University of California Press

Chapter Title: Intraspecific Variation, Adaptation, and Evolution Chapter Author(s): Ryan E. O’Dell and Nishanta Rajakaruna

Book Title: Serpentine Book Subtitle: The Evolution and Ecology of a Model System Book Editor(s): Susan Harrison, Nishanta Rajakaruna Published by: University of California Press. (2011) Stable URL: https://www.jstor.org/stable/10.1525/j.ctt1pnqkb.10

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Intraspecific Variation, Adaptation, and Evolution

Ryan E. O’Dell, Bureau of Land Management Nishanta Rajakaruna, College of the Atlantic

Intraspecific variation refers to the genotypic (genetic) and resulting phenotypic (morphological and physiological) variation found within a species. Variation within a species is crucial for adaptation and evolution by natural selection. Over time, selection can result in genetically distinct populations of a species that are adapted to specific environmental conditions. Such populations are referred to as ecotypes. The study of distinct climatic, elevational, latitudinal, geographic, and edaphic ecotypes has provided much insight into the process of evolution by natu- ral selection (Briggs and Walters, 1997; Levin, 2000; Silvertown and Charlesworth, 2001). Edaphic islands, characterized by distinct soil characteristics, provide unique settings to study the factors and mechanisms promoting the generation and main- tenance of intraspecific variation (Kruckeberg, 1986; Rajakaruna, 2004). Soil char- acteristics are strongly influenced by topography and underlying geology. Topog- raphy affects soil depth, water availability, and solar exposure, whereas geology influences soil development rate (mineral weathering), soil physical properties in- cluding texture and water-holding capacity, and soil chemical properties including plant essential nutrients, as well as toxic elements (Mason, 1946a; Kruckeberg, 1986; Alexander et al., 2007; Rajakaruna and Boyd, 2008). Extreme soil physical or chemical properties are typical of edaphically stressful habitats. Plants often

Serpentine: The Evolution and Ecology of a Model System, edited by Susan Harrison and Nishanta Rajakaruna. Copyright  by The Regents of the University of California. All rights of reproduction in any form reserved.

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This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms 98 serpentine as a model in evolution respond to such edaphic stresses by ecotypic differentiation and subsequent spe- ciation. Various methods have been used to detect ecotypic differentiation. These include reciprocal or unilateral transplant studies conducted in field, glasshouse, or laboratory settings, as well as genetic analysis. Although reciprocal transplant studies (see Chapter 7 in this volume) compare the response of two or more popu- lations to their own, as well as other populations’ environments, unilateral trans- plant studies only examine the response of two or more populations to one of the population’s environments (typically serpentine and nonserpentine populations to serpentine soil). Due to the difficulty in establishing reciprocal or unilateral field transplant studies and the large time investment required to conduct them, trans- plant studies are more commonly carried out in controlled glasshouse or labora- tory settings. Habitat-correlated differences in morphology, growth rates, physiol- ogy, or allele frequencies between populations documented by these studies are regarded as evidence for ecotypic differentiation. This chapter focuses on how distinct soil types have resulted in ecotypic dif- ferentiation and adaptation. We begin with evidence for ecotypic differentiation in chemically stressful edaphic environments, including saline soils and metalliferous mine tailings, and then proceed to highlight ecotypic differentiation in response to serpentine soils. The discussion includes a summary of the adverse physical and chemical characteristics of serpentine soils, key plant morphological and physio- logical mechanisms involved in serpentine soil tolerance, how ecotypic differen- tiation leads to the origin of new species (also see Chapter 4 in this volume), and an extensive review of plant intraspecific variation found within serpentine eco- systems worldwide. We conclude by summarizing major trends in plant adapta- tion to serpentine soils as demonstrated by examples of intraspecific variation.

Saline and Selenium-Enriched Habitats Soils derived from marine sedimentary rocks in arid or semiarid environments are often laden with sodium chloride (NaCl) and selenium (Se), rendering them stressful or toxic to plant establishment and growth (Läuchli and Lüttge, 2002). Some species exhibit ecotypic differentiation, with saline-tolerant ecotypes dis- playing distinct physiological adaptations, such as leaf vacuolar NaCl accumula- tion, to cope with high soil NaCl concentrations (Rozema et al., 1978; Cheeseman, 1988; Staal et al., 1991). Other plant species contain Se-tolerant ecotypes that have adapted to tolerate high concentrations of bioavailable Se in soil (Enberg and Wu, 1995; Wu et al., 1997). In addition to tolerance, some species of Astragalus (Fa- baceae) have evolved selective root uptake and shoot translocation mechanisms to hyperaccumulate Se (Sors et al., 2005). Se hyperaccumulation has been demon- strated to deter herbivory by insects and rodents (Hanson et al., 2004; Freeman et al., 2007, 2009; Quinn et al., 2008).

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METALLIFEROUS Mine Tailings Intraspecific variation has led to ecotypic differentiation of plant species growing on metalliferous mine tailings. Metalliferous mine tailings typically have low water- holding capacity, nutrients, and pH and contain high concentrations of bioavail- able heavy metals, including aluminum (Al), arsenic (As), cadmium (Cd), cobalt (Co), chromium (Cr), copper (Cu), iron (Fe), manganese (Mn), nickel (Ni), lead (Pb), and/or zinc (Zn), depending on the host rock and ore type. These conditions make mine tailings a hostile environment for plant establishment and growth. Due to their widely dispersed, discontinuous distribution and stressful edaphic condi- tions that drastically differ from the surrounding environment, mines may in ef- fect be regarded as isolated island environments. Ecotypic differentiation along mine tailings has been demonstrated in numerous plant species across diverse families. Some species exhibit metal tolerance at low levels within populations not grow- ing on mine tailings, indicating preadaptation (Macnair and Gardner, 1998). Mine ecotypes of some species also exhibit early flowering compared with adjacent nor- mal soil ecotypes, thereby reducing gene flow between the populations (McNeilly and Antonovics, 1968; Macnair and Gardner, 1998; Antonovics, 2006). Physiolog- ical adaptations identified for metal-tolerant ecotypes include metal exclusion or sequestration at the root or shoot level (Smith and Bradshaw, 1979; Macnair, 1983; Baker, 1987; Hall, 2002). Ecotypes of some species exhibit heavy metal hyperac- cumulation (He et al., 2002; Yang et al., 2006), and some metal-hyperaccumulating species show ecotypic differentiation with respect to metal uptake rates (Zhao et al., 2002; Richau and Schat, 2009). Metal-tolerant ecotypes of some species pre- sumably originated independently multiple times via parallel evolution and con- vergence (Nordal et al., 1999). Extensive studies have been conducted on Mimulus guttatus (Phrymaceae; for- merly Scrophulariaceae) found on 150-year-old Cu mines in California (Macnair and Gardner, 1998). These studies have demonstrated ecotypic differentiation with respect to Cu, including the evolution of a new Cu-tolerant species, M. cupriphilus, from the Cu-tolerant ecotypes of M. guttatus (Macnair, 1989a, 1989b; Macnair and Cumbes, 1989; Macnair et al., 1989). The studies of the M. guttatus complex demonstrate the capacity for rapid evolution of both ecotypes and species within a relatively short time span when a species is subjected to the extreme directional selective pressures of an edaphically stressful environment.

Serpentine Soil Properties and Plant Tolerance Mechanisms Serpentine ecosystems bear many similarities to mine tailings and other edaphi- cally unusual soil types. Serpentine soils are often shallow and rocky; frequently

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms 100 serpentine as a model in evolution deficient in nitrogen (N), phosphorus (P), potassium (K), and calcium (Ca); con- tain toxic concentrations of bioavailable magnesium (Mg) and heavy metals in- cluding Ni, Cr, and Co; and exhibit low water-holding capacity (Brooks, 1987; Roberts and Proctor, 1992; Alexander et al., 2007). Exceptionally low Ca:Mg molar ratios (typically ,,1.0) are a key characteristic of serpentine soils. The properties of serpentine soils are inherited from Fe-Mg-rich (ferromagnesian silicates) par- ent materials, which may also contain Ni-, Cr-, and Co-bearing accessory miner- als (Burt et al., 2001). Plant adaptations to serpentine soils are varied and include those that convey tolerance to drought stress, macronutrient deficiency (N, P, K, S, and Ca), macro- nutrient toxicity (Mg), and heavy metal toxicity (Ni, Cr, Co). Typical drought stress adaptations include reduced leaf size, sclerophylly, succulence, and high root:shoot biomass ratio (Kruckeberg, 1984; Tibbetts and Smith, 1992; Brady et al., 2005). Elevated concentrations of cations associated with osmotic adjustment, including K1, Na1, and less frequently, Ni21 as found in some hyperaccumulators, are also associated with drought tolerance (Boyd and Martens, 1992; Bhatia et al., 2005; Kazakou et al., 2008). Inherently slow growth rates and high root:shoot biomass ratios are common strategies to cope with nutrient deficiency (Brady et al., 2005; Kazakou et al., 2008). Enhanced physiological regulation of nutrient uptake, particularly with respect to Ca and Mg, is also a common serpentine tolerance feature (Brady et al., 2005; Kazakou et al., 2008). The exceedingly low Ca:Mg ratios typical of serpentine soils are antagonistic to plant Ca uptake, requiring specialized physiological mecha- nisms to maintain adequate internal concentrations of Ca. In dicotyledonous an- giosperms, a high proportion of the plant’s total Ca is located in the cell wall, bound to pectins at the middle lamella (Demarty et al., 1984; Marschner, 2002). Pectins play an important role in plant cell wall stability and cell-to-cell adhesion (Jarvis, 1984; Jarvis et al., 2003). Typical symptoms of Ca deficiency are cessation of cell wall extension, disintegration of cell walls, and tissue collapse (necrosis) due to pectin breakdown (Demarty et al., 1984; Jarvis, 1984; Marschner, 2002). These symptoms are most apparent in actively growing tissues, such as root tips, new shoots, and leaves, because Ca is a phloem-immobile element. Physiological toler- ance mechanisms to cope with the low Ca:Mg ratio of serpentine soils, including selective Ca uptake and translocation, Mg exclusion, and Mg sequestration, have been identified in diverse serpentine plant species and ecotypes globally (Krucke­ berg, 1950; Brady et al., 2005; Kazakou et al., 2008). Heavy metal tolerance mechanisms identified in serpentine-tolerant plant spe- cies include metal exclusion at the root level, sequestration to various plant or- gans, and toxicity tolerance (Baker, 1987; Shaw, 1990). Some plant species and ecotypes hyperaccumulate heavy metals to exceptionally high concentrations ( 1000 µg/g; Reeves, 1992; Brooks, 1998). Heavy metal hyperaccumulation has

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 101 been demonstrated to reduce insect herbivory in several plant species and eco- types (Brooks, 1998; Boyd, 2007) and also serve in osmotic regulation and drought tolerance (Boyd and Martens, 1992; Bhatia et al., 2005; Kazakou et al., 2008).

Edaphic Ecotype Differentiation and Modes of Speciation Acquisition of environmental stress tolerance and subsequent ecotypic differentia- tion has been proposed as the first steps toward speciation (Figure 5.1; Mason, 1946a, 1946b; Lewis, 1962; Kruckeberg, 1986; Macnair and Gardner, 1998; Raja- karuna, 2004; see Chapter 4 in this volume). In step 1, individuals of the species acquire edaphic tolerance (preadaptive trait) through random genetic mutation that confers serpentine tolerance within a nonserpentine population. In step 2, disruptive selection, catastrophic selection, or gradual divergence separates the species into distinct serpentine-tolerant and serpentine-intolerant ecotypes. In step 3, further genetic divergence in structural and functional traits occurs within the serpentine-tolerant ecotype. At step 4, isolation between serpentine-tolerant

figureFigure 5.1. No: Hypothesized 5.1; Date: speciation 6/3/10; pathway. Pica Redrawnwidth: from24; PeKruckebergrcentage (1986). of original: 91.27

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms 102 serpentine as a model in evolution and serpentine-intolerant ecotypes of the species becomes genetically fixed. Fi- nally, at step 5, further divergence of the serpentine-tolerant ecotype leads to a distinct species. The recurrent process of independent ecotype differentiation for the same adaptations to the same environmental factors within the same species is referred to as parallel evolution (Levin, 2001). Four speciation modes are generally recognized, including parapatric, allopat- ric, peripatric, and sympatric (Figure 5.2; Briggs and Walters, 1997). Parapatric, peripatric, and sympatric speciation are the most logical modes of ecotypic dif- ferentiation in the case of plant species that are not strict serpentine endemics and have populations that may exist on both serpentine as well as adjacent or nearby nonserpentine soils. Allopatric speciation is the most logical mode of differentia- tion in the case of strict serpentine endemic species that are entirely restricted to serpentine (Kruckeberg, 1984). Similar to mines and other edaphically unusual substrates, serpentine soils have a discontinuous, widespread distribution across the landscape and may be regarded as island environments with geographic separation acting as a barrier to gene flow (Figure 5.3). As isolated environments, they are subject to the selective processes that occur with respect to island biogeography (Kruckeberg, 1991). Ex- tension of the concepts of island biogeography to serpentine habitats may be lim- ited because unlike oceanic islands, mainland edaphic islands are surrounded by

figureFigure 5.2. ModesNo: 5.2; of ecotypic Date: differentiation6/3/10; Pica and width: speciation. 24; Pe*designatesrcentage unique of trait ac- quiredoriginal: by ecotype 93.16 and subsequently the resulting distinct species.

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figureFigure 5.3. No:Serpentine 5.3; Date: distribution 6/3/10; (black Pica areas) width: in the northern 27; Pe Sierrarcentage Nevada of moun- tainoriginal: range, California. 84.97 Note the discontinuous, widespread, island-like distribution. This type of discontinuous distribution is typical of most serpentine areas around the world (Brooks, 1987). habitats occupied by potential recruits. On mainland edaphic islands, recruit pre- adaptation to edaphic stress restricts successful colonization. In contrast, recruit dispersal distance is the primary factor that limits successful colonization on oceanic islands. As a result, extension of island biogeography concepts to edaphic islands is most suitable for examining the influence of spatial isolation on

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms 104 serpentine as a model in evolution population genetic attributes, such as reduced gene flow, inbreeding, and genetic drift (Mayer et al., 1994).

Intraspecific Variation: Plant Taxa Used in Studies of Plant Adaptation and Evolution to Serpentine Soils Formal studies of intraspecific variation involving serpentine soils have largely been limited to a small number of genera within a limited number of families. Most plant taxa used are found in the following families: Asteraceae, Brassicaceae, Cary- ophyllaceae, Cupressaceae, Hydrophyllaceae, Lamiaceae, Phrymaceae, Pinaceae, Plantaginaceae, Poaceae, Polemoniaceae, and Rosaceae. Serpentine tolerance ad- aptations vary by species, with some ecotypes having several adaptive traits. A summary of adaptations by species is provided in Table 5.1. Adaptation patterns have emerged within several families. To highlight those patterns, each major plant taxon used for intraspecific variation studies is discussed in turn by plant family.

Asteraceae: Drought Tolerance, Ca and Mg Regulation, Parallel and Convergent Evolution Plant species with widespread distribution on both serpentine and nonserpentine soil types have provided ideal models to study evolutionary concepts related to ecotypic differentiation, as well as the underlying adaptations in ecotypes that con- vey tolerance to serpentine soils. For example, Achillea millefolium, an obligately outcrossing, insect-pollinated species, has a widespread Northern Hemisphere distribution and occurs across a wide range of climates and soil types, including serpentine (Löfgren, 2002). Reciprocal and unilateral transplant studies of A. millefolium serpentine and nonserpentine soil populations in California and Washington revealed that the species has distinct serpentine-tolerant ecotypes (Figure 5.4; Kruckeberg, 1950, 1967; McMillan, 1956; Higgins and Mack, 1987; Cooke, 1994). Identified adaptations include reduced growth rate, nutrient defi- ciency tolerance, and tolerance of low Ca:Mg ratios and elevated Ni. Curiously, some nonserpentine ecotypes were fully or partially serpentine-tolerant, suggest- ing preadaptation (Kruckeberg, 1950). Unilateral transplant glasshouse studies (O’Dell and Claassen, 2008) conducted with a large number of serpentine and nonserpentine ecotypes from North Amer- ica (eastern and western United States and Canada) and Europe (United Kingdom and Austria) revealed the same trend as found by Kruckeberg (1950) on a much broader spatial scale. When grown on serpentine, serpentine ecotype seedlings always exhibited healthy radicle growth, whereas the radicles of nonserpentine

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 105 ecotype seedlings typically became necrotic (O’Dell and Claassen, 2006, 2008). A comparison of nutrient uptake between serpentine and nonserpentine ecotypes revealed that nearly all serpentine-tolerant ecotypes displayed selective Ca uptake at the root level and translocation to the shoot, resulting in a significantly higher shoot Ca:Mg ratio than found in nonserpentine ecotypes, which lack this mecha- nism (O’Dell and Claassen, 2006, 2008). This tolerance mechanism was nearly ubiquitous for serpentine ecotypes of both North America and Europe, suggesting that it has originated multiple times in distant serpentine ecotypes via parallel evolution and convergence (O’Dell and Claassen, 2008). Parallel evolution has also been found for races of the Lasthenia californica complex. L. californica is an obligately outcrossing, insect-pollinated species. It has a widespread distribution in California grasslands, occurring on both serpentine and nonserpentine soils. Two distinct races (races A and C) of this complex have been identified (Desrochers and Bohm, 1995; Rajakaruna and Bohm, 1999). Race A occupies ionically extreme environments (coastal bluffs, alkaline flats, and ser- pentine), whereas race C occurs mostly on more ionically moderate and drier sites (pastures and oak woodland). Although race A is tolerant of the nonserpentine soils that support race C, race C is generally not found on extreme serpentine soils that support race A. Race A has significantly greater Ca uptake and Na and Mg tolerance compared to race C, as measured by root growth, survival, and nutrient uptake (Rajakaruna et al., 2003b). Elevated Na uptake and root-to-shoot translocation in race A is be- lieved to confer greater tolerance of soil ionic stress owing to greater capacity for osmotic adjustment. Elevated Ca uptake and root-to-shoot translocation in race A allows these plants to maintain a favorable shoot tissue Ca:Mg ratio in serpentine soils. The races appear to have evolved barriers to reproduction as well, especially via flowering time differences in their parapatric occurrences in northern Califor- nia (Rajakaruna and Bohm, 1999). Phylogenetic analysis has revealed that the races occur in parallel in two geographically based clades within the complex, sug- gesting the parallel evolution of one or both races in response to contrasting soil conditions under which these races are found (Rajakaruna et al., 2003a).

Phrymaceae and Plantaginaceae: Drought Tolerance, Flower Modification, Early Flowering, Ca and Mg Regulation, Ni Tolerance Genus Mimulus (Phrymaceae; formerly Scrophulariaceae) has emerged as an im- portant model in plant evolution studies (Wu et al., 2008). Mimulus guttatus is insect-pollinated with high variation of pollination mode in individual plants, ranging from fully outcrossing to fully selfing (Gardner and Macnair, 2000). The

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Serpentine Tolerance Adaptations

Flower Modifcation Ca Regulation Mg Regulation Ni Regulation

Higher Comparison Differentiation Reduced Shoot Translocation Comparison Serpentine Supported by Drought Growth Reduced Earlier Selective Translocation Root Level Ca: Mg Rool Level to Shoot Species Family Populations Tolerance Phyogenetics Tolerance Rate Leaf Size Flowering Shape Color Uplake to Shoot Tolerance ExclusionMolar Tolerance Exclusion Sequestration (accumulation) References

Achillea millefolium Asteraceae S and NS S > NS X Xb Xb X Xb X 1–5 Lasthenia califonica Asteraceae S and NS S > NS Xb X X Xb Xb Xb Xb 6–9 Helianthus exilisa Asteraceae S only . . . X X X 10, 11 Erigeron thunbergii Asteraceae S and NS No data Xb 12 Erigeron compositus Asteraceae S and NS S > NS 13 Senecio paupercaulus Asteraceae S and NS S > NS 13 Senecio coronatus Asteraceae S and NS S > NS X 14–16 Berkheya zeyeri Asteraceae S and NS No data Xb X 17 subsp. rehmannii Fragaria virginiana Rosaceae S and NS S > NS 13 Potentilla glandulosa Rosaceae S and NS S > NS 13 Spirea douglasii Rosaceae S and NS S > NS 13 Mimulus guttatus Phrymaceae S and NS S > NS X X X X X X X X 18–22 Mimulus nudatusa Phrymaceae S only . . . X X 23 Collinsia sparsifolia Plantaginaceae S and NS S > NS X X 24, 25 Plantago erecta Plantaginaceae S and NS S > NS 1, 26 Salvia columbariae Lamiaceae S and NS S > NS X X 1 Prunella vulgaris Lamiaceae S and NS S > NS 13 Teucrium Lamiaceae S and NS No data X 27 chamaedrys Silene dioica Caryophyllaceae S andNS S = NS Xb 28–30 Silene vulgaris Caryophyllaceae S and NS S > NS X X X 31, 32 Silene armeria Caryophyllaceae S and NS S > NS 33 Silene paradoxa Caryophyllaceae S and NS S > NS X X 34 Silene italica Caryophyllaceae S and NS S > NS X X 35 Cerastium alpinum Caryophyllaceae S and NS S > NS Xb X X 36–38 Arenaria katoanaa Caryophyllaceae S only . . . X 39 Gilia capitata Polemoniaceae S and NS S > NS X X X 1 Navarretia squarrosa Polemoniaceae S and NS S > NS 40 Navarretia mellita Polemoniaceae S and NS S > NS 40 Navarretia rosulataa Polemoniaceae S only . . . Xb 41 Navarretia jepsoniia Polemoniaceae S only . . . Xb 41 Navarretia jarediia Polemoniaceae S only . . . Xb 41 Leptosiphon Polemoniaceae S and NS S > NS X X X X X 42–44 androsaceus Leptosiphon Polemoniaceae S and NS S > NS X X X 45 parviflorus Leptosiphon bicolor Polemoniaceae S and NS No data X 44 Phacelia californica Hydrophyllaceae S and NS S > NS X X X 1 var. Phacelia dubia Hydrophyllaceae NS only Xc Xc 46 georgianac Agropyron spicatum Poaceae S and NS S > NS X X X X X 47 Festuca rubra Poaceae S and NS S = NS 48, 49 Agrostis stolonifera Poaceae S and NS S = NS 48, 49 Agrostis canina Poaceae S and NS S = NS 48, 49 Agrostis hallii Poaceae S and NS S = NS 40

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Serpentine Tolerance Adaptations

Flower Modifcation Ca Regulation Mg Regulation Ni Regulation

Higher Comparison Differentiation Reduced Shoot Translocation Comparison Serpentine Supported by Drought Growth Reduced Earlier Selective Translocation Root Level Ca: Mg Rool Level to Shoot Species Family Populations Tolerance Phyogenetics Tolerance Rate Leaf Size Flowering Shape Color Uplake to Shoot Tolerance ExclusionMolar Tolerance Exclusion Sequestration (accumulation) References

Achillea millefolium Asteraceae S and NS S > NS X Xb Xb X Xb X 1–5 Lasthenia califonica Asteraceae S and NS S > NS Xb X X Xb Xb Xb Xb 6–9 Helianthus exilisa Asteraceae S only . . . X X X 10, 11 Erigeron thunbergii Asteraceae S and NS No data Xb 12 Erigeron compositus Asteraceae S and NS S > NS 13 Senecio paupercaulus Asteraceae S and NS S > NS 13 Senecio coronatus Asteraceae S and NS S > NS X 14–16 Berkheya zeyeri Asteraceae S and NS No data Xb X 17 subsp. rehmannii Fragaria virginiana Rosaceae S and NS S > NS 13 Potentilla glandulosa Rosaceae S and NS S > NS 13 Spirea douglasii Rosaceae S and NS S > NS 13 Mimulus guttatus Phrymaceae S and NS S > NS X X X X X X X X 18–22 Mimulus nudatusa Phrymaceae S only . . . X X 23 Collinsia sparsifolia Plantaginaceae S and NS S > NS X X 24, 25 Plantago erecta Plantaginaceae S and NS S > NS 1, 26 Salvia columbariae Lamiaceae S and NS S > NS X X 1 Prunella vulgaris Lamiaceae S and NS S > NS 13 Teucrium Lamiaceae S and NS No data X 27 chamaedrys Silene dioica Caryophyllaceae S andNS S = NS Xb 28–30 Silene vulgaris Caryophyllaceae S and NS S > NS X X X 31, 32 Silene armeria Caryophyllaceae S and NS S > NS 33 Silene paradoxa Caryophyllaceae S and NS S > NS X X 34 Silene italica Caryophyllaceae S and NS S > NS X X 35 Cerastium alpinum Caryophyllaceae S and NS S > NS Xb X X 36–38 Arenaria katoanaa Caryophyllaceae S only . . . X 39 Gilia capitata Polemoniaceae S and NS S > NS X X X 1 Navarretia squarrosa Polemoniaceae S and NS S > NS 40 Navarretia mellita Polemoniaceae S and NS S > NS 40 Navarretia rosulataa Polemoniaceae S only . . . Xb 41 Navarretia jepsoniia Polemoniaceae S only . . . Xb 41 Navarretia jarediia Polemoniaceae S only . . . Xb 41 Leptosiphon Polemoniaceae S and NS S > NS X X X X X 42–44 androsaceus Leptosiphon Polemoniaceae S and NS S > NS X X X 45 parviflorus Leptosiphon bicolor Polemoniaceae S and NS No data X 44 Phacelia californica Hydrophyllaceae S and NS S > NS X X X 1 var. Phacelia dubia Hydrophyllaceae NS only Xc Xc 46 georgianac Agropyron spicatum Poaceae S and NS S > NS X X X X X 47 Festuca rubra Poaceae S and NS S = NS 48, 49 Agrostis stolonifera Poaceae S and NS S = NS 48, 49 Agrostis canina Poaceae S and NS S = NS 48, 49 Agrostis hallii Poaceae S and NS S = NS 40

(continued)

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms table 5.1 (continued)

Serpentine Tolerance Adaptations

Flower Modifcation Ca Regulation Mg Regulation Ni Regulation

Higher Comparison Differentiation Reduced Shoot Translocation Comparison Serpentine Supported by Drought Growth Reduced Earlier Selective Translocation Root Level Ca: Mg Rool Level to Shoot Species Family Populations Tolerance Phyogenetics Tolerance Rate Leaf Size Flowering Shape Color Uplake to Shoot Tolerance ExclusionMolar Tolerance Exclusion Sequestration (accumulation) References Avena fatua Poaceae S and NS S > NS X 50 Bromus madritensis Poaceae S and NS S = NS 1 Bromus tectorum Poaceae S and NS S = NS 13 Bromus hordeaceus Poaceae S and NS S > NS X X 1, 40, 51 Bromus diandrus Poaceae S and NS S = NS 1 Bromus laevipes Poaceae S and NS S = NS 1 Elymus multisetus Poaceae S and NS S = NS 1 Vulpia microsatchys Poaceae S and NS S > NS X 52 Aegilops triuncialis Poaceae S and NS S = NS 53,54 Pinus ponderosa Pinaceae S and NS S > NS X X X 55, 56 Pinus contorta Pinaceae S and NS S > NS 13 Pinus sabiniana Pinaceae S and NS S = NS 57 Pinus attenuata Pinaceae S and NS S = NS 40 Pinus virginiana Pinaceae S and NS S = NS 58 Pinus jeffreyii Pinaceae S and NS No data X 59 Pinus balfouriana Pinaceae S and NS No data X 60 subsp. balfouriana Picea glehnii Pinaceae NS only NS 61 Picea jezoensis Pinaceae NS only NS 61 Hesperocyparis Cupressaceae S and NS S = NS 40 macnabiana Juniperus communis Cupressaceae S and NS S = NS 13, 62, 63 Arabidopsis thaliana Brassicaceae NS only CAX mutantc Xc Xc 64 Arabidopsis lyrata Brassicaceae S and NS S > NS Xb X Xb X X Xb 65–68 Erysimum capitatum Brassicaceae S and NS S > NS 1 Streptanthus Brassicaceae S only . . . X X X X X (variation) 69–71 polygaloidesa Streptanthus Brassicaceae S and NS S > NS Xb X X 1, 72–75 glandulosus Caulanthus Brassicaceae S and NS S > NS X X X 76 amplexicaulis Noccaea fendlerii Xd (both Brassicaceae S and NS S = NSd 77–79 subsp. glauca S and NS) Noccaea fendlerii subsp. Brassicaceae S only . . . X 77 siskiyouensea Noccaea fendlerii subsp. Brassicaceae S only . . . X 77 californicaa Xd (both Noccaea goesingense Brassicaceae S and NS S = NSd 80 S and NS) Noccaea Xd (both Brassicaceae S and NS S = NSd 81, 82 cochleariforme S and NS) Noccaea caerulescens Brassicaceae S and NS No data X (S > NS) 83, 84

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms table 5.1 (continued)

Serpentine Tolerance Adaptations

Flower Modifcation Ca Regulation Mg Regulation Ni Regulation

Higher Comparison Differentiation Reduced Shoot Translocation Comparison Serpentine Supported by Drought Growth Reduced Earlier Selective Translocation Root Level Ca: Mg Rool Level to Shoot Species Family Populations Tolerance Phyogenetics Tolerance Rate Leaf Size Flowering Shape Color Uplake to Shoot Tolerance ExclusionMolar Tolerance Exclusion Sequestration (accumulation) References Avena fatua Poaceae S and NS S > NS X 50 Bromus madritensis Poaceae S and NS S = NS 1 Bromus tectorum Poaceae S and NS S = NS 13 Bromus hordeaceus Poaceae S and NS S > NS X X 1, 40, 51 Bromus diandrus Poaceae S and NS S = NS 1 Bromus laevipes Poaceae S and NS S = NS 1 Elymus multisetus Poaceae S and NS S = NS 1 Vulpia microsatchys Poaceae S and NS S > NS X 52 Aegilops triuncialis Poaceae S and NS S = NS 53,54 Pinus ponderosa Pinaceae S and NS S > NS X X X 55, 56 Pinus contorta Pinaceae S and NS S > NS 13 Pinus sabiniana Pinaceae S and NS S = NS 57 Pinus attenuata Pinaceae S and NS S = NS 40 Pinus virginiana Pinaceae S and NS S = NS 58 Pinus jeffreyii Pinaceae S and NS No data X 59 Pinus balfouriana Pinaceae S and NS No data X 60 subsp. balfouriana Picea glehnii Pinaceae NS only NS 61 Picea jezoensis Pinaceae NS only NS 61 Hesperocyparis Cupressaceae S and NS S = NS 40 macnabiana Juniperus communis Cupressaceae S and NS S = NS 13, 62, 63 Arabidopsis thaliana Brassicaceae NS only CAX mutantc Xc Xc 64 Arabidopsis lyrata Brassicaceae S and NS S > NS Xb X Xb X X Xb 65–68 Erysimum capitatum Brassicaceae S and NS S > NS 1 Streptanthus Brassicaceae S only . . . X X X X X (variation) 69–71 polygaloidesa Streptanthus Brassicaceae S and NS S > NS Xb X X 1, 72–75 glandulosus Caulanthus Brassicaceae S and NS S > NS X X X 76 amplexicaulis Noccaea fendlerii Xd (both Brassicaceae S and NS S = NSd 77–79 subsp. glauca S and NS) Noccaea fendlerii subsp. Brassicaceae S only . . . X 77 siskiyouensea Noccaea fendlerii subsp. Brassicaceae S only . . . X 77 californicaa Xd (both Noccaea goesingense Brassicaceae S and NS S = NSd 80 S and NS) Noccaea Xd (both Brassicaceae S and NS S = NSd 81, 82 cochleariforme S and NS) Noccaea caerulescens Brassicaceae S and NS No data X (S > NS) 83, 84

(continued)

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms table 5.1 (continued)

Serpentine Tolerance Adaptations

Flower Modifcation Ca Regulation Mg Regulation Ni Regulation

Higher Comparison Differentiation Reduced Shoot Translocation Comparison Serpentine Supported by Drought Growth Reduced Earlier Selective Translocation Root Level Ca: Mg Rool Level to Shoot Species Family Populations Tolerance Phyogenetics Tolerance Rate Leaf Size Flowering Shape Color Uplake to Shoot Tolerance ExclusionMolar Tolerance Exclusion Sequestration (accumulation) References Noccaea pindicaa Brassicaceae S only . . . X (S > other S) 85 Cochlearia pyrenaica Brassicaceae S and NS No data X 86 Alyssum murale Brassicaceae Ecotypes S > NS X 87 Alyssum bertoloniia Brassicaceae Ecotypes . . . X (S > other S) 88 Alyssum inflatum Brassicaceae Ecotypes S > NS X 89 references: 1: Kruckeberg, 1950; 2: Higgins and Mack, 1987; 3: Cooke, 1994; 4: O’Dell and Claassen, 2006; 5: O’Dell and Claassen, 2008; 6: Desro- chers and Bohm, 1995; 7: Rajakaruna and Bohm, 1999; 8: Rajakaruna et al., 2003a; 9: Rajakaruna et al., 2003b; 10: Sambatti and Rice, 2006; 11: Sam- batti and Rice, 2007; 12: Kawase et al., 2007; 13: Kruckeberg, 1967; 14: Morrey et al., 1992; 15: Mesjasz-Przybyłowicz et al., 2007; 16: Boyd et al., 2008; 17: Williamson et al., 1997; 18: Tilstone and Macnair, 1997; 19: Hughes et al., 2001; 20: Murren et al., 2006; 21: Hendrick, 2008; 22: Selby et al., 2008; 23: Macnair, 1992; 24: Wright et al., 2006; 25: Wright and Stanton, 2007; 26: Espeland and Rice, 2007; 27: Pavlova, 2009; 28: Westerbergh and Saura, 1992; 29: Westerbergh and Saura, 1994; 30: Westerbergh, 1994; 31: Bratteler et al., 2006a; 32: Bratteler et al., 2006b; 33: Lombini et al., 2003; 34: Gon- nelli et al., 2001; 35: Gabbrielli et al., 1990; 36: Nyberg Berglund and Westerbergh, 2001a; 37: Nyberg Berglund and Westerbergh, 2001b; 38: Nyberg Berglund et al., 2003; 39: Kawase et al., 2009; 40: McMillan, 1956; 41: Spencer and Porter, 1997; 42: Proctor and Woodell, 1975; 43: Woodell et al., 1975; 44: Schmitt, 1983; 45: Chapter 4 in this volume; 46: Taylor and Levy, 2002; 47: Main, 1974; 48: Proctor, 1971; 49: Johnston and Proctor, 1981; 50: Har- rison et al., 2001; 51: Freitas and Mooney, 1996; 52: Jurjavcic et al., 2002; 53: Thomson, 2007; 54: Lyons et al., in press; 55: Jenkinson, 1974; 56: Wright, 2007; 57: Griffin, 1962; 58: Miller and Cumming, 2000; 59: Fur nier and Adams, 1986; 60: Oline et al., 2000; 61: Kayama et al., 2005; 62: Ashworth et al., 2001; 63: Adams and Nyugen, 2007; 64: Bradshaw, 2005; 65: Roche et al., 2004; 66: Von Wettberg, 2008; 67: Turner et al., 2008; 68: Turner et al., 2010; 69: Reeves et al., 1981; 70: Rodman et al., 1981; 71: Boyd et al., 2009; 72: Kruckeberg, 1957; 73: Mayer et al., 1994; 74: Mayer and Soltis, 1994; 75: Mayer and Soltis, 1999; 76: Pepper and Norwood, 2001; 77: Reeves et al., 1983; 78: Boyd and Martens, 1998; 79: Peer et al., 2006; 80: Reeves and Baker, 1984; 81: Mizuno et al., 2003; 82: Mizuno et al., 2005; 83: Assunção et al., 2003; 84: Assunção et al., 2008; 85: Taylor and Macnair, 2006; 86: Nagy and Proctor, 2001; 87: Asemaneh et al., 2006; 88: Galardi et al., 2007; 89: Ghasemi and Gahderian, 2009. a Strict serpentine endemic species. b Evidence of parallel and convergent evolution within species. c Example of preadaptation. d Constitutive trait.

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms table 5.1 (continued)

Serpentine Tolerance Adaptations

Flower Modifcation Ca Regulation Mg Regulation Ni Regulation

Higher Comparison Differentiation Reduced Shoot Translocation Comparison Serpentine Supported by Drought Growth Reduced Earlier Selective Translocation Root Level Ca: Mg Rool Level to Shoot Species Family Populations Tolerance Phyogenetics Tolerance Rate Leaf Size Flowering Shape Color Uplake to Shoot Tolerance ExclusionMolar Tolerance Exclusion Sequestration (accumulation) References Noccaea pindicaa Brassicaceae S only . . . X (S > other S) 85 Cochlearia pyrenaica Brassicaceae S and NS No data X 86 Alyssum murale Brassicaceae Ecotypes S > NS X 87 Alyssum bertoloniia Brassicaceae Ecotypes . . . X (S > other S) 88 Alyssum inflatum Brassicaceae Ecotypes S > NS X 89 references: 1: Kruckeberg, 1950; 2: Higgins and Mack, 1987; 3: Cooke, 1994; 4: O’Dell and Claassen, 2006; 5: O’Dell and Claassen, 2008; 6: Desro- chers and Bohm, 1995; 7: Rajakaruna and Bohm, 1999; 8: Rajakaruna et al., 2003a; 9: Rajakaruna et al., 2003b; 10: Sambatti and Rice, 2006; 11: Sam- batti and Rice, 2007; 12: Kawase et al., 2007; 13: Kruckeberg, 1967; 14: Morrey et al., 1992; 15: Mesjasz-Przybyłowicz et al., 2007; 16: Boyd et al., 2008; 17: Williamson et al., 1997; 18: Tilstone and Macnair, 1997; 19: Hughes et al., 2001; 20: Murren et al., 2006; 21: Hendrick, 2008; 22: Selby et al., 2008; 23: Macnair, 1992; 24: Wright et al., 2006; 25: Wright and Stanton, 2007; 26: Espeland and Rice, 2007; 27: Pavlova, 2009; 28: Westerbergh and Saura, 1992; 29: Westerbergh and Saura, 1994; 30: Westerbergh, 1994; 31: Bratteler et al., 2006a; 32: Bratteler et al., 2006b; 33: Lombini et al., 2003; 34: Gon- nelli et al., 2001; 35: Gabbrielli et al., 1990; 36: Nyberg Berglund and Westerbergh, 2001a; 37: Nyberg Berglund and Westerbergh, 2001b; 38: Nyberg Berglund et al., 2003; 39: Kawase et al., 2009; 40: McMillan, 1956; 41: Spencer and Porter, 1997; 42: Proctor and Woodell, 1975; 43: Woodell et al., 1975; 44: Schmitt, 1983; 45: Chapter 4 in this volume; 46: Taylor and Levy, 2002; 47: Main, 1974; 48: Proctor, 1971; 49: Johnston and Proctor, 1981; 50: Har- rison et al., 2001; 51: Freitas and Mooney, 1996; 52: Jurjavcic et al., 2002; 53: Thomson, 2007; 54: Lyons et al., in press; 55: Jenkinson, 1974; 56: Wright, 2007; 57: Griffin, 1962; 58: Miller and Cumming, 2000; 59: Fur nier and Adams, 1986; 60: Oline et al., 2000; 61: Kayama et al., 2005; 62: Ashworth et al., 2001; 63: Adams and Nyugen, 2007; 64: Bradshaw, 2005; 65: Roche et al., 2004; 66: Von Wettberg, 2008; 67: Turner et al., 2008; 68: Turner et al., 2010; 69: Reeves et al., 1981; 70: Rodman et al., 1981; 71: Boyd et al., 2009; 72: Kruckeberg, 1957; 73: Mayer et al., 1994; 74: Mayer and Soltis, 1994; 75: Mayer and Soltis, 1999; 76: Pepper and Norwood, 2001; 77: Reeves et al., 1983; 78: Boyd and Martens, 1998; 79: Peer et al., 2006; 80: Reeves and Baker, 1984; 81: Mizuno et al., 2003; 82: Mizuno et al., 2005; 83: Assunção et al., 2003; 84: Assunção et al., 2008; 85: Taylor and Macnair, 2006; 86: Nagy and Proctor, 2001; 87: Asemaneh et al., 2006; 88: Galardi et al., 2007; 89: Ghasemi and Gahderian, 2009. a Strict serpentine endemic species. b Evidence of parallel and convergent evolution within species. c Example of preadaptation. d Constitutive trait.

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figure 5.4. Response of serpentine (S) and nonserpentine (NS) ecotypes of Achillea millefolium (Asteraceae) grown on serpentine (top) and nonserpentine (bottom) soil (photo credit: Kruckeberg, 1950). Note the poor growth of the NS ecotypes on the serpen- tine soil and normal growth of all ecotypes on the nonserpentine soil. Also note, however, that sporadic individuals of the NS ecotypes appear to have tolerance to the serpentine soil. This is particularly evident for ecotype 206 (fifth row from left) where most if not all individuals of the nonserpentine ecotype appear to be serpentine-tolerant. species has a widespread North American distribution and is found in a variety of climates and on diverse soil types, including serpentine. Reciprocal and unilateral transplant studies of serpentine and nonserpentine populations of M. guttatus in California have revealed that the species has under- gone ecotypic divergence with respect to serpentine tolerance. M. guttatus serpen- tine ecotypes have an earlier flowering time, reduced leaf size and shoot biomass, and higher Ca uptake and root-to-shoot translocation when compared to nonser- pentine ecotypes (Hughes et al., 2001; Murren et al., 2006; Hendrick, 2008; Selby et al., 2008). Additionally, serpentine ecotypes have greater Ni tolerance than non- serpentine ecotypes (Tilstone and Macnair, 1997). M. guttatus is believed to be the progenitor of M. nudatus, a strict serpentine endemic in California (Macnair, 1992; Safford et al., 2005). Although M. nudatus does not appear to differ significantly with respect to Ca and Mg regulation as

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 113 compared to nonserpentine M. guttatus ecotypes, the species does have drought tolerance mechanisms, including significantly earlier flowering and reduced leaf size (Macnair and Gardner, 1998; Gardner and Macnair, 2000). Reciprocal field transplant studies of Collinsia sparsifolia and Plantago erecta (Plantaginaceae; formerly Scrophulariacae) showed that both species display eco- typic differentiation, with serpentine ecotypes displaying greater tolerance of ser- pentine soils than nonserpentine ecotypes (Kruckeberg, 1950; Wright et al., 2006; Espeland and Rice, 2007). C. sparsifolia ecotypes display differentiation of floral traits, including larger flowers with more intense pigmentation in the serpentine ecotype (Wright and Stanton, 2007; Chapter 7 in this volume).

Caryophyllaceae: Ca and Mg Regulation, Ni Tolerance, Parallel and Convergent Evolution Ecotypic differentiation has been studied in several species of Silene in Europe. S. dioica is an insect-pollinated, dioecious, obligately outcrossing species (Wester- bergh and Saura, 1994). In Sweden, serpentine and nonserpentine populations of S. dioica do not significantly differ in either serpentine or Ni tolerance in a glass- house setting (Westerbergh, 1994). Both serpentine and nonserpentine popula- tions were equally tolerant of serpentine soils, and thus serpentine tolerance is regarded as a constitutive trait in the species. Phylogenetic analysis of several dis- tant serpentine and nonserpentine populations revealed that although both popu- lations in northern Sweden were genetically similar, these populations in southern Sweden exhibited genetic differentiation (Westerbergh and Saura, 1992). Although the serpentine and nonserpentine populations do not display phenotypic differen- tiation with respect to serpentine tolerance, genetic differentiation between them suggests that differences between the environments have restricted gene flow be- tween the populations (Westerbergh and Saura, 1994). S. dioica primarily grows on fragmented habitat types, including serpentine and meadows. It is hypothe- sized that differences in the dominant vegetation cover between the serpentine (grassland) and nonserpentine (forest) environments have restricted the move- ment of pollinator guilds of Silene between them (Westerbergh and Saura, 1994). Quantitative trait loci analysis of serpentine and nonserpentine populations of S. vulgaris in Switzerland showed that the populations display ecotypic differentia- tion with respect to Ni tolerance and morphological traits, including leaf size and shoot length (Bratteler et al., 2006a, 2006b). S. armeria exhibits ecotypic differen- tiation with the serpentine ecotype displaying greater tolerance of low substrate Ca:Mg ratios than the nonserpentine ecotype (Lombini et al., 2003). Likewise, S. paradoxa exhibits ecotypic differentiation, with the serpentine ecotype display- ing greater tolerance of Ni (as measured by root growth) than the nonserpentine

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms 114 serpentine as a model in evolution ecotype (Gonnelli et al., 2001). The Ni tolerance mechanism identified in the ser- pentine ecotype was root sequestration. S. italica also appears to use root seques- tration as a Ni tolerance mechanism (Gabbrielli et al., 1990). Unlike S. dioica, Cerastium alpinum from Sweden, Norway, and Finland does display ecotypic differentiation with serpentine ecotypes having greater Ni and Mg tolerance (Nyberg Berglund et al., 2003). Phylogenetic analysis has revealed that serpentine ecotypes are genetically more similar to nonserpentine ecotypes within the same region than other distant serpentine ecotypes, suggesting that like S. dioica, the species has repeatedly evolved serpentine-tolerant ecotypes via paral- lel evolution (Nyberg Berglund, 2005; Nyberg Berglund and Westerbergh, 2001a, 2001b; Nyberg Berglund et al., 2003).

Polemoniaceae: Early Flowering, Ca and Mg Regulation, Parallel and Convergent Evolution Gilia capitata has a widespread distribution on numerous soil types in western North America (Kruckeberg, 1950). Reciprocal transplant glasshouse studies of G. capitata from California have revealed the presence of distinct serpentine and nonserpentine ecotypes (Kruckeberg, 1950). Shoot tissue analysis of serpentine and nonserpentine ecotypes grown on serpentine soil revealed that the serpentine ecotype had significantly lower Mg concentrations than the nonserpentine eco- type (Kruckeberg, 1950). This suggests that the primary tolerance mechanism used by the serpentine ecotype to maintain a balanced shoot Ca:Mg ratio is re- stricted root-to-shoot translocation of Mg. Under natural field conditions,G. capi- tata growing on serpentine soil flowers at least two weeks earlier than plants grow- ing on nonserpentine soil in the same region, alluding to another possible serpentine tolerance trait (Kruckeberg, 1950). Leptosiphon androsaceus, L. parviflorus, and L. bicolor have a limited western U.S. distribution, with populations occurring on both serpentine and nonserpen- tine soil (Woodell et al., 1975; Schmitt, 1983; Chapter 4 in this volume). Serpen- tine populations of L. androsaceus typically have purple flowers, whereas nonser- pentine populations typically bear white flowers (Proctor and Woodell, 1975). Reciprocal field transplant studies of serpentine and nonserpentine populations in southern California showed that the species has undergone ecotypic differentia- tion (Woodell et al., 1975). Serpentine ecotypes grew significantly larger on ser- pentine soil than did nonserpentine ecotypes. The tolerance mechanism identified in the serpentine ecotype was the ability to maintain an adequate shoot Ca:Mg ratio via selective root-level Ca uptake and translocation to the shoot. Like L. androsaceus, L. parviflorus has serpentine and nonserpentine popula- tions that differ in flower color (Chapter 4 in this volume). Serpentine populations

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 115 in northern California typically bear pink flowers, whereas adjacent nonserpen- tine populations bear white flowers. Reciprocal transplant studies involving ser- pentine and nonserpentine populations in the glasshouse demonstrated ecotypic differentiation with the nonserpentine populations being almost completely intol- erant of serpentine soil. Genetic analysis of these populations revealed that flower color, flowering phenology, plant size, and serpentine tolerance were strongly in- fluenced by soil type and that the trait differences may be reinforced by differing flowering time and restricted gene flow between the two populations. Studies of pollen dispersal between serpentine and nonserpentine ecotypes of L. bicolor in northern California have showed that the serpentine ecotype flowers two weeks earlier than the nonserpentine ecotype (Schmitt, 1983). L. bicolor is self-compatible, facultatively autogamous, and primarily pollinated by beeflies (Bombylliidae). Flowering time difference between the two populations had im- portant consequences for the availability of pollinators and outcrossing between the populations. The early flowering serpentine population received significantly fewer visits from beeflies (44%) compared with the later flowering nonserpentine population (95%) since the serpentine population’s flowering period was largely outside of the beeflies’ activity period. As a result, the proportion of flowers avail- able to outcross with the other population was only 7% for the serpentine popula- tion and 88% for the nonserpentine population. The study suggests that because serpentine soils tend to dry faster than nonserpentine soils, early flowering of the serpentine ecotype may have been selected for, at the expense of outcrossing op- portunity, in a response to mortality from drought (Schmitt, 1983). Like L. bicolor, L. androsaceus serpentine ecotypes have also been noted to flower earlier than nonserpentine ecotypes (Schmitt, 1983).

Hydrophyllaceae: Ca and Mg Regulation, Ni Tolerance Genus Phacelia occurs throughout North and South America and consists of her- baceous annual or perennial species. Reciprocal glasshouse transplant studies of serpentine and nonserpentine populations of P. californica from California re- vealed ecotypic differentiation (Kruckeberg, 1950). Serpentine ecotypes main- tained a significantly higher shoot Ca:Mg concentration than nonserpentine eco- types, primarily by selective Ca uptake and translocation to the shoot. P. dubia is found throughout the eastern United States and includes three allo- patric edaphic endemic varieties: var. georgiana, var. interior, and var. dubia. P. dubia var. georgiana grows on shallow granite soils, var. interior grows on shallow limestone soils, and var. dubia grows on relatively fertile woodland soils (Taylor and Levy, 2002). All three varieties were tested for serpentine soil tolerance as well as low Ca:Mg ratio and elevated Ni in culture solution (Taylor and Levy, 2002). Of

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms 116 serpentine as a model in evolution them, only var. georgiana displayed some tolerance of serpentine soil, low Ca:Mg ratios, and elevated Ni. Since var. georgiana is not known to naturally grow on serpentine soils, elevated tolerance of serpentine soil factors in the variety is re- garded as an example of preadaptation to serpentine soils.

Poaceae: Ca and Mg Regulation, Type II Cell Wall, Drought Tolerance Poaceae is a global family of herbaceous, wind-pollinated, monocotyledon angio- sperms. Studies of serpentine tolerance and ecotypic differentiation in Poaceae have largely focused on species found in North America and Europe. Serpentine and nonserpentine populations of Agropyron spicatum from Washington grown in serpentine soil and solution culture simulating the chemical conditions of serpen- tine soil have been shown to exhibit ecotypic differentiation (Kruckeberg, 1967; Main, 1974). The serpentine ecotype was able to produce significantly more shoot biomass than the nonserpentine ecotype at lower solution Ca:Mg ratios. Shoot tissue analysis revealed that the serpentine ecotype was able to maintain a signifi- cantly higher shoot Ca:Mg ratio due to selective Ca uptake at the root and translo- cation to the shoot, as well as Mg exclusion at the root or shoot level. Studies of serpentine and nonserpentine populations of Festuca rubra, Agrostis stolonifera, and A. canina from the United Kingdom revealed little difference be- tween the populations with respect to serpentine soil tolerance or Ca, Mg, and Ni uptake in culture solution (Proctor, 1971; Johnston and Proctor, 1981). Like A. stolonifera and A. canina, serpentine and nonserpentine populations of A. hallii from California were also found not to differ in serpentine tolerance (McMillan, 1956). Likewise, growth comparisons of serpentine and nonserpentine popula- tions of Bromus hordeaceus, B. madritensis, B. tectorum, B. diandrus, B. laevipes, and Elymus multisetus (Figure 5.5) from California on serpentine soil did not re- veal any ecotypic differentiation within those species (Kruckeberg, 1950, 1967). Some reciprocal transplant studies involving Avena fatua (Harrison et al., 2001), Bromus hordeaceus (Freitas and Mooney, 1996; Harrison et al., 2001), Vulpia mi- crostachys (Jurjavcic et al., 2002), and Aegilops triuncialis (Thomson, 2007; Lyons et al., in review) from California revealed some ecotypic differentiation, but the evidence implicated soil texture and water availability (drought tolerance) rather than soil chemistry. Drought stress tolerance of the B. hordeaceus serpentine eco- type was attributed to its ability to maintain a slow growth rate and high root:shoot biomass ratio (Freitas and Mooney, 1996). Species in order Poales have type II cell walls (Jarvis et al., 1988; Marschner, 2002; Vogel, 2008). Studies have demonstrated that species of order Poales have a lower Ca requirement than dicot angiosperms (type I cell walls), primarily due to lower cell wall pectin content (Jarvis et al., 1988; Marschner, 2002; Broadley et al.,

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figure 5.5. Response of serpentine (S) and nonserpentine (NS) ecotypes of Elymus mul- tisetus (Poaceae; monocotyledon) to being grown on serpentine (top) and nonserpentine (bottom) soil (photo credit: Kruckeberg, 1950). Note equal growth of NS and S ecotypes on serpentine soil. Species of Poaceae have type II cell walls, which contain much less Ca than the type I cell walls possessed by dicotyledonous plants. The lower Ca requirement of monocotyledonous plants like grasses confers tolerance of the low Ca:Mg ratio of serpen- tine soils. As a result, many or most grass species typically display tolerance of serpentine soil even if they do not naturally grow on serpentine.

2003; Vogel, 2008). Compared to the higher Ca requirement of dicot angiosperms, the lower Ca requirement of species in order Poales could represent a physiologi- cal advantage when growing on Ca-deficient serpentine soils. Wind pollination with its high outcrossing potential (Freidman and Barrett, 2009) combined with the low Ca requirement may explain why species of Poales infrequently exhibit differentiation into distinct serpentine and nonserpentine ecotypes (general indif- ference to serpentine) and why there are relatively few serpentine endemic species within the order.

Pinacae and Cupressaceae: Ca and Mg Regulation, Low Pectin Type I Cell Wall Like Poaceae, ecotypic differentiation of species in the wind-pollinated gymno- sperm genera Pinus (Pinaceae), Picea (Pinaceae), Hesperocyparis (formerly

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Cupressus; Cupressaceae), and Juniperus (Cupressaceae) appears to be weak to nonexistent. Reciprocal transplant field studies of Pinus ponderosa in California have shown that distinct serpentine-tolerant ecotypes exist, however, ecotypic dif- ferentiation is weak and only became evident after 20 years of growing serpentine and nonserpentine ecotypes on serpentine soil (Jenkinson, 1974; Wright, 2007). The serpentine ecotype was shown to maintain significantly higher shoot Ca con- centrations than the nonserpentine ecotype by selective Ca uptake by roots and translocation to the shoot (Jenkinson, 1974). Similar delayed expression of serpentine and nonserpentine ecotypes has been demonstrated for P. contorta in Washington (Kruckeberg, 1967). In shorter recip- rocal transplant studies (1 year) comparing serpentine tolerance of serpentine and nonserpentine populations of P. sabiniana (Griffin, 1962) and P. attenuata (McMillan, 1956) in California and P. virginiana in Maryland (Miller and Cum- ming, 2000), no ecotypic differentiation was detected. Similarly, nonserpentine populations of Picea glehnii and P. jezoensis were found to be equally tolerant of serpentine and nonserpentine soils in Hokkaido, Japan (Kayama et al., 2005). Al- though serpentine and nonserpentine populations of P. jeffreyi (Furnier and Adams, 1986) and P. balfouriana ssp. balfouriana (Oline et al., 2000) from Califor- nia exhibit genetic differentiation, no serpentine soil tolerance studies have been conducted with those species to verify ecotypic differentiation. Like Pinaceae, ecotypic differentiation of species in Cupressaceae is weak to nonexistent. Little difference in serpentine tolerance was found for serpentine and nonserpentine populations of Hesperocyparis macnabiana (formerly Cupressus macnabiana) from California (McMillan, 1956). Juniperus communis has a wide- spread, Northern Hemisphere distribution. Taxonomy of the species is confounded with disagreement of specific recognition of variety and subspecies ranks (Ash- worth et al., 2001; Adams and Pandey, 2003; Adams et al., 2003). At least one rec- ognized variety, var. jackii, is believed to be exclusive to serpentine soils in the northwestern United States (Kruckeberg, 1984; Adams and Nyugen, 2007). The variety exhibits a unique growth form of the species consisting of elongated, sparsely branched, lateral branches (Kruckeberg, 1984). Although some phyloge- netic analyses support taxonomic recognition of var. jackii (Adams and Nguyen, 2007), others do not (Ashworth et al., 2001). Additionally, comparison of serpen- tine tolerance of serpentine and nonserpentine populations has revealed that both are equally serpentine-tolerant (Kruckeberg, 1967). When var. jackii was grown on nonserpentine soil, its growth form reverted to that more typical of nonserpen- tine var. alpina, suggesting that the unique growth form of var. jackii is environ- mentally induced rather than genetically fixed (Ashworth et al., 2001). Similar to Poacae and other species in order Poales, the general indifference of species of Pinaceae and Cupressaceae to serpentine soils may be related to cell wall pectin content. Gymnosperms possess type I cell walls like dicotyledon

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 119 angiosperms, however, studies of Pinaceae and Cupressaceae cell wall composi- tion have shown that they have a substantially lower pectin content (15–35%) than dicotyledon (40–60%) angiosperms (Simson and Timell, 1978a, 1987b; Thomas et al., 1987; Edashige et al., 1995; Piro and Dalessandro, 1998; Hafrén et al., 2000). The lower cell wall pectin content of species of Pinaceae and Cupres- saceae may confer a lower Ca requirement. Low Ca requirement has been noted for Pinus radiata growing on serpentine in New Zealand (Mead and Gadgil, 1978; Lee et al., 1991).

Brassicaceae: Flower Modification, Ca and Mg Regulation, Ni Tolerance and Hyperaccumulation, Parallel and Convergent Evolution Arabidopsis thaliana is one of the most widely used models in the study of plant genetics and development. In a study to identify the underlying physiological tol- erance mechanisms involved in serpentine tolerance, mutants of A. thaliana were screened for tolerance to low solution Ca:Mg ratios (Bradshaw, 2005). Intolerant mutants perished at the cotyledon stage. Survivors were grown to provide tissue for genetic analysis. Genetic analysis of surviving mutants revealed that tolerance of the mutants to low solution Ca:Mg ratios was due to the loss of Ca21–H1 anti- porter CAX1 function. CAX1 is a high capacity Ca-proton antiporter that helps maintain cytoplasmic homeostasis by pumping excess Ca21 from the cytoplasm into the vacuole (Cheng et al., 2003). Mutation of CAX1 effectively acts as a Mg21 exclusion mechanism, allowing the plant to maintain adequate cytoplasmic Ca21 concentrations when subjected to low substrate Ca:Mg ratios. Arabidopis lyrata has been found to have natural serpentine-tolerant ecotypes (Turner et al., 2008; Von Wettberg, 2008). Recently identified candidate genes for serpentine tolerance in A. lyrata serpentine populations from the eastern United States and United Kingdom include K1 transporters (KUP9 and KUP10), cation Ca21 exchanger CCX1 (formerly CAX7; Shigaki et al., 2006), CorA-like Mg21 transporters (MRS2-2, MRS2-6, and MRS2-7), voltage-gated Ca21 channel 1S, and metal tolerance proteins (MTPc3, MTPc1) (Turner et al., 2008, 2010). These can- didate genes suggest that Ca and Mg regulation and heavy metal detoxification are important tolerance mechanisms in the serpentine ecotypes. Loci sequencing of A. lyrata from the United Kingdom has revealed evidence of parallel and conver- gent evolution of serpentine tolerance within ecotypes of the species. Comparison studies of the growth rates of serpentine and nonserpentine ecotypes of A. lyrata on potting soil showed that the serpentine ecotype had significantly slower growth rate than the nonserpentine ecotype, revealing another serpentine tolerance trait in the species (Roche et al., 2004).

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Brassicaceae has a large number of serpentine endemic species, particularly in the genus Streptanthus. Eighteen out of a total of 44 species, subspecies, and varie­ ties in the genus Streptanthus in California are strict serpentine endemics (Safford et al., 2005; Al-Shehbaz and Mayer, 2008). Streptanthus polygaloides is a strict serpentine endemic and the only known species in the genus that hyperaccumulates Ni (Reeves et al., 1981; Boyd et al., 2009). Ni hyperaccumulation in the species has been demonstrated to deter insect herbivory (Jhee et al., 2005). The range of S. polygaloides is limited to the Sierra Nevada mountains in California (Figure 5.6). There is substantial ecotypic varia- tion, especially for flower color and leaf morphology, within Streptanthus species (Table 5.2). S. polygaloides has at least four flower sepal color morphs, including yellow (Y), purple (P), yellow maturing to purple (Y-P), and yellow undulate (U) throughout its range (Boyd et al., 2009). Whereas Y and P morphs have broad but elevationally divided (Y at lower elevations; P at higher elevations) distributions in the northern Sierra Nevada mountains, the Y-P and U populations are geographi- cally restricted, especially the U population, which is almost 100 airline kilometers away from any other color morph populations. Plants of all four color morphs of S. polygaloides were grown in a glasshouse on soil treated with Ni to examine elemental uptake and morphological differences. Significant differences in elemental uptake and leaf morphology were found. The P morph accumulated significantly higher shoot Ni and Mg concentrations than the other three morphs. The geographically isolated U morph had significantly higher K and Zn than the other morphs. Significant differences in plant height and leaf morphology were also found between all four flower color morphs. Morpho- logical and physiological differences between the color morphs suggest that the species is diverging under genetic isolation (Boyd et al., 2009). Chemotaxonomic analysis of two widely separated populations of S. polygaloides within the Y and Y-P morph population range strongly suggests that at least two taxonomically dis- tinct variants are present within the species (Rodman et al., 1981). Further evidence of genetic isolation is provided by phylogenetic studies of the Streptanthus glandulosus complex (Mayer et al., 1994; Mayer and Soltis, 1994, 1999; Mayer and Beseda, 2010). S. glandulosus is found throughout the Coast Range mountains in California (Figure 5.6). The species complex consists of ten subspecies with great variation in flower color, leaf morphology, and affinity for serpentine soils (Table 5.2). Phyogenetic studies have resulted in the identification of four major clades, each corresponding to a geographic area: Northwest, East, Bay Area, and South (Mayer and Soltis, 1994, 1999). Subspecies are distributed within the clades as follows: Northwest—raichei, hoffmanii, and sonomensis; East— arkii; Bay Area—secundus, pulchellus, and niger; South—albidus and glandulosus (Mayer and Beseda, 2010). Subspecies josephinensis is a distant disjunct, occurring in southwestern Oregon, USA. Serpentine soil tolerance tests of serpentine and

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms figure 5.6. Distribution of Streptanthus polygaloides flower color morphs and S. glandu- losus subspecies in California. Gray areas 5 serpentine, S 5 serpentine population, NS 5 nonserpentine population. All populations of S. polygaloides (strict serpentine en- demic) are from serpentine. S. glandulosus ssp. josephinensis, a serpentine endemic dis- junct located in southwestern Oregon, is not shown. It occurs approximately 200 airline km north of the northernmost known populations (approximately delineated by the top edge of this figure) of S. glandulosus in California. Figure drawn from Kruckeberg (1957), Mayer and Soltis (1994), and Boyd et al. (2009).

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms table 5.2 Features of Morphs, Subspecies, and Varieties of Streptanthus polygaloides, S. glandulosus complex, and Caulanthus amplexicaulis

Streptanthus polygaloides (Boyd et al., 2009)

Species Morph Flower Color Leaf Dimensions Serpentine Endemicity* S. polygaloides P purple shorter, narrower Strict S. polygaloides Y yellow shorter, narrower Strict S. polygaloides Y-P yellow maturing to purple longer, wider Strict S. polygaloides U yellow, undulate longer, narrower Strict

Streptanthus glandulosus complex (Kruckeberg, 1957; Mayer and Beseda, 2010)

Species Subspecies Flower Color Leaf Pubescence Serpentine Endemicity* S. glandulosus raichei rose, magenta, purple hairy - hispid Weak S. glandulosus hoffmanii rose, magenta pubescent - hispid Broad S. glandulosus sonomensis pale yellow, creamy white pubescent - hispid Strong S. glandulosus arkii dark maroon, purplish black hairy - hispid Weak S. glandulosus secundus yellow, creamy white pubescent - hispid Strong S. glandulosus pulchellus red, reddish purple pubescent - hispid Broad S. glandulosus niger purplish black glabrous Strict S. glandulosus albidus cream, greenish white glabrous Broad S. glandulosus glandulosus rose, lavender, violet, purple pubescent - hispid Weak S. glandulosus josephinensis white, creamy white pubescent - hispid Strict

Caulanthus amplexicaulis (Pepper and Norwood, 2001)

Species Variety Flower Color Serpentine Endemicity* C. amplexicaulis amplexicaulis purple Weak C. amplexicaulis barbarae yellow, creamy white Strict source: *Safford et al., 2005.

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 123 nonserpentine populations of S. glandulosus showed that the serpentine popula- tions were entirely serpentine tolerant, whereas the nonserpentine populations did not display any tolerance (Kruckeberg, 1950). Response of the serpentine and nonserpentine populations to Ca-reconstituted serpentine soils with increasing Ca:Mg ratios, showed that the serpentine populations were significantly more tol- erant of lower soil Ca:Mg ratios than the nonserpentine populations (Kruckeberg, 1950). Hybrid cross and fertility analysis studies of the subspecies revealed that most subspecies crosses had low interfertility (Kruckeberg, 1957). Only crosses of sub- species raichei and arkii (both formerly S. glandulosus ssp. glandulosus) with se- cundus and pulchellus generally resulted in relatively high interfertility. Within subspecies crosses, interfertility was inversely correlated with distance, with geo- graphically closer populations having greater interfertility than populations fur- ther from each other. Phylogenetic analysis of S. glandulosus subspecies support the findings of hybrid crosses, with genetic identity being correlated with geo- graphical distance between populations (Mayer et al., 1994). Furthermore, serpen- tine populations of ssp. raichei, arkii, and glandulosus were found to have greater genetic identity with nonserpentine populations found within the same region than more distant regions (Mayer and Soltis, 1994, 1999). It appears that serpen- tine tolerance has either evolved numerous times within the genus or been repeat- edly lost among the descendants of a serpentine tolerant ancestor (Mayer and Soltis, 1994). Caulanthus and closely related Streptanthus are both members of the Streptan- thoid complex. Caulanthus amplexicaulis consists of two varieties including the serpentine endemic var. barbarae and the nonserpentine var. amplexicaulis (Pep- per and Norwood, 2001). C. amplexicaulis var. barbarae has yellow flowers and is restricted to serpentine in the far southern Coast Range of California. C. amplexi- caulis var. amplexicaulis has purple flowers and is found exclusively on nonserpen- tine soils at least 75 km away to the east from the nearest var. barbarae popula- tions. Although the two varieties are ecologically and geographically isolated, they are completely interfertile (Pepper and Norwood, 2001). Phylogenetic analysis of populations of the two C. amplexicaulis varieties revealed that the varieties are distinct and monophyletic. Analysis of other species of Caulanthus and Streptan- thus included in the study, however, revealed that the two genera are not mono- phyletic. This result is supported by chemotaxonomic studies of both genera (Rodman et al., 1981). Additionally, serpentine endemism is not monophyletic, suggesting that serpentine tolerance and subsequent endemism has arisen inde- pendently several times in the Streptanthoid complex (Rodman et al., 1981; Pep- per and Norwood, 2001). Many reassigned species of genus Thlaspi (tribes Thlaspidae and Noccaeeae; Brassicaceae) are Ni hyperaccumulators and serpentine endemics (Brooks, 1998;

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Peer et al., 2003). Recent phylogenetic studies of genus Thlaspi and closely related genera revealed that Thlaspi is not monophyletic (Koch and Mummenhoff, 2001; Koch and Al-Shehbaz, 2004). All New World and most Old World Thlaspi have since been reassigned to genus Noccaea (Koch and Al-Shehbaz, 2004). Although not discussed by the authors, the phylogenies suggest either that serpentine toler- ance and heavy metal hyperaccumulation has evolved numerous times within tribe Noccaeeae, or that the ancestral state of Noccaeeae is presence of serpentine tolerance and heavy metal hyperaccumulation, with some derivative species hav- ing lost one or both of the traits. Noccaea fendleri includes ssp. glauca, idahoense, siskiyouense (all formerly vari- eties of Thlaspi montanum), and californica (formerly Thlaspi californicum). N. fendlerii has a widespread western U.S. distribution. N. fendlerii ssp. glauca popu- lations occur on both serpentine and nonserpentine substrates in California, Or- egon, and Washington (Kruckeberg, 1967; Reeves et al., 1983; Boyd and Martens, 1994). N. fendlerii ssp. siskiyouense and californica are localized serpentine endem- ics in southwestern Oregon and northern California, respectively (U.S. Fish and Wildlife Service, 2003). Like most other Noccaea species, N. fendleri ssp. glauca, siskiyouense, and californica are all known to be Ni hyperaccumulators (Reeves et al., 1983). Serpentine and nonserpentine populations of N. fendleri ssp. glauca from Washington exhibit ecotype differentiation with respect to serpentine soil toler- ance in both the glasshouse and the field (Kruckeberg, 1967). Comparison of the Ni uptake of serpentine and nonserpentine populations of N. fendleri ssp. glauca from California in the glasshouse showed that both serpentine and nonserpentine populations hyperaccumulate Ni (Boyd and Martens, 1998). Ni hyperaccumula- tion in the species has been shown to deter insect herbivory (Boyd and Martens, 1994). Field-collected plant tissue of N. fendleri ssp. glauca from serpentine (high Ni soil) and nonserpentine (very low Ni soil) populations were analyzed, and only plants from the serpentine population were found to have Ni concentrations high enough to indicate hyperaccumulation (Boyd and Martens, 1998). When plants of both serpentine and nonserpentine populations were grown on the same Ni-rich substrate in a glasshouse and the tissue was analyzed, however, plants of both pop- ulations exhibited Ni hyperaccumulation (Boyd and Martens, 1998; Peer et al., 2006). It has been suggested that the nonserpentine populations of N. fendleri ssp. glauca are effectively preadapted for serpentine tolerance, and Ni hyperaccumula- tion is a constitutive trait (Boyd and Martens, 1998). Similar results were found for Noccaea goesingense (formerly Thlaspi goesingense; Austria; Reeves and Baker, 1984) and Noccaea cochleariforme (formerly Thlaspi japonicum; Japan; Mizuno et al., 2003, 2005) serpentine and nonserpentine populations. Noccaea caerulescens (formerly Thlaspi caerulescens) from Europe exhibits dif- ferentiation with serpentine ecotypes displaying significantly greater Ni tolerance

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 125 and Ni uptake than nonserpentine ecotypes (Assunção et al. 2003, 2008). Signifi- cant variation in Ni uptake has also been found between serpentine populations of the serpentine endemic Noccaea pindica (formerly Thlaspi pindicum) from Greece (Taylor and Macnair, 2006). Some populations of the species display significantly greater Ni uptake and root-to-shoot translocation than other populations when grown in the same substrate (Taylor and Macnair, 2006). Like Noccaea, many species of genus Alyssum (Brassicaceae) are also Ni hyper- accumulators and serpentine endemics (Brooks, 1998). Comparison of serpentine and nonserpentine populations of A. murale grown in culture solution with Ni showed that the serpentine population was significantly more tolerant of elevated Ni concentrations than the nonserpentine population (Asemaneh et al., 2006). Tissue Ni analysis of nine populations of the serpentine endemic A. bertolonii grown in culture solution containing Ni revealed ecotypic variation with respect to both Ni tolerance and uptake (Galardi et al., 2007), similar to Noccaea pindica. Phylogenetic analysis of species and subspecies in Alyssum has revealed that the Ni hyperaccumulation mechanism has been lost or acquired several times (Mengoni et al., 2003). Studies of serpentine and nonserpentine populations of A. inflatum, a Ni hyperaccumulator from Iran, showed that plants from the serpentine popula- tion were more tolerant of low solution Ca:Mg ratios than plants of the nonserpen- tine population when grown in culture solution (Ghasemi and Ghaderian, 2009). There was no difference between plants of the two populations with respect to Ni tolerance or uptake.

Adaptation Trends As shown with the many diverse taxa, several recurring adaptations to serpentine soils are evident. These adaptations include (1) inherently slower growth rate; (2) high root:shoot mass ratios; (3) early flowering time; (4) regulation of Ca and Mg, including selective Ca uptake and enhanced root-to-shoot translocation and Mg exclusion at the root level or restricted root-to-shoot translocation; and (5) heavy metal regulation, including root-level exclusion or sequestration, and varia- tion in heavy metal uptake and root-to-shoot translocation in heavy metal hyperaccumulators. Inherently slower growth rate, high root:shoot mass ratios, and early flowering time appear to be strongly associated with drought tolerance, although slower growth rate and high root:shoot mass ratio may also help cope with low macronu- trient (N, P, K) availability. Regulation of Ca and Mg uptake is associated with the maintenance of balanced tissue Ca and Mg concentrations in response to very low Ca and high Mg soil concentrations. Species of order Poales contain type II cell walls, which have much lower Ca concentrations than type I cell walls in most other vascular plant species. As a result, species of Poales are effectively preadapted

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms 126 serpentine as a model in evolution to low substrate Ca concentrations and do not appear to display ecotypic differen- tiation with respect to Ca and Mg regulation as frequently as species in other plant families. Root-level heavy metal exclusion and sequestration are common toler- ance mechanisms to the elevated bioavailable heavy metal concentrations in ser- pentine soils. Evolution of heavy metal hyperaccumulation has been demonstrated in ecotypes of a number of species. Studies have shown that metal hyperaccumula- tion confers protection from insect herbivory and serves a role in osmotic regula- tion and drought tolerance in some species. Recurring evolutionary trends in selection for serpentine tolerance are evi- dence for preadaptation and ecotypic differentiation occurring by catastrophic selection. This mode of selection appears to be especially common for tolerance mechanisms involving Ca and Mg regulation. Convergent and parallel evolution of ecotypes numerous times within the same species also appears to be common and is represented by several species within diverse families. The role of parallel evolution in the origin of species (Levin, 2001) is at an early stage of discovery, and we believe that serpentine ecosystems can provide model settings for the study of parallel speciation (i.e., the parallel origins of traits that confer adaptations and reproductive isolation) in plants.

Summary Plant adaptation and evolutionary mechanisms on serpentine soils bear many similarities to other phytotoxic soils such as alkaline, saline soils and acidic, metal ore mine tailings. Unlike mine tailings and serpentine, saline soils can often have a broad, continuous, widespread distribution, especially in arid inland environ- ments and coastlines (Läuchli and Lüttge 2002). In contrast, mine tailings and serpentine areas are limited and typically have a discontinuous, widespread distri- bution across the landscape, resembling island environments.

Although metal ore mine tailings globally have the same general conditions of low pH and high bioavailable metal concentrations, these conditions can be highly variable due to mine host rock type, ore type, and quantity of associated elemental sulfur and sulfate-bearing minerals (Quilty, 1975; Williamson et al., 1982; Tordoff et al., 2000). Compared to metal ores and their host rock types, serpentine has a relatively consistent mineral composition (Brady et al., 2005; Alexander et al., 2007; Kazakou et al., 2008). Regardless of climate, topography, and associated or- ganisms, serpentine consistently weathers to produce shallow, rocky soils, low in bioavailable N, P, K, and Ca and high in bioavailable Mg, and often Ni, Cr, and Co. Low Ca:Mg ratio is the defining characteristic of serpentine soils worldwide. Serpentine soils have a widespread distribution, occurring on every continent and found in virtually every type of climate (Kruckeberg, 1984; Brooks, 1987; Roberts and Proctor, 1992). Serpentine also supports a high diversity of plant

This content downloaded from 129.65.41.106 on Wed, 16 Oct 2019 01:33:07 UTC All use subject to https://about.jstor.org/terms intraspecific variation 127 species and vegetation types. Additionally, it is a natural phytotoxic environment as opposed to being an unnatural, anthropogenic, phytotoxic environment like mine tailings. Widespread, island-like distribution; consistency of adverse soil chemical conditions; diversity of climate, plant species, and vegetation type; and natural, relatively undisturbed conditions make serpentine soils ideal model eco- systems for studies of intraspecific variation and resulting plant evolution.

Acknowledgments We thank Mike Mayer (University of San Diego) and Ihsan Al-Shehbaz (Missouri Botanical Garden) for taxonomic information regarding Streptanthus and Noc- caea, Tanner Harris for assisting in the literature searches, Kelly Bougher for com- piling literature cited, and Jessica Wright and Bob Boyd for useful feedback during manuscript preparation.

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