) and stored at 80 °C. Voucher specimens were preserved in 80% ethanol and stored at 20 °C for at least four weeks. The vouchers have been deposited in the Auburn University Museum of Natural History (AUMNH; Auburn, AL, USA) collection. Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

Turret-building tarantula diversification Molecular techniques Genomic DNA was isolated from leg tissue using the DNeasy Tissue Kit (Qiagen, Valencia, CA, USA) and stored at 20 °C prior to amplification. We amplified the mtDNA COI gene using the protocol outlined in Hendrixson et al. (2013). Primers used to amplify this region of COI included C1-J-1751 ‘SPID’ (50 -GAG CTC CTG ATA TAG CTT TTC C-30 ) and C1-N-2776 (50 -GGA TAA TCA GAA TAT CGT CGA GG-30 ) (Hedin & Maddison, 2001). Thermal cycler parameters were: initial denaturation at 95 °C for 2 min; 30 cycles of denaturation at 96 °C for 30 s, annealing at 48 °C for 30 s, and extension at 72 °C for 1 min; and a final extension at 72 °C for 2 min. PCR products were purified using a 6:1 ratio of PCR product to ExoSAP-IT (GE Healthcare, Piscataway, NJ, USA) and then sequenced in both directions using the amplification primers on an ABI 3130 Genetic Analyzer with the ABI Big Dye Terminator ver. 3.1 Cycle Sequencing Ready Reaction Kit. All double-stranded DNA fragments were manually edited in Geneious Pro 4.8.5. (Biomatters Ltd, Auckland, New Zealand). Phylogenetic analyses Sequences were aligned in Geneious using muscle (Edgar, 2004) and the best-fitting models of nucleotide substitution were selected for each COI codon position using jModelTest 0.1.1 (Posada, 2008). Collapse 1.2 (Posada, 2004) was used to remove redundant haplotypes. Unique haplotypes were analysed phylogenetically; the trees were rooted using Aphonopelma eutylenum Chamberlin, 1940, Aphonopelma hentzi (Girard, 1852), Aphonopelma paloma Prentice, 1993, Aphonopelma nayaritum Chamberlin, 1940 and Aphonopelma reversum Chamberlin, 1940. Appropriate substitution models were used to assess phylogenetic patterns via Bayesian inference (BI) implemented in MrBayes 3.2.2 (Ronquist & Huelsenbeck, 2003) and maximum likelihood (ML) implemented with RAxML (Stamatakis, 2006). Both analyses were conducted on the Cyberinfrastructure for Phylogenetic Research cluster (CIPRES Gateway 3.1) at the San Diego Supercomputer Center. The ML analyses consisted of 1000 bootstrap pseudoreplicates for the best tree. The BI analysis was run for 40 million generations with four chains (one cold, three heated), with model parameters unlinked across character partitions, and sampling every 4000 generations. We modified the heating parameter until state-swap frequencies were between 10% and 70%, and discarded the first 25% of trees as burn-in. We verified the convergence of all Markov chains on similar model parameters with Tracer 1.6 (Rambaut & Drummond, 2007). Network analyses We used the program Network 4.5.1.6 (Fluxus Technology, Clare, UK; available at: http://www.fluxus-engineering.com/) Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

to construct median-joining networks of the mtDNA haplotypes for five of the six major lineages recovered in the phylogenetic analyses. We did not construct a network for samples for a unique clade from the Owens Valley (site 11, see Results) because they contained identical haplotypes. We constructed each network with transversions/transitions weighted 3:1 and used the parsimony option to remove excessive links (Polzin & Daneshmand, 2003). Demographic history Genetic indices for groups identified by the phylogenetic and network analyses were estimated in Arlequin 3.11 (Excoffier et al., 2005). We used Fu’s FS (Fu, 1997) and mismatch analyses (Rogers, 1995) to test for genetic signals of demographic expansion within predefined groups. Because some clade and subclade sample sizes were small, we also used DnaSP 5.10.01 (Librado & Rozas, 2009) to conduct R2 tests, which perform well when detecting demographic growth in small sample sizes (Ramos-Onsins & Rozas, 2002). The significance of the R2 values were assessed using 5000 coalescent simulations. We inferred changes in effective population size over time for each clade using the Bayesian skyline method (Ho & Shapiro, 2011) implemented in beast 1.7.2 (Drummond & Rambaut, 2007). Time and population size were calibrated with a strict clock prior with a substitution rate of 0.0169 site1 lineage1 Myr1, which is a strongly supported COI rate for arthropods (Papadopoulou et al., 2010). Strict clock priors are appropriate for the estimation of divergences in datasets with low variation (Brown & Yang, 2011). The bestfitting model of nucleotide substitution was determined for each clade using jModelTest. Analyses were run using the substitution models for 20 million steps and sampled every 1000 steps. All operators were optimized automatically. Effective sample size (ESS) values were assessed and results of the analyses were visualized using Tracer 1.6 (Rambaut & Drummond, 2007). Subclades I, IIb and IIc were omitted from the analyses because of a low number of polymorphic sites and small sample sizes. Divergence dating and lineages-through-time plots We generated a time-calibrated phylogeny with beast using a relaxed molecular clock. beast analyses were performed using the best-fitting substitution model for each COI codon position as previously estimated. We unlinked substitution and clock models across data partitions (but with a linked tree model), and conducted all analyses using an uncorrelated lognormal relaxed clock. No appropriate tree prior was obvious because our dataset spans divergent lineages that represent at least three different species (Hendrixson et al., 2013), so we replicated analyses using the tree priors ‘speciation: birth–death incomplete sampling’, ‘speciation: Yule process’ and ‘coalescent: constant size’ (following the methods of Hedin et al., 2013). We conducted three replicate 1055

M. R. Graham et al. Markov chain Monte Carlo (MCMC) runs for 50 million generations each, and sampled every 1000 generations. We used Tracer to assess convergence among runs and to confirm that ESS values exceeded 200 for all parameters. We used LogCombiner to combine separate tree files and TreeAnnotater (beast package) to construct a maximum clade credibility tree. Also as in Hedin et al. (2013), we calibrated the molecular clock with a mean SD of 0.0169 0.0019 (Papadopoulou et al., 2010). To assess the overall tempo of diversification, we used a lineages-through-time (LTT) plot to visualize the number of lineages in the phylogeny as a function of clade age. These plots are constructed from a previously inferred phylogeny by working backwards through time, counting the accumulation in number of ancestral lineages. If diversification has been constant over time, then a straight line is predicted when the number of lineages is plotted on a logarithmic scale; significant deviations from this expectation indicate that rates of diversification or extinction may be changing through time. To generate the LTT plot, we used the ape (Paradis et al., 2004) and geiger (Harmon et al., 2008) packages, as well as the function ‘rbdtree.n3’ (Nick J. Matzke; available at http://ib.berkeley.edu/courses/ib200b/labs/ lab12/rbdtree.n3.R), in the R computing environment. We imported the beast consensus tree, pruning the outgroups so that only the A. mojave group and immediate sister-lineages were represented, and log-transformed the data along the y-axis. In order to evaluate whether the LTT plot deviated from a null distribution, we plotted null distributions under Yule and pure-birth processes, using the branching times and lambda (k) from the dated tree. We also evaluated the LTT plot by running 1000 simulations under different parameters (e.g. pure birth vs. birth = 0.3 and death = 0.1) and plotting the A. mojave group LTT against those results. Species distribution models We used a total of 163 unique occurrence points representing collection sites from our genetic samples, as well as georeferenced records from the American Museum of Natural History and the collection of Tom Prentice (AUMNH) for species distribution modelling. We used the program Maxent 3.3.3 (Phillips et al., 2006) to construct species distribution models, following the procedures outlined in Graham et al. (2013a). In brief, we screened 19 bioclimatic layers (http://www.worldclim.org/bioclim) by using values from grid cells containing occurrence records to identify and remove highly correlated (Pearson’s correlation coefficient > 0.75) layers. After screening for correlation and putative biological relevance, the final predictor layers comprised the following 10 bioclimatic layers: BIO1, annual mean temperature; BIO2, mean diurnal temperature range; BIO7, annual temperature range; BIO8, mean temperature of the wettest quarter; BIO9, mean temperature of the driest quarter; BIO13, precipitation of the wettest month; BIO14, pre1056

cipitation of the driest month; BIO15, precipitation seasonality; BIO17, precipitation of the driest quarter; and BIO18, precipitation of the warmest quarter. We ran Maxent using logistic output by starting with default settings and random seeding for 10,000 iterations. We used a default prevalence of 0.9 to incorporate our a priori knowledge of the natural history of the A. mojave group. We applied an equal training sensitivity and specificity threshold and tested model robustness by cross-validation over 10 replicates. We relied on the method available in Maxent for determining the area under the receiver operating characteristic curve (AUC) to assess model performance. To examine the distribution of suitable climate for the A. mojave group during glacial periods, we then projected the models onto simulated climates for the LGM (hindcasting) derived from the Community Climate System Model (CCSM; Otto-Bliesner et al., 2006) and the Model for Interdisciplinary Research on Climate (MIROC; Hasumi & Emori, 2004), using the same settings as above. Climatic suitability was displayed in ArcGIS 10.1 (ESRI, Redlands, CA, USA) by converting continuous Maxent outputs into binary grids using the maximum training sensitivity plus specificity threshold, which has performed well in comparisons of various threshold criteria (Liu et al., 2005; Jimenez-Valverde & Lobo, 2007). RESULTS Phylogenetic analyses We sequenced 924 bp of COI from 40 individuals of A. mojave (sensu lato) and two specimens of A. joshua (GenBank accession numbers JX946016–JX946107 and KP677292– KP677333). Combined with the samples from Hendrixson et al. (2013) and Hamilton et al. (2014), our sampling effort yielded a total of 128 samples of A. mojave (sensu lato) and six of A. joshua. In the 42 new samples collected for this study, we identified 24 novel haplotypes, totalling 67 unique haplotypes for the entire dataset. Substitution models selected under the AIC for each codon position were as follows: first, HKY+I+G; second, GTR+I+G; third, GTR+I+G. The COI topologies obtained by BI and ML analyses were nearly identical. The BI and ML phylogenies (Fig. 2) strongly supported the monophyly of the A. mojave group and recovered six major geographically structured clades that were strongly supported by both analyses. Two of these clades – a unique clade near the Owens Valley (‘Owens clade’) and a clade centred on the Amargosa River valley (‘Amargosa clade’) – were not recovered in Hendrixson et al. (2013). Other geographically delimited groups of A. mojave (sensu lato) consisted of individuals distributed as follows: in the north-western Mojave Desert just south of the southern Sierra Nevada (‘north-western clade’); along the south-western Mojave Desert north of the San Gabriel and San Bernardino mountains (‘south-western clade’); and widespread throughout the eastern Mojave Desert and part Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

Turret-building tarantula diversification

Figure 2 Majority rule (50%) consensus tree depicting results of a Bayesian phylogenetic analysis of mtDNA (COI) sequences for the Aphonopelma mojave group in the south-western USA. Vertically arranged bars indicate geographically structured clades. Bayesian posterior probability values are displayed above nodes and maximum likelihood bootstrap values are below nodes.

of the northern Sonoran Desert east of the Colorado River (‘eastern clade’). Samples of A. joshua from the southern Mojave Desert also formed a distinct clade. Although all of these main clades were strongly supported by BI and ML analyses, neither criterion could resolve the relationships among them with strong support. Network analyses Haplotype networks constructed from the COI data (Fig. 3) revealed a high degree of phylogeographical structure. The largest group, the eastern clade, was geographically structured into three main subclades: a group of haplotypes in the northwestern and central Mojave Desert (subclade I), haplotypes in Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

the central and northern Mojave (subclade II), and haplotypes east of the Colorado River (subclade III). Haplotypes from subclade II were organized into three additional subclades: subclade IIa along the border of California and Nevada, subclade IIb in the northern Mojave Desert from north of the Spring Mountains north-east to south-western Utah, and subclade IIc from the western part of the Mojave National Preserve. Samples from subclade IIb formed a star-shaped pattern with the most common haplotype found at all the sample sites throughout the north-eastern Mojave Desert. Most of the haplotypes in subclade III were divided into two additional subclades: subclade IIIa nested between the Gila, Bill Williams and Colorado rivers, and subclade IIIb distributed north of the Bill Williams River and south and east of the Colorado River. 1057

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Figure 3 Map of the south-western USA and networks constructed from mtDNA (COI) sequences from major lineages within the Aphonopelma mojave group. Each circle in the network represents a unique haplotype. Circle size in both the map and network are proportional to sample size. Colours in the map correspond to the colours of each of the groups identified in the haplotype network. The black circle in the map indicates a divergent lineage (the Owens clade) that was only discovered at one sample site and contained only one haplotype. Small black circles in the networks represent hypothetical haplotypes.

Subclade IIIa and subclade IIIb both comprised star-shaped clusters of haplotypes. Haplotypes from the south-western clade formed a large star shape with long branches. Samples from east of the San Bernardino Mountains and close to localities of A. joshua were slightly differentiated from the other haplotypes of the southwestern clade. Samples from the Coxcomb Mountains (site 46) were highly differentiated (67 mutational steps) from other haplotypes of the south-western clade. Haplotypes from the north-western clade were differentiated into northern and southern haplotypes, with northern haplotypes forming a starshaped cluster and southern samples all possessing the same haplotype. The three samples of the Amargosa clade all possessed unique haplotypes with long branches grouped together at hypothetical ancestral haplotypes. Haplotypes from A. joshua were connected by ancestral haplotypes to samples from the westernmost locality (site 49), separated from all other A. joshua haplotypes by at least 28 mutational steps. Demographic history Values of p for each clade and subclade ranged from less than 0.001 to 0.054, and values of h ranged from 0.591 to 0.957. Values of Fu’s FS were negative for subclades IIb and IIIa (Table 2), indicating deviations from mutation–drift equilibrium, as would be expected for populations that had undergone recent expansion or selection (Fu, 1997); FS was positive for all other subclades. No R2 values were statistically significant. A unimodal mismatch distribution, which also indicates recent demographic expansion or selection (Rogers & Harpending, 1992), was observed only in subclade IIb. Distribution curves were multimodal for all other subclades, signifying that the populations may be at equilibrium. 1058

Parametric bootstraps resulted in significant sums of squared deviations (SSD) under the sudden-expansion model for subclades I, II, III, IIIa and IIIb and the north-western clade. Subclade III and the north-western clade exhibited significant SSD values under the spatial expansion model. Values of Harpending’s raggedness index (r) were significant for subclades II, III, IIIa and IIIb and the north-western clade for the sudden-expansion model, indicating that the model was not a good fit. No r values were significant under a model of spatial expansion, indicating that the model was a good fit for all clades. Bayesian skyline plots differed between clades (see Appendix S2). The plots suggest that population sizes of the eastern clade and subclades II, III and IIIb recently (< 100 ka) decreased and subsequently expanded, whereas the southwestern and north-western clades appear to have experienced recent population declines. The analyses suggest that the population sizes of subclades IIa and IIIa remained relatively stable during the late Pleistocene. Overall, the demographic analyses provided evidence of demographic expansion for all analysed clades except subclade IIa and the south-western clade. (See Table 2 for summary statistics and results from demographic analyses.) Divergence dating and lineages-through-time plot The topology resulting from the beast analysis (Fig. 4) was mostly identical to that generated with MrBayes and RAxML (Fig. 2). Mean estimates of the time to the most recent common ancestor (TMRCA) for the entire A. mojave group placed it in the late Miocene, ranging from 8.04 to 6.62 Ma in analyses run using the four different tree priors (Table 3). Analyses using all but the Yule tree prior suggested Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

0.005 0.243 0.064 0.285 0.177 – 0.044 0.112 0.091 0.037 0.033

0.810 0.370 0.650 0.390 0.380 – 0.280 0.080 0.510 0.710 0.330

multimodal multimodal multimodal multimodal unimodal – multimodal multimodal multimodal multimodal multimodal

that the subsequent diversification, which resulted in four additional nodes, occurred during the late Miocene. Only two nodes were estimated for the Pliocene – one representing the common ancestor of the eastern clade (2.69–2.66 Ma) and another indicating the common ancestor of the southwestern clade (3.45–2.99 Ma) when a divergent sample from the Coxcomb Mountains (site 46) was included; the mean TMRCA for the south-western clade was estimated to be 1.37–1.34 Ma when the divergent sample was not included. The remaining 60 nodes had mean estimates in the Pleistocene. The TMRCA for samples from three major lineages also fell within the Pleistocene: A. joshua (1.11–1.08 Ma), the Amargosa clade (0.94–0.88 Ma), and the north-western clade (0.66–0.70 Ma). The LTT plots (inset in Fig. 4, and see Appendix S3) depicted a clear deviation from the null distribution, with a distinct increase in lineage formation during the late Miocene. Species distribution models

0.004 0.092 0.023 0.108 0.005 – 0.042 0.044 0.044 0.032 0.111

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*Distribution curves were visually assessed using the sudden expansion model. †We did not perform all analyses for subclade IIc due to an inadequate sample size.

89 11 37 12 19 6 38 16 20 17 16 Eastern clade Subclade I Subclade II Subclade IIa Subclade IIb Subclade IIc† Subclade III Subclade IIIa Subclade IIIb South-western clade North-western clade

43 5 15 5 5 5 19 10 9 10 5

176 58 58 17 3 17 68 22 25 114 27

0.044 0.022 0.021 0.009 < 0.001 0.011 0.054 0.007 0.009 0.044 0.012

0.957 0.764 0.869 0.758 0.591 0.933 0.949 0.933 0.874 0.904 0.800

2.01 7.39 3.93 3.79 -3.66 – 1.83 -0.97 1.63 7.67 7.32

0.754 0.996 0.899 0.945 < 0.001 – 0.786 0.303 0.801 0.993 0.994

0.115 0.158 0.144 0.221 0.108 0.266 0.156 0.139 0.157 0.153 0.204

0.799 0.557 0.853 0.975 0.103 0.773 0.919 0.500 0.794 0.747 0.974

0.005 0.147 0.060 0.189 0.005 – 0.037 0.047 0.055 0.048 0.137

0.190 < 0.001 < 0.001 0.080 0.530 – < 0.001 0.020 < 0.001 0.060 0.050

0.005 0.243 0.064 0.285 0.177 – 0.044 0.112 0.091 0.037 0.331

0.040 0.010 < 0.001 0.120 0.420 – < 0.001 0.010 0.020 0.120 0.010

0.770 0.270 0.830 0.200 0.360 – < 0.001 0.090 0.170 0.450 0.030

Distribution curve* P r P SSD P n Lineage

Haps

PS

p

h

FS

P

R2

P

SSD

P

r

Spatial expansion Sudden expansion

Table 2 Nucleotide diversity (p), haplotype diversity (h) and results of Fu’s FS, R2, and mismatch analyses with sum of squared deviations (SSD) and Harpending’s raggedness values (r) for tarantulas of the Aphonopelma mojave group in the south-western USA. PS, number of polymorphic sites (out of 912); Haps, number of haplotypes.


High AUC scores for both training and testing data (both > 0.95) indicated that the species distribution model performed significantly better than random (Raes & ter Steege, 2007). The species distribution model constructed under current climatic conditions (Fig. 5a) depicted largely contiguous suitable climate across the majority of the Mojave Desert and part of the northern Sonoran Desert. Climate was predicted to be unsuitable at the highest elevations and the extremely hot, low regions along the Colorado River Valley and the Death Valley drainage. Although there are no records of the A. mojave group from the region, climate was predicted to be suitable in the Peninsular Ranges south of the San Bernardino Mountains. LGM models based on CCSM and MIROC (Fig. 5b,c) both predicted suitable climate for the A. mojave group in the western Mojave Desert, the north-western Sonoran Desert, and the southern San Joaquin Valley. The CCSM model depicted a reduction in suitable habitat in the northern Mojave Desert and northern areas east of the Colorado River. Both LGM models predicted suitable climate along the Colorado River Valley, but suitable regions extended further north in the MIROC model to include the north-eastern Mojave Desert and extending into extreme south-western Utah. In summary, both models suggested that the LGM experienced an increase in suitable habitat in the north-western Sonoran Desert, and a downward shift in the elevation of suitable habitat, especially along the Colorado River Valley. Although both LGM models highlighted southern portions of the San Joaquin Valley as suitable, no tarantulas of the A. mojave group have been documented from the region and the models do not predict any connectivity with the Mojave Desert where they currently occur. The precipitation of the driest month played a large role in modelling the distribution of the A. mojave group, as it contributed 32% of the current model, 32.3% of the CCSM model, and 31.7% of the MIROC model. 1059

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Figure 4 Maximum clade credibility tree resulting from beast analysis of mtDNA (COI) sequences from tarantulas of the Aphonopelma mojave group in the south-western USA using a birth–death tree prior (see Table 3 for results using different tree priors). Inset: lineages-through-time (LTT) plot. The null distribution under a Yule process is depicted, using the branching times and lambda (k) from our dated tree, with white and grey shading representing the confidence intervals (see Appendix S3 for alternative null distributions). LTT plots represent the number of lineages with living descendants in a clade against time. If l = 0 and k are constant, LTT plots will show exponential growth. Significant deviations from this expectation indicate that diversification is changing with time. Our data suggest that an increased shift in diversification occurred in the late Miocene.

DISCUSSION Pre-Pleistocene divergence Despite the relatively narrow distribution of the A. mojave species group, species were highly structured, with no fewer than six major clades identified in BI and ML analyses

(Fig. 2). Although samples from each of these tarantula clades appear to share common ancestors in the late Pliocene and early Pleistocene, molecular-clock estimates suggest that they all have origins in the late Miocene (Fig. 4). Importantly, variances around mean date estimates (95% highest posterior density) do not include any time spans more recent than the middle Pliocene. Therefore, even if the molecular

Table 3 Comparison of the time to most recent common ancestors for major mtDNA lineages within the Aphonopelma mojave group in the south-western USA estimated with beast using alternative tree priors. Mean estimates are provided in millions of years (Ma) with 95% highest posterior density (HPD) intervals provided in parentheses. Data are provided for the south-western clade with (1) and without (2) disjunct samples from the Coxcomb Mountains in California (site 46). Lineage

Birth–death (Ma)

Birth–death incomplete (Ma)

Yule (Ma)

Coalescent (Ma)

Entire A. mojave group Eastern clade South-western clade (1) South-western clade (2) North-western clade A. joshua Amargosa clade

7.99 2.67 3.45 1.35 0.65 1.08 0.88

7.94 2.66 3.43 1.34 0.65 1.08 0.88

6.62 2.68 2.99 1.37 0.70 1.11 0.94

8.04 2.69 3.40 1.36 0.66 1.09 0.90

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(5.71–10.50) (1.92–3.53) (2.20–4.81) (0.88–1.84) (0.35–1.00) (0.64–1.57) (0.49–1.31)

(5.75–10.39) (1.88–3.49) (2.20–4.82) (0.88–1.83) (0.36–0.99) (0.64–1.56) (0.50–1.32)

(4.69–8.72) (1.88–3.58) (1.80–4.25) (0.89–1.92) (0.35–1.12) (0.61–1.65) (0.50–1.45)

(5.75–10.59) (1.91–3.55) (2.12–4.74) (0.89–1.87) (0.35–1.00) (0.64–1.59) (0.51–1.35)



(a)

(b)

(d)

(c)

(e)

Figure 5 Maps displaying species distribution models (a–c) and the geographical correlation between major clades of the Aphonopelma mojave group in the south-western USA (d) and displacement of mountain ranges during the Miocene (e). Species distribution models represent climate predicted as suitable (dark shading) under (a) current conditions, and under (b) CCSM and (c) MIROC simulations of late glacial climates. Arrows indicate displacement with respect to stable North America from 8–6 Ma. Black dots indicate locations of genetic samples used in this study. The approximate distributions of the Mojave and Sonoran deserts are displayed as black dotted lines. The kinematic model (e) was adapted from McQuarrie & Wernicke (2005).

clock estimate is slightly off, the A. mojave group can be confidently added to the growing list of Mojave and Sonoran desert taxa that exhibit pre-Pleistocene divergences. The data now suggest that over half (9 out of 16) of the arid-adapted fauna in the Mojave and Sonoran deserts are the products of ancient rather than recent diversification. Of those that exhibit pre-Pleistocene divergences, the A. mojave group is the only taxon that cannot be categorized into one of the three genetic patterns outlined by Wood et al. (2012): (1) pre-Pleistocene divergences across the Colorado River; (2) pre-Pleistocene divergences across the Mojave/Sonoran ecotone; and (3) pre-Pleistocene divergences across both regions. Instead, pre-Pleistocene divergences in the A. mojave group increased in number from east to west. Samples from the eastern half of the distribution consisted entirely of haplotypes of the eastern clade and, although highly structured, diversification among eastern populations occurred almost entirely during the Pleistocene (Fig. 4). The western half of the Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

distribution, however, comprises five major clades, each occupying a range less than half the area of the eastern clade. This geographical scattering of major clades with relationships dating to the Miocene is more complex than the three patterns outlined by Wood et al. (2012), and instead appears to be correlated with geological features such as mountain ranges. Interestingly, the late Miocene time frame during which the western clades were estimated to have diverged from each other is precisely when tectonic activity is known to have displaced regional mountain ranges to the north-west (McQuarrie & Wernicke, 2005). We therefore propose that the increased number of pre-Pleistocene divergences in the western half of the distribution of the A. mojave group can be attributed to Miocene extensional tectonics. Like other species of Aphonopelma in North America, the A. mojave group is well suited to arid environments, but they do not inhabit many of the hottest low-elevation regions, such as Death Valley and the Colorado River Valley. These tarantulas 1061

M. R. Graham et al. are generally found on alluvial deposits along the edges of such valleys at elevations a few hundred metres higher than the valley floor, where they construct shallow burrows with distinct turret-like mounds surrounding the entrances (Prentice, 1997). Hendrixson et al. (2013) tested this elevational pattern by conducting surveys at various elevations in the central Mojave Desert near Death Valley and only found tarantulas of the A. mojave group at elevations above 950 m. Thus, dispersal among populations of turret-building tarantulas is likely to be limited by hot, low-elevation valleys; landscape deformations such as crustal extensions may thus have caused the fragmentation and subsequent allopatric diversification in this group. Miocene tectonics in the western Mojave and Sonoran deserts are best understood in the context of the palaeogeography of the larger Basin and Range Province, an expanse of valleys trending north–south and mountain ranges spanning North American aridlands from Idaho and Oregon south to north-western Mexico. The Basin and Range Province began to form in the Oligocene, when tectonic activity along the Pacific–North American boundary caused the region to collapse away from the more static Colorado Plateau through the formation of metamorphic core complexes and crustal extension. In a detailed synthesis of kinematic (displacement) data, McQuarrie & Wernicke (2005) demonstrated that the Basin and Range Province began to form with east-to-west extension in the Great Basin Desert through the Oligocene. The Mojave and Sonoran deserts then developed as the region underwent extension trending north-east to south-west during the early and middle Miocene. During the late Miocene – the time frame for which we estimated A. mojave group diversification – deformation of the Mojave and Sonoran deserts underwent a shift. Mountain ranges in the east remained stable, while those in the west underwent rapid displacements trending south-east to north-west (see McQuarrie & Wernicke, 2005 for details and an animated version of this model). Intriguingly, this region of increased and directionally shifted displacement is geographically congruent with the distribution of major clades in the A. mojave group (Fig. 5d,e). We therefore hypothesize that the six clades of the A. mojave group are the products of vicariance as populations became isolated when displaced mountain ranges in the western Mojave and Sonoran deserts formed new low-elevation valleys that operated as barriers to dispersal. The same model of diversification was proposed to explain the origins of major clades of mid-elevation scorpions in the Eastern California Shear Zone (in the north-western Mojave Desert) during the Pliocene (Graham et al., 2013b). As far as we know, however, this is the first study to invoke Miocene tectonic activity to explain contemporary phylogeographical patterns in taxa of the Mojave and Sonoran deserts. Mojave Assembly Model The Mojave Assembly Model predicts that terrestrial fauna with ancient distributions spanning the Colorado River would have been sundered by development of the Bouse For1062

mation and Colorado River between 9 and 4 Ma (reviewed in Mulcahy et al., 2006). The distribution of the eastern clade does appear to be constrained by the Colorado River to the south, but the northern portion of the eastern clade stretches well beyond the river, extending as far west as the Panamint Range near Death Valley. Subclades within the eastern clade do form a discontinuity at the Colorado River, but the molecular-clock analysis places this split during the Pleistocene (Fig. 4), which is much too recent to be a product of vicariance at the Bouse Embayment. The Mojave Assembly Model also predicts that desert taxa would have been isolated in basins in the western Mojave Desert and along the Lower Colorado River Valley 4–2 Ma. The northwestern clade, south-western clade and eastern clade currently inhabit these basins. This part of the model does not, however, explain the distributions of the Amargosa clade, the Owens clade and A. joshua, all of which diverged in the Miocene and obviously did not go extinct during the Pliocene. The Mojave Assembly Model predicts that arid-adapted taxa were repeatedly fragmented during the Pleistocene, first as isolated desert basins began to develop, and then as climate fluctuations repeatedly produced pluvial periods that further fragmented desert habitats. The model concludes with range expansions out of arid Pleistocene refugia as climates warmed during the Holocene. LGM species distribution models for tarantulas of the A. mojave group predict that suitable climate was greatly reduced in the north-eastern Mojave Desert (Fig. 5b,c). This hypothesis is strongly supported by genetic data from populations of subclade IIb that currently inhabit the north-eastern Mojave Desert. A negative value of Fu’s FS, unimodal mismatch distribution and star-shaped haplotype network (Table 2, Fig. 3) all suggest that this clade underwent a recent demographic expansion. Northern regions were thus only recently colonized by turret-building tarantulas, subsequent to the LGM as climates warmed. Unlike predictions from the Mojave Assembly Model, LGM species distribution models suggest that climates remained suitable for tarantulas throughout much of the remaining Mojave and Sonoran deserts (Fig. 5b,c). Demographic analyses of COI data mostly contradict this interpretation, however, providing evidence that all clades except the south-western clade underwent recent demographic expansions (samples of the Owens clade were not included in these analyses). Furthermore, subclades IIIa and IIIb of the eastern clade both form star-shaped clusters in the haplotype network (Fig. 3), which is indicative of recent population expansion. Given these patterns and the distribution of suitable habitat in the MIROC hindcast (Fig. 5c), turret-building tarantulas east of the Colorado River were probably restricted to the Lower Colorado River Valley at the LGM. A similar trend was recently discovered in Arizona hairy scorpions, which are thought to have exploited southern and northern refugia along the Lower Colorado River Valley during Pleistocene glacial cycles (Graham et al., 2013a). In summary, the phylogeography of the A. mojave group can only be partly explained by the Mojave Assembly Model. Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

Turret-building tarantula diversification The model failed to produce meaningful predictions for late Miocene and Pliocene levels of diversification and only moderately predicted phylogeographical patterns for the Pleistocene. We therefore propose that the Mojave Assembly Model should be expanded to include Miocene and Pliocene tectonic activity as additional vicariance events that influenced the diversification of Mojave and Sonoran desert taxa. Furthermore, we suggest that for the period 4–2 Ma, the model should incorporate the following additional regions as potential early desert basins: the Amargosa Valley, the Owens Valley and low elevations within and around Joshua Tree National Park. Taxonomic considerations Sampling of 18 new locations resulted in the discovery of two new major mitochondrial (maternal) lineages within the A. mojave group, one from the Owens Valley and the other from areas adjacent to the Amargosa Valley. Hendrixson et al. (2013) had already identified the remaining clades and subjected them to five different species-delimitation approaches. The number of species identified by each approach varied, but the authors suggested that the group contained no fewer than three and as many as five species. If their analyses were conducted using our expanded sampling, we suspect that the Owens clade and Amargosa clade would add at least two additional species to their totals. Thus, current taxonomy based largely on morphology significantly underestimates species-level diversity in this group, which is in dire need of systematic revision. CONCLUSIONS Incorporating extensional tectonics into models of evolution in the Mojave and Sonoran deserts could precipitate a radical shift in our understanding of the historical assembly of aridadapted biotas. Previous authors repeatedly uncovered prePleistocene divergences within taxa inhabiting these deserts, but most attributed those patterns to vicariance at the Bouse Embayment (reviewed in Mulcahy et al., 2006). Others suggested that steep environmental gradients could maintain genetic structure along the Sonoran/Mojave ecotone (Wood et al., 2012), as seen in models of parapatric speciation (Doebeli & Dieckmann, 2003). We advocate a novel hypothesis, that many pre-Pleistocene divergences, like those uncovered in the A. mojave group, may actually be the result of vicariance associated with extensional tectonics. Recent geological reconstructions, including animated hypotheses of landscape evolution in the North American south-west (McQuarrie & Wernicke, 2005; McQuarrie & Oskin, 2011), have been underutilized and would provide an ideal framework for testing biogeographical hypotheses in the Mojave and Sonoran deserts and the larger Basin and Range Province. The six pre-Pleistocene divergences uncovered in tarantulas of the A. mojave group emphasize the influence that ancient geological events may have had on current patterns of biodiversity in North American deserts. Journal of Biogeography 42, 1052–1065 ª 2015 John Wiley & Sons Ltd

ACKNOWLEDGEMENTS We thank the National Park Service (Joshua Tree National Park, JOTR-2008-SCI-0017 and JOTR-2010-SCI-0015; Mojave National Preserve, MOJA-2010-SCI-0032), the State of California Departments of Fish and Game (802021-05) and Parks and Recreation [9-2195(08III2010)] for providing permits to collect the tarantulas used in this study. We also thank Tom Prentice and Wendell Icenogle for passing on their vast knowledge of the group. George Graham provided logistical support. Funding for this study was provided by the National Science Foundation (DEB-0841610 to J.E.B. and B.E.H.), Millsaps College (to B.E.H.) and Auburn University (to J.E.B.). REFERENCES Alvarez-Casta~ neda, S.T. (2010) Phylogenetic structure of the Thomomys bottae–umbrinus complex in North America. Molecular Phylogenetics and Evolution, 54, 671–679. Axelrod, D.I. (1983) Paleobotanical history of the western deserts. Origin and evolution of deserts (ed. by S.G. Wells and D.R. Haragan), pp. 113–129. University of New Mexico Press, Albuquerque, NM. Bell, K.C., Hafner, D.J., Leitner, P. & Matocq, M.D. (2010) Phylogeography of the ground squirrel subgenus Xerospermophilus and assembly of the Mojave Desert biota. Journal of Biogeography, 37, 363–378. Brown, R.P. & Yang, Z. (2011) Rate variation and estimation of divergence times using strict and relaxed clocks. BMC Evolutionary Biology, 11, 271. Cox, B.F., Hillhouse, J.W. & Owen, L.A. (2003) Pliocene and Pleistocene evolution of the Mojave River, and associated tectonic development of the Transverse Ranges and Mojave Desert, based on borehole stratigraphy studies and mapping of landforms and sediments near Victorville, California. Paleoenvironments and paleohydrology of the Mojave and southern Great Basin deserts (ed. by Y. Enzel, S.G. Wells and N. Lancaster), pp. 1–42. GSA Special Papers 368. Geological Society of America, Boulder, CO. Crews, S.C. & Hedin, M. (2006) Studies of morphological and molecular phylogenetic divergence in spiders (Araneae: Homalonychus) from the American southwest, including divergence along the Baja California Peninsula. Molecular Phylogenetics and Evolution, 38, 470–487. Doebeli, M. & Dieckmann, U. (2003) Speciation along environmental gradients. Nature, 421, 259–264. Drummond, A.J. & Rambaut, A. (2007) BEAST: Bayesian evolutionary analysis by sampling trees. BMC Evolutionary Biology, 7, 214. Edgar, R.C. (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Research, 32, 1792–1797. Edwards, S. & Bensch, S. (2009) Looking forwards or looking backwards in avian phylogeography? A comment on Zink and Barrowclough 2008. Molecular Ecology, 18, 2930–2933. 1063

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Editor: Melodie McGeoch

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