The evolution of a single toe in horses - NSF PAR

The evolution of a single toe in horses: causes, consequences, and the way forward 1

Brianna K. McHorse,1,*,†,‡ Andrew A. Biewener,*,† and Stephanie E. Pierce*,‡ 2

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*Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, MA 02138, USA; 4

†Concord Field Station, Harvard University, Bedford, MA 01730, USA; ‡Museum of Comparative Zoology, 5

Harvard University, Cambridge, MA 02138, USA 6

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1 [email protected] 8

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Abstract 10

Horses are a classic example of macroevolution in three major traits—large body size, tall-11

crowned teeth (hypsodonty), and a single toe (monodactyly)—but how and why monodactyly evolved is 12

still poorly understood. Existing hypotheses usually connect digit reduction in horses to the spread and 13

eventual dominance of open-habitat grasslands, which took over from forests during the Cenozoic; digit 14

reduction has been argued to be an adaptation for speed, locomotor economy, stability, and/or 15

increased body size. In this review, we assess the evidence for these (not necessarily mutually exclusive) 16

hypotheses from a variety of related fields, including paleoecology, phylogenetic comparative methods, 17

and biomechanics. Convergent evolution of digit reduction, including in litopterns and artiodactyls, is 18

also considered. We find it unlikely that a single evolutionary driver was responsible for the evolution of 19

monodactyly, because changes in body size, foot posture, habitat, and substrate are frequently found to 20

influence one another (and to connect to broader potential drivers, such as changing climate). We 21

conclude with suggestions for future research to help untangle the complex dynamics of this remarkable 22

morphological change in extinct horses. A path forward should combine regional paleoecology studies, 23

Evolution of a single toe in horses 2

quantitative biomechanical work, and make use of convergence and modern analogs to estimate the 24

relative contributions of potential evolutionary drivers for digit reduction. 25

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Total words in text: 7608 27

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Introduction 29

Horse evolution and grasslands 30

Horses are the only living members of the family Equidae, which today comprises just six species 31

in the genus Equus (including zebras, asses, and caballine horses, the group to which domestic horses 32

belong). In contrast to today’s paucity of species, the equid fossil record includes nearly 50 genera and 33

hundreds of species over the last 58 million years (MacFadden 1994). The earliest equids were only dog-34

sized, with four toes on the foreleg and three on the hind leg (MacFadden 1994). Today’s horses are 35

large, long-legged grazers with a single toe on each leg, which is enclosed in a hard hoof. An enlarged 36

third digit makes up the bulk of the distal limb, with considerably reduced metapodials II and IV present 37

as splint bones fused to the center metapodial (Figure 1). Recent work has shown that vestiges of digits I 38

and V may still be present as ridges and wings in the proximal metapodial and distal phalanx (Solounias 39

et al. 2018). Despite their large size and long, slender limbs, horses are considerably athletic, reaching a 40

recorded top racing speed of 70 km/hr (“Fastest speed for a race horse” 2019); the highest jump 41

recorded by a domestic horse and rider is 2.47 m (“Highest jump by a horse” 2019). That horses can 42

accomplish such feats on a single toe, which evolved millions of years prior to human influence, is 43

remarkable. 44

Fossil horses played a critical role in both supporting Darwin’s theory of evolution and, later, the 45

Modern Synthesis (Simpson 1951). In the 1870s, O.C. Marsh had made a considerable collection of fossil 46

horses, which he then arranged into a series of small to large, three-toed to one toe, low-crowned teeth 47


to high-crowned teeth (Marsh 1874). This proposed evolutionary series was so striking for its time that 48

after seeing it, T.H. Huxley, “Darwin’s Bulldog,” rewrote an address to be given at the New York 49

Academy of Sciences to include these fossil horses as evidence of evolution (Schuchert 1940). At the 50

time, orthogenesis—an evolutionary “progression” in a straight line towards some ideal form—was a 51

popular conception of evolution, and this arrangement of horses supported that view (Figure 2). Thus 52

the classic story of horse evolution was formed: as grasslands took over from forests, the horse 53

gradually evolved larger body size (perhaps to better defend against predators), taller-crowned teeth to 54

handle abrasive grasses, and long, monodactyl limbs to race away from predators in their newly open 55

habitat (Figure 2; Matthew 1926). 56

Despite subsequent recognition that equid evolution was in fact more like a bush than a straight 57

line (Simpson 1951; MacFadden 1994), it is still portrayed in a linear fashion in many museums and 58

textbooks (MacFadden et al. 2012). Some trends in equid evolution do appear to exist by gestalt—59

today’s horses are indeed much larger, hypsodont, and have reduced digits relative to the earliest 60

horses. Monodactyly had two separate evolutions, one in the Dinohippus/Equus lineage and one in the 61

Pliohippus/Astrohippus lineage, strongly suggesting at least some adaptive utility and selection for this 62

condition. It therefore requires careful attention to discuss the evolution of horses without slipping into 63

verbal orthogenesis by drawing a straight line between the earliest horse and the lone surviving genus 64

today, particularly given that trends of digit reduction and increasing hypsodonty do exist in at least 65

some parts of the horse tree (Janis 2007). But evidence from diet, habitat, tooth morphology, and digit 66

state do not match the orthogenetic pattern: decreasing body size was common in lineages such as the 67

Archaeohippus or Nannippus; not all tridactyl horses browsed; and not all hypsodont, monodactyl 68

horses grazed (MacFadden 1994; MacFadden et al. 2012). 69

Beyond the pattern itself, the classic explanations for why horses evolved the way they did is 70

tremendously “sticky” (Schimel 2012). Long after the complexity of the equid tree and the nonlinearity 71


of trait evolution was acknowledged, the initial explanations for each horse trait still held the weight of 72

established fact rather than reasonable hypothesis. The evolutionary story of horses has seen several 73

advances in understanding over the last few decades, particularly as powerful quantitative methods 74

emerge and we accumulate more available specimens through fieldwork, museum cataloguing, and 75

especially digitization (Marshall et al. 2018). With these new data and methods, untested explanations 76

for horse trait evolution have been challenged one by one. 77

The simple causal relationship between abrasive grass and hypsodonty has been shown to be 78

complicated, with grasslands predating hypsodonty by at least four million years in horses, rodents, and 79

lagomorphs (Strömberg 2002, 2006; Jardine et al. 2012). Tooth mesowear, a macroscopic measure of 80

tooth wear that can record information about diet, is highly variable within fossil horse populations and 81

does not always match up directly with grasslands and hypsodonty (Mihlbachler et al. 2011); 82

furthermore, fresh grazing can cause mesowear similar to browsing (Winkler et al. 2019). In extant taxa, 83

hypsodonty correlates more with habitat openness (Mendoza and Palmqvist 2008)than with the 84

proportion of grass consumed, and feeding height (which relates to the amount of soil grit consumed) 85

drives microwear more than diet (Mainland 2003). The study of tooth enamel isotopes has also 86

complicated the relationship between hypsodonty and diet; for example, in one locality, the hypsodont 87

horse species were likely browsers while the species with low-crowned teeth were consuming more 88

grasses (MacFadden et al. 1999), and individual variation in isotope values can be high in large 89

herbivores (Green et al. 2018). Hypsodonty seems to be driven mostly by grit (via phytolith, dirt, or 90

volcanic ash), not grass alone, and evolved under much less of a straightforward evolutionary arms race 91

than initially thought. 92

Increasing body size, which was initially explained as a defense mechanism against predation 93

(Matthew 1926), has also been suggested as an example of Cope’s rule (that lineages tend to increase in 94

maximum body size through time) in equids (Martin 2018), perhaps as a result of ecological 95


specialization (Raia et al. 2012). Others have argued that because grass is generally less nutritious than 96

browse (but see Codron et al. 2007), larger body size was beneficial because it increased total digestive 97

capacity, thus allowing the animal to process larger quantities of low-quality food (Demment and Van 98

Soest 1985; Lovegrove and Mowoe 2013). A recent study showed strong evidence for transitions to an 99

unguligrade foot posture being associated with rapid increases in body size, and rate of body size 100

evolution, across mammals (Kubo et al. 2019), pointing to the possibility that unguligrady supports 101

larger body sizes. Kubo et al. (2019) suggested that larger body sizes may provide a release from higher 102

levels of predation, again connecting predator pressures to horse evolution. However, many equid 103

lineages retained similar body sizes through evolutionary time or even became smaller (MacFadden 104

1994), irrespective of expanding grasslands. Recent work suggested that more than 90% of changes in 105

horse body size can be explained simply by diffusion, a random walk of evolution, rather than 106

competition for niches (Shoemaker et al. 2013), so the pattern of increasing body mass may not be a 107

trend at all. 108

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Modern hypotheses for digit reduction 110

Like hypsodonty and body size, the story of digit reduction is complicated. The classic 111

explanation was that high speeds were necessary for predator escape in open grasslands, with reduced 112

digits on elongated limbs providing this speed (Matthew 1926; Simpson 1951). But high-speed pursuit 113

predators such as wolves did not evolve until approximately 20 million years after many ungulates, 114

including equids, evolved lengthened limbs and reduced digits (Janis and Wilhelm 1993), providing 115

evidence against this hypothesis. Subsequent years have seen three other major hypotheses about the 116

proximate driving force behind digit reduction in horses: 117

1) The locomotor economy hypothesis: Open, arid grasslands required longer travel distances to 118

access patchy resources, such as water, and elongated limbs decreased the energetic cost of 119


locomotion by increasing stride length (Janis and Wilhelm 1993). While reduced digits were not 120

explicitly discussed in the referenced paper, reducing mass in the distal limb would decrease 121

moment of inertia and thus the energetic cost of swinging the leg (Hildebrand 1960; Myers and 122

Steudel 1985; Browning et al. 2007; Kilbourne et al. 2016), and has been argued to be a driver of 123

digit reduction in archosaurs (de Bakker et al. 2013). 124

2) The stability hypothesis: Forests required lateral dodging movements on soft ground, whereas 125

grasslands required high-speed, straight-line movements on hard ground; a tridactyl and 126

monodactyl foot were respectively better suited to stability in those environments (Shotwell 1961). 127

Effectively, Shotwell proposed two separate hypotheses: 2a) the soft substrate stability hypothesis, 128

that the tridactyl foot is adapted for stability in lateral dodges on soft ground, and 2b) the hard 129

substrate stability/speed hypothesis, that the monodactyl foot is adapted for stability and speed in 130

straight lines on hard ground. The superior speed of the monodactyl foot on hard ground was also 131

proposed by Matthew (1926), who focused on the rigid hoof and spring-like tendon-ligament system 132

of extant equids. 133

3) The body size hypothesis: Evolutionary increases in body mass produced greater bending forces on 134

the limbs, and a single digit resists bending forces better than several smaller digits of the same total 135

size (Thomason 1986). 136

None of these hypotheses are mutually exclusive. The first two assume that grasslands act as 137

the ultimate driver of digit reduction (via the proximate causes of increased locomotor demands and 138

changing substrate conditions, respectively). The third hypothesis suggests body mass as a proximate 139

cause. As discussed previously, proposed drivers of body mass itself in horses range from evolutionary 140

diffusion (Shoemaker et al. 2013) to grasslands (Illius and Gordon 1992), consistent with the first two 141

hypotheses. Body mass increase in response to the evolution of grasslands might also partially be 142

related to cooling climate (Lovegrove and Mowoe 2013), but climate cooling should also be considered 143


as one of many potential drivers of grasslands themselves (Strömberg 2011). Therefore, if the body size 144

hypothesis is correct, the ultimate driver of digit reduction could be any or a combination of other 145

changes in forage and climate. 146

In this review, we survey what evidence exists to support or refute these digit reduction 147

hypotheses. First, we give a brief overview of horse evolution with a focus on digits. Next, we discuss an 148

analytical method for quantifying the degree of digit reduction to provide a continuous metric for 149

evolutionary analyses. We follow this with a review of research from biomechanics, macroevolution, and 150

other subdisciplines that has brought new insights into digit reduction in recent years. Finally, we 151

conclude with a discussion of critical gaps in our understanding and make suggestions for avenues of 152

future study. 153

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Overview of horse digit evolution 155

The phylogenetic relationships of the earliest equids have seen considerable changes since 156

Hyracotherium/Eohippus was universally considered the first horse (Figure 3). Recent phylogenetic work 157

has split Hyracotherium into H. leporinum, now considered a basal palaeothere (outside of Equidae), and 158

an array of new genera for basal true equids, including Sifrhippus and Arenahippus (Froehlich 2002). 159

Another recent phylogenetic analysis has placed Ghazijhippus (found in Pakistan) and Cymbalophus and 160

Pliolophus (found in Europe) at the base of the equid tree, basal to the North American Sifrhippus and 161

Arenahippus (Bai et al. 2018). Regardless which genus is most basal, the earliest equids were small, dog-162

sized creatures that had four digits on the forefoot and three on the hind foot (semi-tetradactyl). They 163

had low-crowned (brachydont) teeth indicative of a diet of fruits and soft leaves, and although they 164

seem “primitive” relative to extant horses, a remarkably complete skeleton of the basal horse 165

Arenahippus grangeri (previously H. grangeri; considered a junior synonym of Sifrhippus by Secord et al. 166

2012, but left separate here) shows that it was fairly derived for the time in its foot posture 167


(subunguligrade) and somewhat elongate metapodials (Kitts 1956; Wood et al. 2011). Although the 168

shoulder and hip joints had considerable range of movement, the distal limb in A. grangeri was already 169

primarily restricted to parasagittal motion, as in later equids (Wood et al. 2011). 170

Coeval with this Arenahippus was another, similarly small equid, Orohippus, which had even 171

more restricted movements in the distal limb due to more stable carpal and tarsal articulations. Wood 172

et al. (2011) hypothesized that Orohippus may have occupied more open terrain than A. grangeri, which 173

was found in tropical forests—reminiscent of Shotwell’s (1961) stability vs. substrate hypotheses. Wood 174

et al. (2011) suggest the following chain of events: as climate changed in the Paleogene (66-12 Ma), 175

equid diets shifted from high-quality fruits and leaves to lower-quality browse and graze; such a dietary 176

shift drove increases in body size (consistent with results from Secord et al. (2012)) to allow processing 177

greater amounts of food; and finally, increased body size drove a need for more centrally-located, 178

upright limbs (reminiscent of Camp and Smith’s (1942) argument for lineage-scale digit reduction). 179

Like earlier equids, the three-toed (tridactyl) species Mesohippus, Miohippus, and Anchitherium, 180

were also likely subunguligrade, with all distal phalanges contacting the ground and supported by a foot 181

pad (Camp and Smith 1942; Sondaar 1968; Thomason 1986). However, relative to earlier equids, their 182

limbs became more restricted to a pendulum-like motion in the parasagittal plane via limb bone fusion 183

(radius-ulna, tibia-fibula) and changes in joint articulations (Sondaar, 1968). Along with increasingly 184

parasagittal motion came the lengthening of the limb, particularly distally. In later lineages, beginning 185

with Parahippus at the base of the grazing radiation, limb elongation continued, and the lateral digits 186

were reduced; the side toes likely did not touch the ground at rest (Sondaar, 1968). In the monodactyl 187

or nearly-monodactyl lineages, such as Pliohippus and Equus, a tendon-and-ligament suspensory 188

apparatus and a ‘springing’ foot evolved, with markedly elongate phalanges and considerable elastic 189

energy storage (Biewener 1998) that may have benefited them on hard ground in open grasslands 190

(Matthew 1926; Sondaar 1968; Janis and Wilhelm 1993). 191


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Quantifying digit reduction 193

Until recently, one challenge of studying digit reduction in horses was the lack of a quantitative 194

way to measure digit reduction. The discrete categories of semi-tetradactyl (four toes in front and three 195

behind), tridactyl (three toes), and monodactyl (one toe) are useful, but fail to capture a wide variety of 196

morphological (and probably functional) diversity throughout the main body of the equid phylogeny 197

(Figure 3). Furthermore, many modern analyses require continuous variables to reconstruct the mode or 198

rate of evolution (O’Meara and Beaulieu 2014). We addressed this gap in two recent papers, where we 199

introduced the Toe Reduction Index (TRI), a continuous measure of digit reduction for perissodactyls 200

(McHorse et al. 2017; Parker et al. 2018). TRI is measured as the ratio of side digit length to center digit 201

length in the proximal phalanx (Equation 1, Figure 4), taking the average of side digits if they are both 202

available, and is best calculated for each individual before averaging across a species or genus. The index 203

ranges from 0 (no side digits, as in Equus) to 1 (all digits equal in length): 204

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Equation 1. 𝑇𝑅𝐼 = 𝑠𝑝𝑒𝑐𝑖𝑒𝑠 𝑚𝑒𝑎𝑛 (𝑚𝑒𝑎𝑛(𝑃𝑃𝑙𝑒𝑛𝑔𝑡ℎ𝐼𝐼,𝑃𝑃𝑙𝑒𝑛𝑔𝑡ℎ𝐼𝑉)

𝑃𝑃𝑙𝑒𝑛𝑔𝑡ℎ𝐼𝐼𝐼) 206

207

where PPlength refers to the maximum articular length of the proximal phalanx in digit II, IV, or III 208

according to the subscript; the mean of PPlength is first taken for all available side digits (II and IV), then 209

divided by the mean of PPlength for digit III. This provides the individual-level TRI, which is then 210

averaged across all individuals in a species to provide a species-level TRI (Equation 1). Values greater 211

than 1 are theoretically possible and would correspond to lateral digits greater in length than the center 212

digit; however, this seems unlikely to occur. 213


With the Toe Reduction Index, we can quantitatively represent the real morphological variation 214

present in equid digits (Figure 5). Whereas previously all tridactyl horses would be coded the same in 215

categorical data, TRI values range from nearly 1 (all three digits of equal size) to less than 0.3 (side digits 216

⅓ the size of the center digit), illuminating variation that was previously unavailable to quantitative 217

analyses. These new data make it possible to address questions such as whether digit reduction 218

correlates with changes in other traits, e.g., hypsodonty or body mass (Parker et al. 2018). Furthermore, 219

TRI can be used in the future to explore digit reduction in a variety of other groups, including 220

artiodactyls, with appropriate modification to account for paraxonic symmetry vs. mesaxonic symmetry 221

(i.e., artiodactyls have symmetrical enlarged digits III and IV with the axis of symmetry running between 222

them, whereas TRI was developed for a single, symmetrical center digit III). A TRI dataset expanded to 223

other taxa would open up the possibility of both more quantitative taxon-specific studies and more 224

broadly comparative studies. 225

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Biomechanical investigation of digit reduction 227

As terrestrial quadrupeds, horses primarily use their limbs to interact with the environment 228

through locomotion. The forces that act on bones during an animal’s life can be a powerful source of 229

selection; the geometry of bones can often indicate, for example, locomotor style or other functional 230

uses for that part of the body (Swartz et al. 1992; Anyonge 1996; Doube et al. 2018). In domestic horses, 231

many biomechanical studies have linked skeletal morphology to performance in competition, 232

connecting form to function (Barrey et al. 2002; Gnagey et al. 2006; Weller et al. 2006; Hobbs et al. 233

2010; Kristjansson et al. 2016). An extra load of just 2.4 kg on a horse’s distal limb has been shown to 234

increase cost of transport by nearly 7% (Wickler et al. 2004), providing a direct connection to the 235

locomotor economy hypothesis of digit reduction. 236


Studies examining the biomechanical and physiological consequences of limb morphology in 237

extant horses rarely connect to the fossil record and to equid evolution, but a combination of these 238

disciplines offers considerable insight into outstanding questions like the driver of equid digit reduction. 239

Although biomechanical performance data cannot be obtained for extinct animals, musculoskeletal 240

modeling is a powerful tool to reconstruct soft-tissue dynamics, forces, and ultimately provide insight 241

into performance in extinct species (Hutchinson and Garcia 2002; Pierce et al. 2012, 2013; Nyakatura et 242

al. 2019). Such studies usually make use of detailed skeletal data from extinct species, sometimes 243

combining it with experimental biomechanical data on extant taxa. Biomechanical studies can therefore 244

help fill in the relationship between morphology, performance, and the environment, helping to connect 245

patterns evident at the macro-level (morphological and taxonomic change over millions of years) to the 246

individual level (where morphology and performance determine fitness; Figure 6). 247

Beam bending is a mechanical engineering approach used to calculate stresses on structural 248

beams. When applied to skeletons, beam bending analyses determine the stresses experienced by a 249

bone using the bone’s own internal geometry, the forces it experiences from muscle contractions and 250

external sources (such as a food item being bitten or the ground contacting a foot), and the angles and 251

moment arms at which those forces act. While frequently used to explore the effects of bite forces in 252

the skull (e.g., Van Valkenburgh and Ruff 1987; Busbey 1995; Therrien 2005), beam bending has also 253

been used to estimate locomotor forces in the limbs of extant and extinct animals (Biewener 1983; 254

Alexander 1985; Blob and Biewener 2001), including in horses (Biewener et al. 1983). In extinct equids, 255

beam bending was first applied to the center metapodial to explore locomotor stresses in Mesohippus 256

(subunguligrade), Merychippus (unguligrade), and modern Equus (unguligrade), using in-vivo strain 257

gauge data recorded from metapodials of living horses to ground-truth the method (Thomason 1985). 258

Using broken metacarpals to assess internal geometry and reducing forces in Mesohippus by 50% to 259

account for load-bearing side digits, Thomason (1985) found midshaft metacarpal stresses to be similar 260


in the extinct and extant horse. Stresses were highest in the unguligrade grazer Merychippus, suggesting 261

that size increase alone did not drive the transition from the subunguligrade to the fully unguligrade 262

foot; Thomason suggested that habitat could have been the major other factor driving this 263

morphological change in the distal limb (Thomason 1985). 264

In a recent study, we used CT scans of fossil metapodials to apply beam bending to the same 265

question in higher anatomical resolution and across much more of the equid phylogeny (McHorse et al. 266

2017). Midshaft stress under high-performance locomotion in the center metapodial was calculated for 267

extinct species in twelve equid genera, first with a full body-weight load on the center digit and then 268

with body-weight load reduced proportional to the size of the side digits using TRI (scaled as in Equation 269

2): 270

271

Equation 2. 𝑙𝑜𝑎𝑑𝑇𝑅𝐼 = 𝑙𝑜𝑎𝑑𝑏𝑜𝑑𝑦 ∗ (1

(2 ∗ 𝑇𝑅𝐼) + 1) 272

273

where loadTRI is the load on the center digit, loadbody is a species average body weight or the body weight 274

of the individual, and TRI is the Toe Reduction Index as calculated by Equation 1. The scaling is such that, 275

at a TRI of 1, the body weight is distributed equally among the three toes, and at a TRI of 0, the entire 276

body weight is on the center digit. 277

Our results supported and expanded on Thomason’s (1985) work, showing that when the side 278

digits bear load proportional to their size, bone safety factors (ratio of failure stress to peak locomotor 279

stress) were in the range of those found for extant mammals from mice to elephants (2 to 4; Biewener 280

1991). In contrast, when the side digits did not reduce the load on the center metapodial, stresses close 281

to or surpassing the tensile fracture stress of bone were reached in taxa as late as Parahippus, indicating 282

that side digits were mechanically critical for resisting stress until at least the beginning of the grazing 283


radiation (Figures 2 and 4). Furthermore, the center metapodial is positively allometric relative to body 284

mass in its cross-sectional geometry (i.e. resistance to compressive and bending forces), meaning that 285

the center digit compensated for reduced digits through evolutionary time (McHorse et al. 2017). This 286

positive allometry lends some support to the body size hypothesis, that a single large digit is a response 287

to increasing body sizes and can better resist the increased loads. The evolution of unguligrady, which 288

has been connected to increased rates of body size evolution (Kubo et al. 2019), could have spurred 289

these changes indirectly. The allometry results of this study (and the timing of side digits becoming 290

unnecessary for load-bearing) are also consistent with the locomotor economy hypothesis; it is possible 291

that as longer strides and thus longer limbs were favored by selection, the inertial costs of maintaining 292

side toes began to outweigh any remaining stabilizing or load-bearing benefit. 293

294

Evidence from macroevolution, biogeography, and ecology 295

Selection for digit reduction on an evolutionary scale requires morphological changes to 296

influence fitness by changing how the animal performs in its environment (Figure 6; Arnold 1983). In 297

other instances of digit reduction, new selective pressures frequently come from new ecologies or 298

locomotor modes, such as the cetacean transition into water (Shapiro et al. 2007) or the evolution of 299

ricochetal locomotion coupled with out-in-the-open foraging in jerboas (Moore et al. 2015). In horses, 300

various proximate causes of digit reduction have been suggested, as illustrated by the hypotheses set 301

forth earlier in this paper. Yet the most generally accepted ultimate cause of digit reduction in this 302

group, the one that makes for a new relationship between morphology and fitness, is the evolution of 303

grasslands. 304

Virtually no macroevolutionary work has explicitly addressed digit reduction in horses, but many 305

studies focus on horse macroevolution more generally. Of particular interest is the Miocene grazing 306

radiation of horses in North America (18 to 15 Ma), which began with Parahippus—the same genus 307


found to be among the last in which side toes were critical for mechanical support (Figures 2 and 4; 308

McHorse et al. 2017). Diversification rates were high and at least 19 new species originated quite 309

rapidly, although rates of morphological evolution were not elevated (MacFadden and Hulbert 1988; 310

Cantalapiedra et al. 2017). Though the radiation was suggested to be in response to grasslands, rapid 311

diversification lagged grasslands by several million years (Strömberg 2006; Cantalapiedra et al. 2017). 312

Most speciation events during the Miocene radiation were in fact via dispersal into new regions 313

(Maguire and Stigall 2008). Because dispersal was the main driver of speciation, factors that facilitated 314

movement—such as habitat fragmentation due to tectonic and climatic events—promoted speciation 315

(Stigall 2013). If digit reduction promoted greater economy of locomotion, it could therefore have 316

indirectly supported speciation. 317

There is no denying the scope of environmental change that accompanied the approximately 318

58-million-year history of horse evolution. In North America, temperatures swung from the warmth of 319

the Paleocene-Eocene thermal maximum, through a gradual, bumpy cooling spanning the Eocene, 320

Oligocene, and Miocene (periodically interrupted by warmer peaks lasting a few million years), and 321

finally dropped into the cyclical chill of Ice Ages (Figure 7; Zachos et al. 2001). These thermal changes 322

accompanied precipitation changes, from the wet tropical forest of warmer periods through increasing 323

aridity as the climate cooled (Janis 1993). It is against this climatic backdrop that grasslands evolved in 324

North America, becoming regionally dominant ecosystems approximately 22 million years ago and 325

dominant across North America by 7-11 million years ago (Strömberg 2005, 2006). 326

As the environmental landscape changed, varied habitats appeared. This variety is the basis of 327

Shotwell’s (1961) hypothesis that tridactyl horses and monodactyl horses were better suited to different 328

habitats—woodland-savanna and grasslands, respectively—and thus partitioned habitats accordingly. 329

Shotwell (1961) tracked the biogeography of the genera Hipparion (tridactyl) and Pliohippus 330

(monodactyl) in the Pliocene, connecting it to patterns of faunal change in the Northern Great Basin of 331


North America. In most faunas, including the Southern Great Basin, the monodactyl grazer Pliohippus is 332

found first in the middle Miocene (Clarendonian, approximately 13.6 to 10.3 million years ago), but in 333

the Northern Great Basin, it is not found until its immigration in the late Miocene and early Pliocene 334

(Hemphillian, approximately 10.3 to 4.9 million years ago). Shotwell connected this late appearance of 335

Pliohippus to the coeval spread of semi-arid plains and prairie grasslands from the Southern into the 336

Northern Great Basin, arguing that Pliohippus migration tracks this habitat. Similarly, he suggested that 337

Hipparion tracked woodland-savanna habitat, going locally extinct at the end-Hemphillian as savanna 338

habitats were reduced or eliminated but persisting longer where such habitats remained a major feature 339

of the landscape for a longer time (Shotwell 1961). To further support his claim of partitioning by 340

habitat, Shotwell noted that the relative abundance of hipparionines was not different in regions with or 341

without Pliohippus present, supporting the idea that the genera were not in direct competition. 342

In a recent study that aimed to investigate Shotwell’s (1961) hypothesis more quantitatively, we 343

tested for niche partitioning among different groups—in this case, tridactyl and monodactyl equids—344

using site occupancy (Parker et al. 2018). With approximately 3500 fossil horse occurrences that could 345

be assigned to a North American Land Mammal Age and to a paleohabitat (forest/swamp, forest, 346

woodland, woodland-savanna, savanna, grassland-savanna, or grassland), we tested whether tridactyl 347

and monodactyl genera were found in the same habitat type significantly less often than by random 348

chance (which would support habitat partitioning). In fact, overlap between these groups was higher 349

than expected by chance in all North American Land Mammal Ages except the Blancan, where it was 350

indistinguishable from random (Parker et al. 2018). Rather than partitioning by habitat, tridactyl and 351

monodactyl horses were found together more often than by chance. Consequently, at the spatial and 352

temporal scale examined, horses were more similar by shared ancestry than different by digit state, and 353

digit state did not correlate with hypsodonty or body size (Parker et al. 2018). These results suggest that 354

the three classic equid traits did not coevolve under a single, grassland-specific selective regime, but 355


rather were the product of multiple selective pressures that varied across the diverse habitats available 356

to equids in the Miocene and Pliocene. This lack of correlation or habitat partitioning points to the 357

conclusion that whatever drove digit reduction was not identical to the driver of other important equid 358

traits, so the cause of digit reduction was probably multifaceted (because, e.g., grasslands as the driving 359

factor of all three traits would likely lead to correlated evolution among them). 360

361

Insights from convergent evolution of digit reduction 362

Digit reduction is widespread in tetrapods, including theropod dinosaurs, marsupials, rodents, 363

squamates, and ungulates. Although some forms of digit reduction are arguably related to very different 364

drivers than in equids (e.g., in hopping, bipedal rodents), others may be more closely related. Some of 365

the most ecologically and morphologically convergent examples to horses are in the Artiodactyla, the 366

second major clade of North American ungulates (along with the Perissodactyla, in which Equidae is a 367

family). Many artiodactyl taxa evolved elongated limbs and reduced digits throughout the Cenozoic 368

(Janis and Wilhelm 1993), and they have been characterized as a competitor group to perissodactyls, 369

perhaps partially responsible for the decline of the perissodactyl order by outcompeting them (Illius and 370

Gordon 1992), although a qualitative comparison has found that competition leading to replacement 371

was not at play (Cifelli 1981). As with hypsodonty and limb evolution in horses, parallel arguments have 372

been made in artiodactyls for the evolutionary benefit of hypsodonty (DeMiguel et al. 2014) and the 373

unguligrade distal limb (Clifford 2010). However, recent work has shown that the small herbivorous 374

“condylarths,” which were replaced by artiodactyls and perissodactyls, also had cursorial species, 375

suggesting that cursoriality was not the only driver of success in ungulates (Gould 2017). 376

Developmental studies have also shown that digit reduction is accomplished via different 377

mechanisms in these two ungulate clades. For example, horses (Perissodactyla), camels (Artiodactyla), 378

and jerboas (Rodentia) form the beginnings of all five digits in early embryonic limb patterning but then 379


show a spike in cell death around the reduced digits in later post-patterning stages (Cooper et al. 2014). 380

In contrast, pigs (Artiodactyla) and cattle (Artiodactyla) developmentally reduce digits via restricted 381

expression of Ptch1, with no noticeable increase in cell death (Sears et al. 2011; Cooper et al. 2014; 382

Lopez-Rios et al. 2014). These results suggest that, developmentally at least, there may be more than 383

one way to evolve reduced digits, encouraging a comparative approach to further determine what 384

similarities and differences exist among convergent evolutions of digit reduction. 385

In convergence with the foot structure of early horses, two genera of caviomorph rodents, 386

Hydrochaeris and Cavia, have three toes on the hind foot (digits II, III, and IV) and in the forefoot have 387

eliminated digit I, reduced digit V to nonfunctionality, and evolved a digit-III-dominant foot (Rocha-388

Barbosa et al. 2007). Ground reaction forces and effective mechanical advantage of the limbs have been 389

characterized in capybara (Biewener 2005); they were found to have a less erect posture than goats, and 390

so may be a suitable comparison for basal equids, whose posture was not so erect as extant horses. 391

Even more striking convergence can be found in the South American Litopterna, an order 392

recently found to be sister to Perissodactyla (Buckley 2015; Welker et al. 2015; Westbury et al. 2017). 393

The litoptern genus Thoatherium evolved a monodactyl condition even more extreme than equids by 394

eliminating even the remnant “splint” metapodials II and IV. However, litopterns and horses show an 395

opposite relationship between digit reduction and body mass. Equids with the most reduced digits are 396

the largest species and, in general, less reduced digits are found on smaller species (McHorse et al. 397

2017). Conversely, the monodactyl litoptern species are the smallest and those with extremely robust 398

side digits are the largest (Janis 2007). This difference suggests that even if the body size hypothesis 399

remains plausible as a driver of digit reduction in horses, body size is likely not driving digit reduction in 400

litopterns. Despite these divergent body mass patterns, tracing the convergent evolution of digit 401

reduction in litopterns, together with studies of habitat and climate in South America (e.g., Strömberg et 402

al. 2013), has the potential to reveal whether other potential drivers are parallel in the two groups. 403


404

Future directions 405

The future of studying digit reduction in equids is promising. Here we lay out the steps we 406

believe are necessary to support, or reject, existing hypotheses, as well as new ideas for what may have 407

driven the evolution of such a remarkable trait as monodactyly. It is important to recognize that most of 408

the working hypotheses are not mutually exclusive, and as has become clear from previous work, we 409

should expect interrelationships between the potential causes of digit reduction (e.g., cooler climate 410

may affect body size directly, which is a hypothesized driver of digit reduction, but cooler climate also 411

leads to more open habitats, which is a different hypothesized driver). However, we argue that the way 412

that these selective drivers interact has a significant effect on how we conceptualize the “why” behind 413

digit reduction, and therefore it is a valuable endeavor to uncover the primary driver or drivers of 414

monodactyly and digit reduction as a whole. 415

416

Locomotion and biomechanics 417

If the primary driver of digit reduction was the need for better economy while covering long 418

distances, as hypothesized by Janis and Wilhelm (1993), then digit reduction should virtually always go 419

hand-in-hand with limb elongation. Research manipulating moment of inertia (MOI) in limbs has shown 420

that decreasing MOI does reduce cost of transport (Martin 1985; Myers and Steudel 1985; Wickler et al. 421

2004), and added distal limb mass increases cost of transport in extant horses (Wickler et al. 2004), so 422

some degree of energetic savings is almost certainly a consequence of digit reduction. The logical next 423

step is a theoretical exploration of the magnitude of energetic savings from 1) reduction of MOI at the 424

distal limb due to loss of digits and 2) elongation of the limb over 3) a range of body sizes and taxa. The 425

results from these calculations could be used to create a theoretical cost of transport morphospace that 426

connects changes in limb length, relative differences in segment MOI, estimated limb swing frequency, 427


and body mass to a resulting cost of transport. That morphospace could be used to calculate, e.g., how 428

much locomotor economy was improved between different taxa and whether the difference gained by 429

digit reduction constitutes a significant energetic savings. To evaluate significance of energetic savings, it 430

would be necessary to relate the results of the calculations to known costs associated with swinging legs 431

in living animals (e.g., Fedak et al. 1982). Tracking these changes through time would further allow 432

testing of whether quantitative improvements in locomotor economy over evolutionary time tracked (or 433

slightly lagged) aridification and the spread of grasslands. This analysis is ideally suited to include 434

artiodactyls as a comparative group, because they also evolved longer limbs, reduced digits, and would 435

have experienced the same pressures at the same times where they overlapped spatially with horses. 436

The idea that a tridactyl foot is more stable for lateral dodging on soft substrate, whereas a 437

more rigid single hoof is more stable for and provides faster straight-line locomotion on hard substrate 438

(Shotwell 1961), remains untested. The soft substrate hypothesis is concerned primarily with stability, 439

whereas the hard substrate may have considerably more to do with elastic energy storage and energy 440

dissipation. Whether theoretical or practical (as in biorobotics), this hypothesis requires a test of 441

whether tridactyl feet are indeed more stable on softer substrates. Examination of locomotor 442

performance in extant taxa with reduced digits would provide an interesting first step, which could be 443

complemented by studies using modeling and simulation to explore the effect of digit number on 444

stability in the equid distal limb; a combination of simulation and biorobotics would offer considerable 445

flexibility (e.g., Nyakatura et al. 2019). Biomechanical modeling work such as this can be more powerful 446

when ground-truthed with locomotion data, such as speed, joint kinematics, and forces from extant 447

animals—a challenge when the vast majority of equid diversity is extinct. Monodactyl equids are 448

straightforward to study in that domestic horses are anatomically extremely similar, particularly in the 449

distal limb, to both wild Equus and to other extinct monodactyl taxa. 450


Several taxa are potential modern analogs for tridactyl or semi-tetradactyl horses, and would 451

therefore be suitable for study of locomotor biomechanics (directly testing dodging stability on softer 452

substrates), for ground-truthing the proposed simulation and biorobotics work, and for comparative 453

studies of distal limb anatomy (including internal geometry, i.e., resistance to bending, torsion, and 454

compressive forces). Tapirs (Tapiridae) are one of three extant families in the perissodactyl order, the 455

others being equids and rhinoceroses, and are similar to basal equids in their digit state: four digits on 456

the front leg and three in the back. Tapirs may therefore offer a convenient semi-tetradactyl species for 457

locomotion studies of substrate-based stability; they have been used for comparative anatomical and 458

biomechanical studies (McHorse et al., 2017), and we have collected kinetics and kinematics data from 459

Baird’s tapir (Tapirus bairdii) that will provide locomotor forces to scale for future finite element 460

modeling work. A recent study has argued that the semiaquatic tapir benefits from lateral splaying in 461

phalanges II and IV, which could allow for greater stability on soft, muddy surfaces beneath water (Endo 462

et al. 2019), echoing the substrate stability hypothesis itself. 463

While tapirs provide the closest phylogenetic match for a study of digits and stability, they may 464

not provide the best biomechanical one. In contrast to equids, tapirs are semiaquatic (Nowak and 465

Paradiso 1999), and although some tapir species may provide morphological analogs for other basal 466

perissodactyls such as palaeotheres (MacLaren and Nauwelaerts 2019), the range of body sizes differs 467

considerably in the two families. Most basal equids had a body mass between 5 kg and 20 kg 468

(MacFadden 1986; Secord et al. 2012), but even the smallest extant tapir species have body masses over 469

130 kg (Nowak and Paradiso 1999). Therefore, caviomorph rodents and small artiodactyls may provide a 470

closer biomechanical analog in terms of the forces generated during locomotion. The two genera of 471

caviomorph rodents with remarkable convergence towards the equid foot condition are much closer in 472

size: species tend to average 30-60 kg and 1 kg in Hydrochaeris and Cavia respectively (Biewener 2005; 473

Ferraz et al. 2005). Similarly, some species of artiodactyl are quite small (e.g., tragulids and moschids are 474


generally less than 10 kg; Nowak and Paradiso 1999) and may provide a more suitable biomechanical 475

comparison than tapirs. Furthermore, extant artiodactyls are considerably more taxonomically diverse 476

than perissodactyls, and they underwent many similar changes in the evolution of unguligrady and 477

reduced digits—often argued to be an adaptation for fast and efficient locomotion (Clifford 2010). As 478

with all non-domesticated animals, the greatest challenge here may lie in access, and the most suitable 479

biomechanical comparison might require a compromise between the anatomy, preferred habitat, and 480

the availability (and behavioral temperament) of various species. 481

482

Macroevolutionary and regional analyses 483

While biomechanical studies can help untangle why different morphologies might be adaptive 484

for different substrates, macroevolution can give us perspective on how it happened. If grasslands were 485

the ultimate driver of monodactyly, we would expect genera with more reduced digits to be found more 486

often and/or to be more evolutionarily successful in open grassland-dominated habitats. Conversely, we 487

would expect genera that retained considerable side digits, such as hipparionines, to be found more 488

often in softer, forested environments and/or to be more successful there. Differential habitat use has 489

been shown in several taxa to provide diverging selective demands and thus to influence diversification 490

(Losos 2009; Collar et al. 2011; Price et al. 2011), although the groups in those studies (anoles, monitor 491

lizards, and labrid fishes) are all much smaller than equids. A recent spatially and temporally broad study 492

of habitat partitioning found no such evidence for tridactyl and monodactyl horses dividing habitats 493

where they overlapped (Parker et al. 2018). 494

Explicitly hypothesis-testing the niche partitioning aspect of Shotwell’s (1961) argument was an 495

important first step in tackling these ideas quantitatively, but the study by Parker et al. (2018) had some 496

limitations in scope. First, both temporal and spatial bins were coarse; North American Land Mammal 497

ages can be several million years long, which represents considerable time-averaging, and the spatial 498


resolution of habitat identification was typically at the county level. Since modern counties certainly 499

cover sufficient land to contain multiple habitat types, it is likely that fossil localities within the same 500

county could have had very different habitats. Second, the relative distribution of habitats in the study 501

was fairly constant through time, despite evidence that, on the whole, grasslands became the far more 502

dominant ecosystem. It is possible that the data analyzed by Parker et al. (2018) do not sufficiently 503

capture ecological reality, perhaps reflecting a bias in preservational environments that led to the 504

fossilization of horses. 505

Regional analyses may offer a solution to the limitations of spatial and temporal data, 506

particularly given how regionally-specific climate and habitat can be (Chen et al. 2015). We propose that 507

the Great Plains region of North America, which includes parts of several states including Montana, 508

Wyoming, North and South Dakota, and Nebraska, offers an ideal place to test more carefully the 509

relationship between horses, climate, and habitat (Figure 8). The horse fossil record is dense, 510

particularly in Nebraska (Figure 8; MioMap search for “Equidae” returns more than 1000 specimens, 511

Carrasco et al. 2005). The phytoliths (grass species indicators) and paleosols (C3 vs. C4 grass indicators) 512

are also well-characterized in the area (Figure 8; Fox and Koch 2003; Strömberg 2005, 2011). Other 513

potential candidate regions include the John Day region of Oregon and the state of Florida, both with 514

remarkable fossil records of horse evolution and extensive research into climatic and habitat change 515

through time (Stock 1946; Macfadden and Cerling 1996; MacFadden et al. 1999; Retallack 2004; 516

Maguire and Stigall 2008; Maguire 2015). However, there are tradeoffs to choosing regional-scale 517

studies: in exchange for better-controlled data and more power to detect trends locally, one gives up 518

some amount of power to explain global trends. In other words, trends at one scale cannot necessarily 519

be extrapolated to others—a critical challenge of macroevolutionary studies in general (Jablonski 2008). 520

Although qualitatively the trend appears to point toward monodactyly being ‘optimal,’ at least 521

in grasslands, this pattern has yet to be quantitatively tested—and as we know from work showing that 522


body size evolution was likely not directional in horses, apparent trends can be deceiving. Phylogenetic 523

comparative methods offer a way to explicitly test the evolutionary mode of trends like these. 524

Evolutionary model-fitting can compare the fit of models such as Brownian Motion (a random walk), an 525

Ornstein-Uhlenbeck (OU) process (a model where the trait is being pulled with some strength towards 526

an adaptive peak of some ‘optimal’ value), or a multi-peak OU model, which allows for multiple optima 527

that may correspond to another feature such as habitat (Hansen 1997; Butler and King 2004). In an in-528

progress study, we explicitly test digit reduction in this framework, investigating whether digit reduction 529

is pulled to some adaptive optimum for all of equids (i.e., some degree of digit reduction is “optimal”) or 530

whether that optimum varies based on habitat type (e.g., forest-dwelling species are pulled towards 531

some moderate value of TRI whereas grassland dwellers are pulled towards monodactyly). Alternatively, 532

if different habitats drive different rates of digit evolution but there is no trait optimum, a multi-rate 533

model would be more appropriate (Collar et al. 2010). A study such as this would also be suited to a 534

more broadly comparative context, evaluating whether the mode of digit reduction evolution is similar 535

in other taxa (e.g., artiodactyls or litopterns). 536

537

Conclusion 538

The evolution of monodactyly in horses is remarkable and is unique among extant animals, but 539

fortunately for scientists, the themes of digit reduction, habitat change, and body size change are 540

repeated many times in the fossil record. Reviewing the available evidence makes it clear that we are 541

unlikely to find a single evolutionary driver to be solely responsible for the evolution of monodactyly, 542

because open habitat, changes in substrate, changes in foot posture, and changes in body size can all tie 543

to one another and to broader ecological drivers such as changing climate. However, we argue that by 544

combining finer-scale regional studies, quantitative biomechanical studies, and careful analysis of 545

convergent clades, it will be possible to estimate the relative contributions of these evolutionary drivers. 546


Even if digit reduction is ultimately not driven by the same factors in each clade (as may be the case with 547

horses vs. litopterns), such a discovery would be a considerable leap forward in our understanding of 548

how—and why—horses evolved a single toe. 549

550

Funding 551

This work was supported by the National Science Foundation [DGE-1144152 to B.K.M, DEB-552

1701656 to B.K.M and S.E.P]. 553

554

Acknowledgments 555

The authors would like to thank Zachary Morris and other members of the Pierce and Biewener 556

labs for productive discussion; Talia Moore for help conceptualizing Figure 6; Samantha Hopkins and 557

Edward Davis, who first encouraged the equid evolutionary line of thinking and have provided ongoing 558

thoughts; and Abigail Parker and Tristan Reinecke, whose work contributed to ideas mentioned in this 559

review. Hayley O’Brien suggested looking into caviomorph rodents. Zhijie Jack Tseng and an anonymous 560

reviewer provided helpful comments that improved the manuscript. Finally, the authors would like to 561

thank Samantha Price and Martha Muñoz, who organized the symposium on Biomechanics in the Era of 562

Big Data, to which this paper is a contribution. 563

564

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Figure Captions 826

Figure 1. The anatomy of modern Equus metapodials; proximal articular views are of the metacarpal (A) 827

and metatarsal (B), and the metacarpal and phalanges are shown in anterior (C) and posterior (D) views. 828

Abbreviations: digits II, III, and IV are shown for the metacarpal and metatarsal; PP is proximal phalanx; 829

MP is medial phalanx; DP is distal phalanx. Sesamoids are indicated with lines. 830

Figure 2. The linear progression of horses (small to large, many toes to one toe, low-crowned teeth to 831

high-crowned teeth), a view that dominated early narratives about equid evolution. Modified from 832

Matthew (1926). 833


Figure 3. A simplified cladogram of horse genera with tapir as an outgroup. Topology after Froehlich 834

(2002), Fraser et al. (2015), Jones (2016), and Bai et al. (2018). Subclades are highlighted by color and 835

are after Famoso and Davis (2014) and Cantalapiedra et al. (2017), but are frequently paraphyletic (e.g., 836

the Merychippus-Grade Equinae). Note that “Merychippus” is a known polyphyletic group, and here we 837

include only one of several phylogenetic positions for taxa called Merychippus; see Fraser et al. (2015). 838

Digit state is shown by lines (solid black for semi-tetradactyl, thin gray for tridactyl, dotted black for 839

monodactyl). Size is indicated by a circle, scaled based on the base-10 logarithm of body mass (where 840

available). Extant taxa are marked with an asterisk. 841

Figure 4. An illustration of the measurements used to calculate TRI. First the lengths of the proximal side 842

phalanges (PPlengthIV and PPlengthII) are averaged, then this value is divided by the length of the 843

proximal center phalanx (PPlengthIII). Illustration modified from Matthew (1926). 844

Figure 5. A phylogenetic tree of some horse genera showing discrete categories of digit state (left) vs. 845

Toe Reduction Index (TRI), a continuous measure of digit reduction (right). TRI captures considerable 846

variation within tridactyl horses that is missed by discrete categories. Modified from Parker et al. (2018). 847

Figure 6. Morphology interacts with the environment to create a given performance, which then 848

(modulated by competition) determines fitness in that environment. Selection acts according to fitness, 849

driving evolutionary change in morphology. 850

Figure 7. Global temperature through time, with significant biotic and abiotic events highlighted. 851

Temperature, climatic event, and ice sheet data from Zachos et al. (2001); equid data from Bai et al. 852

(2018), MacFadden and Hulbert (1988), and Janis (2007), with equid and litoptern data from MacFadden 853

(1994); grassland and hypsodonty data from Strömberg (2005, 2011); pursuit predator data from Janis 854

and Wilhelm (1993). Ages indicated by annotations are approximate, and in many cases (e.g., the spread 855

of grasslands) are ± several million years. Note that oxygen isotope to degrees Celsius relationships are 856


calculated for an ice-free ocean, so temperature estimates are only valid until approximately 35 Ma 857

(Zachos et al. 2001). 858

Figure 8. Equid occurrences (orange, Paleocene, Eocene and Oligocene; yellow, Miocene, Pliocene, and 859

Pleistocene) across a section of North America. Size of the circle is scaled to number of occurrences. The 860

densely sampled Great Plains region is highlighted in pale orange. Sites characterized for C3 vs. C4 861

grasses (Fox and Koch 2003) are shown by magenta circles; sites characterized for grassland indicator 862

phytoliths (Strömberg 2005, 2011) are shown by blue and purple circles. 863

The evolution of a single toe in horses - NSF PAR

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