Bone Growth Strategies and Skeletochronological Age Estimates of Desert Tortoise

(Gopherus agassizii) Populations

A Thesis

Submitted to the Faculty

Drexel University

Amanda Jane Curtin

in partial fulfillment of the

requirements for the degree

Doctor of Philosophy

May 2006

Dedications

For my grandparents, Peggy Lillian and David Croll Pollock and Crystal Pamela and

Harold Desmond Curtin who always nurtured and encouraged my inquisitive nature and gave me a strong sense of my place in this world.

In special remembrance of my grandmother, Peggy, who inspired my love of nature and also my grandfather, Harold, who inspired my love of knowledge and history....I would not have come this far without the determination and imagination that they fueled in me.

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Acknowledgements

Special thanks to my advisor, Dr. Jim Spotila, for his friendship, support and encouragement. I’d also like to express my gratitude and appreciation to the other members of my committee: Dr Hal Avery (I could not have asked for a better teacher on desert ecology), Dr. Mike O’Connor, Dr. Peter Dodson and lastly Dr. George Zug, whose guidance, encouragement and friendship meant more than words can express. Thanks to the following people and institutions for donating carcasses used in this study: Dr. Philip

Medica and the United States Geological Service (USGS), Dr. Roy Averill-Murray

(Arizona Fish and Wildlife Service), Dr. William Boarman (USGS), the Desert Tortoise

Conservation Center (Clark County, Nevada), and the Mojave National Preserve,

California. To all the undergraduate and graduate students from Drexel University who helped with carcass collection and histological processing, I am most grateful. Also thanks to the Bioscience staff and faculty at Drexel for their support and encouragement, especially Dr. Laura Duwel, Dr. Jeremy Lee, Ms. Gerry Marekova and Ms. Brenda

Jones-Bowden. Lastly to my friends and family, especially my mother, Louise, and step- father, George Loudon - without your love, strength and encouragement I would not have been able to achieve this dream. This research was funded by the Panaphil Foundation and Betz Chair Endowment.

Table of Contents

LIST OF TABLES……………………………………………………………………. viii

LIST OF FIGURES………………………………………………………………….. xi

ABSTRACT………………………………………………………………………….. xvi

1. GENERAL INTRODUCTION…………………………………………………... 1

The desert tortoise (Gopherus agassizii)..……………………………….……… 2

Skeletochronology….…….…………………………………………..…...... 4

General methodology…………………….………………….……………...... 6

Ranking protocol……...... 7

Correction factor………..…………………………………….……….….. 9

General problems and considerations………………………………………… 9

2. CHAPTER 2. KNOWN-AGED DESERT TORTOISES (GOPHERUS AGASSIZII) FROM ROCK VALLEY, NEVADA VALIDATE SKELETOCHRONOLOGICAL ANALYSIS...…………………………………………………………………….. 13

Introduction…………………………………………………………………... 13

Materials and methods……………………………………………………….. 15

Skeletochronology...... 17

Results...... 18

Age and longevity...... 18

Age at sexual maturity...... 19

Discussion...... 19

Age and longevity...... 19

Age at sexual maturity..…………………………………………………... 21

Conclusions and implications for future research……....……………….……… 23

3. AGE AND GROWTH STRATEGIES OF SONORAN DESERT AND WEST MOJAVE DESERT TORTOISES (GOPHERUS AGASSIZII)...... 40

Introduction...... 40

West Mojave Desert Tortoises...... 43

Sonoran Desert Tortoises...... 44

Materials and methods...... 46

Materials...... 46

Skeletochronology...... 46

Statistical analysis...... 47

Results...... 48

Body size...... 48

Bone morphometry...... 49

Resorption core diameters...... 51

Age and longevity...... 52

Age at sexual maturity...... 54

Discussion...... 55

Body size and bone morphometry...... 55

Resorption core diameters...... 59

Longevity and age at maturity...... 62

Growth strategies, ecological implications and conclusions...... 63

4. AGE ESTIMATES AND GROWTH STRATEGIES IN EASTERN MOJAVE DESERT TORTOISE (GOPHERUS AGASSIZII) POPULATIONS...... 101

Introduction...... 101

Materials and methods...... 102

Results...... 104

Body size...... 104

Bone morphometry...... 105

Resorption core diameters...... 105

Age and longevity...... 106

Age at sexual maturity...... 107

Discussion...... 107

Body size and bone morphometry...... 107

Growth strategies, ecological implications and conclusions...... 109

5. CHAPTER 5. GROWTH STRATEGIES IN DESERT TORTOISES POPULATIONS: ECOLOGICAL IMPLICATIONS AND FUTURE TRENDS...... 142

Introduction...... 142

Materials and methods...... 146

Results...... 147

Discussion...... 150

Size, growth and productivity...... 150

Ecological implications and conclusions...... 155

BIBLIOGRAPHY...... 167

VITA...... 180

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LIST OF TABLES

1. Shell measurements for Gopherus agassizii carcasses from the East Mojave Desert, Nevada……………………………………………………………………………… 25

2. Bone measurements for Gopherus agassizii carcasses from the East Mojave Desert, Nevada; CL = carapace length, H = humerus, F = femur, I = ilium, S = scapula, J = juvenile, F = female, M = male…………………………………………….……… 27

3. Periosteum diameter (PD) and growth layer (GL) ranking protocol and Correction Factor (CF) age estimates for Gopherus agassizii from the East Mojave Desert (Nevada); J = juvenile, F = female, M = male…………………………………….. 29

4. Maximum age estimates for Gopherus agassizii from the East Mojave Desert (Nevada) using the periosteum diameter (PD) and growth layer (GL) ranking protocol and correction factor (CF) skeletochronology methods. Age estimates in bold are equal or closest to the known age (where available). The value in brackets is the difference between the age estimate and known age; CL = carapace length……… 32

5. Student’s t-test analysis and mean difference (+ one standard error [SE] of the mean) between known age versus skeletochronology age estimates in Gopherus agassizii from Rock Valley, Nevada. Mean squared error (MSE) and root mean squared error (RMSE) are given to show which aging method provided ages with the least error and bias to known ages; PD = periosteum diameter, GL = growth layers, CF = correction factor……………………………………………………………………………..... 34

6. Rainfall comparisons between Goffs, California and Rock Valley, Nevada during identical time periods between 1982 and 1986. The (+) accentuates which location was greater during the time period measured………………………………...…… 35

7. Population samples of desert tortoises (Gopherus agassizii) from the Sonoran Desert, Arizona……………………………………………………………………………. 67

8. Location, elevation and habitat type for populations of Gopherus agassizii from Arizona. NSD = northern Sonoran Desert; SSD = southern Sonoran Desert……. 69

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9. Analysis of variance in bone length between adult Gopherus agassizii from the Sonoran Desert, Arizona and West Mojave Desert, California. df = degrees of freedom………………………………………………………………...…………. 70

10. Details of regression lines for carapace length (CL) versus total bone length (TBL) in Gopherus agassizii from the Sonoran Desert, Arizona and the West Mojave Desert, California…………………………………………………………………………. 71

11. Regression statistics of carapace length (CL) versus resorption core diameter (RCD) for Sonoran Desert and West Mojave Desert tortoises…………….…………….. 72

12. Age estimation for Sonoran Desert Gopherus agassizii using the Correction Factor (CF) method and Ranking Protocol (RP) method. J = juvenile; F = female; M = male………………………………………………………………………………. 73

13. Age estimation for West Mojave Gopherus agassizii using the Ranking Protocol (RP) and Correction Factor (CF) method……………………………………………… 76

14. Size and age estimates of Gopherus agassizii from the Sonoran Desert, Arizona determined by the Correction Factor (CF) method and Ranking Protocol (RP). Age in bold represents the more accurate age estimate; CL = carapace length; CW = carapace width; PL = plastron length; J = juvenile, M = male; F = female; Unk = unknown sex………………………………………………………………………...……… 79

15. Size and age estimates of Gopherus agassizii from West Mojave Desert, California determined by the Correction Factor (CF) method and Ranking Protocol (RP). Age in bold represents the more accurate age estimate; CL = carapace length, PL = plastron length…………………………………………………………………………….. 82

16. Analysis of variance in age within size classes between Gopherus agassizii from the Sonoran Desert, Arizona and West Mojave Desert, California; CL = carapace length. Values in bold represent a statistically significant difference between size classes from the two localities…………………………………………………………… 85

17. Shell measurements of Gopherus agassizii from two localities in the East Mojave Desert: the Desert Tortoise Conservation Center (DTCC) in Nevada and Ivanpah Valley, Mojave National Preserve (MNP) in California; CL = carapace length, CW = carapace width, PL = plastron length……………..…………………………….. 112

18. Bone length and width measurements for Gopherus agassizii from the East Mojave Desert (CA and NV); J = juvenile, F = female, M = male; Unk = unknown sex; H = humerus, I = ilium, F = femur, S = scapula……………………………………… 114

19. Analysis of variance (ANOVA) in proximal, midshaft and distal bone width between adult male and female Gopherus agassizii from the East Mojave Desert, CA and NV……………………………………………………………………………….. 117

20. Regression statistics of carapace length (CL) versus resorption core diameter (RCD) in juvenile and adult Gopherus agassizii from the East Mojave Desert (CA and NV)………………………………………………………………………………. 118

21. Skeletochronological age estimates for Gopherus agassizii from the East Mojave Desert (CA and NV) using the Correction Factor (CF) and Ranking Protocol (RP) methods; CL = carapace length; J = juvenile, F = female, M = male; Unk = unknown sex……………………………….……………………………………………….. 119

22. Maximum skeletochronological age estimates for Gopherus agassizii from the East Mojave Desert (CA and NV) determined by the Correction Factor (CF) and Ranking Protocol (RP) methods; CL = carapace length (mm); unk = unknown sex……… 122

23. Regression statistics for age versus carapace length (CL) in juvenile and adult Gopherus agassizii from the East Mojave Desert (CA and NV); CL = carapace length. Values in bold are statistically significant……………………………….. 125

24. Summary of life-history traits and annual growth rates for Gopherus agassizii from the West and East Mojave Desert in California and southern Nevada and from the Sonoran Desert in Arizona………………………………………………………. 160

LIST OF FIGURES

1. Diagrammatic representation of a cross section through the mid-shaft area of a long bone. Arrows represent diameter measurements. The initial solid arrow represents the resorption core diameter and subsequent dashed arrows represent complete growth layer (GL) diameters. The initial two GLs are partially resorbed and diameters are this not available. All diameters were recorded along the dorsal-ventral plane, but only where the entire circumference of a growth layer was visible; mca = medullary cavity area……………………………………………………………… 12

2. A comparison of skeletochronology age estimates with known-ages in Gopherus agassizii from Rock Valley, Nevada; PD = periosteum diameter, GL = growth layers, CF = correction factor…………………………………………………………….. 36

3. Box plot of skeletochronology age estimates and known age of Gopherus agasizzii from Rock Valley, Nevada; PD = periosteum diameter, GL = growth layers, CF = correction factor. Vertical lines within the box plots represent the median age and whiskers the 10th and 90th percentile………………………….………………….. 37

4. Regression of known age versus skeletochronology age estimates in Gopherus agassizii from Rock Valley, Nevada; PD = periosteum diameter (solid line), GL = growth layers (dashed line), CF = correction factor (dotted line)………….…….. 38

5. Regression of age versus carapace length (CL) in Gopherus agassizii from Rock Valley, Nevada; PD = periosteum diameter, GL = growth layers, CF = correction factor………………...……………………………………………………………. 39 .

6. Regression of carapace length (CL) versus carapace width (CW) and plastron length (PL) in adult male and female Gopherus agassizii from A) the Sonoran Desert, Arizona and B) the west Mojave Desert, California. Solid lines = adult females, broken lines = adult males……………………………………………...………… 86

7. Shell measurements of adult male and female Gopherus agassizii from A) the Sonoran Desert, Arizona and B) the west Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers……………………………………………….. 87

8. Regression of carapace length (CL) versus carapace width (CW) and plastron length (PL) in A) adult female and B) adult male Gopherus agassizii from the Sonoran Desert (SD), Arizona and the west Mojave (WM) Desert, California. Solid lines represent Sonoran Desert tortoises, broken lines represent West Mojave tortoises. 88

9. Regression of carapace length versus carapace width (CW) and plastron length (PL) in juvenile Gopherus agassizii from the Sonoran Desert (SD), Arizona and the west Mojave (WM) Desert, California. Solid lines represents SD, broken lines represent WM adults………….…………………………………………………………….. 89

10. Bone lengths for adult male (M) and female (F) Gopherus agassizii from A) the Sonoran Desert, Arizona and B) the west Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers. Numbers above box plots represent sample sizes……………………………………………………………………………….. 90

11. Bone widths for adult female Gopherus agassizii from the Sonoran Desert (SD), Arizona and west Mojave Desert (WM), California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers………………………………………………………………….. 91

12. Bone widths for adult male Gopherus agassizii from the Sonoran Desert, Arizona and west Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers…………………………………………………………………………….. 93

13. Resorption core diameters for adult A) female and B) male Gopherus agassizii from the Sonoran Desert, Arizona and the West Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers…………………..……………………….. 95

14. Regression of carapace length versus resorption core diameters in A) humeri and B) ilia for Gopherus agassizii from the Sonoran Desert, Arizona (AZ) and the West Mojave (WM) Desert, California. Solid lines represent the SD sample and dashed lines WM sample……………………………………………..…………………… 96

15. Correction Factor age estimates for A) juvenile and Ranking Protocol age estimates for B) adult Gopherus agassizii from the Sonoran Desert (SD), Arizona and the west Mojave (WM) Desert, California. Horizontal lines within each box represent the

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median value and whiskers represent the 10th and 90th percentile. Dots represent outliers………………………………………………..…………………………… 97

16. Regression of skeletochronology age estimates versus carapace length for Gopherus agassizii from the Sonoran Desert, Arizona (solid line) and the West Mojave Desert, California (dashed line). Fine dotted line represents estimated age of sexual maturity at ~180 mm CL based on the regression lines for both populations……….……… 98

17. Regression of Ranking Protocol age estimates versus carapace length for adult female and adult male Gopherus agassizii from the A) Sonoran Desert (SD), Arizona and the B) West Mojave (WM) Desert, California……………………….……………….. 99

18. Box plots of age range within size classes for Gopherus agassizii from the Sonoran Desert (SD), Arizona and the West Mojave (WM) Desert, California. Horizontal lines within the box plots represent the median age and whiskers the 10th and 90th percentile. Outliers are represented by black dots………………...... …………… 100

19. Regression of carapace length (CL) vs carapace width (CW) and plastron length (PL) for Gopherus agassizii from the Desert Tortoise Conservation Center (DTCC; solid lines), Nevada (NV) and the Mojave National Preserve (MNP; dashed lines), California (CA)…………………………………………………………………… 126

20. Carapace length (CL) vs carapace width (CW) and plastron length (PL) in adult male (dashed lines) and female (solid lines) Gopherus agassizii from the East Mojave Desert (CA and NV)……………………………………………………………… 127

21. Shell size of A) adult female and B) male Gopherus agassizii from the East Mojave Desert (CA and NV). Males are significantly larger than females for all shell measurements (p < 0.05). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers..…… 128

22. Bone length for adult male and female Gopherus agassizii from the East Mojave Desert (CA and NV). In all cases males had significantly longer bones than females (p < 0.05). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers…………………… 129

23. Regression of carapace length versus bone length in Gopherus agassizii from the East Mojave Desert (CA and NV)…………………………………………………….. 130

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24. Bone widths of adult male and female Gopherus agassizii from the East Mojave Desert (NV and CA). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers………… 131

25. Resorption core diameters for adult Gopherus agassizii from the East Mojave Desert (CA and NV). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers….……… 133

26. Regression of carapace length versus resorption core diameter for Gopherus agassizii from the East Mojave Desert (CA and NV)……………………………………… 134

27. Age estimates adult male and female Gopherus agassizii from the East Mojave Desert (CA and NV). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers………… 136

28. Regression of age versus body size (carapace length) in Gopherus agassizii from the East Mojave Desert (CA and NV)……………………………………………….. 137

29. Regression of age versus size in adult male and female Gopherus agassizii from the East Mojave Desert (CA and NV)……………………………………………….. 138

30. Age range within the size classes of Gopherus agassizii from the East Mojave Desert (CA and NV). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers….……… 139

31. Regression of size (plastron length) versus age for East Mojave Desert tortoises (Gopherus agassizii) from Rock Valley, Nevada (data from Table 1; Turner et al., 1987), Clark County, Nevada and Ivanpah Valley, Mojave National Preserve, California………………………………………………………….……………… 140

32. Average annual rainfall for the lower and higher elevation study plots (800 – 1100 m) in Ivanpah Valley, Mojave National Preserve, California (Avery, unpubl. data) between 1997 and 2003…………………………………………………………... 141

33. Carapace length (CL), carapace width (CW) and plastron length (PL) of A) adult female and B) adult male Gopherus agassizii from the West and East Mojave Desert

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(California and Nevada) and Sonoran Desert (SD; Arizona); WM = West Mojave, EM = East Mojave……………………..………………………………………… 162

34. Age versus body size in Gopherus agassizii from the West Mojave Desert, California (long dashed line), East Mojave Desert, California and Nevada (short dashed line) and Sonoran Desert, Arizona (solid line)………………………………………… 163

35. Age versus body size in juvenile Gopherus agassizii from the West Mojave Desert, California (long dashed line), East Mojave Desert, California and Nevada (short dashed line) and Sonoran Desert, Arizona (solid line)…………………………… 164

36. Age versus body size in A) adult female and b) adult male Gopherus agassizii from the West Mojave Desert, California (long dashed line), East Mojave Desert, California and Nevada (short dashed line) and Sonoran Desert, Arizona (solid line)…………………………………………………………………….………… 165

37. Box plot of annual growth rates of juvenile and adult Gopherus agassizii from the Sonoran Desert (SD) in Arizona and West (WM) and East Mojave (EM) Desert in California and Nevada. Horizontal lines represent median values, whiskers represent the 5th and 95th percentile and dots represent outliers…………………………… 166

Abstract Bone Growth Strategies and Skeletochronological Age Estimates of Desert Tortoise (Gopherus agassizii) Populations Dr. James R. Spotila

Turtles are among the longest-lived vertebrates and concepts related to aging are important because they may have a substantial impact on life history evolution. Age at sexual maturity and longevity are essential baseline data which all demographic studies need for the conservation of extant species. The desert tortoise (Gopherus agassizii) is a threatened species and seems to be long-lived, although population longevity is mostly unknown. Skeletal growth layers can be used to estimate age, and skeletochronology has been shown to accurately age many reptiles. My main objective was to determine whether bone, obtained from carcasses, could be used to indicate differences in longevity and growth strategies among wild desert tortoise populations. I used two skeletochronological methods (the Correction Factor and Ranking Protocol) to estimate ages for tortoises from the Mojave and Sonoran Deserts. Using a sample of 9 known- aged tortoises from Rock Valley, Nevada, I determined that the Correction Factor method provided the most accurate age estimates for juveniles, whereas the Ranking Protocol provided the most accurate age estimates for adults. West Mojave tortoises had the smallest females, highest growth rates, youngest age at sexual maturity but shortest longevity of the three populations. In contrast, Sonoran tortoises had the largest females, lowest growth rates, oldest age at sexual maturity and greatest longevity. East Mojave tortoises showed intermediate traits. Annual growth rates in West Mojave adults were similar to those of juveniles, whereas East Mojave, and especially Sonoran adults had significantly reduced growth rates after the attainment of sexual maturity. West Mojave

xvi tortoises may have higher overall growth because of the higher desert productivity and more abundant forage after wet years. The cost for Mojave Desert tortoises (especially western populations), without dependable and predictable annual forage and water for health and growth maintenance, may be higher mortality and reduced life expectancy, which females have compensated for by evolving a younger age at sexual maturity.

Desert tortoises are potential models for the consequences of increased desertification on desert fauna, especially in species that have not evolved as desert specialists but rather have learned to adapt to these extreme environments.

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CHAPTER 1. GENERAL INTRODUCTION

The desert tortoise (Gopherus agassizii)

The four species of living tortoises (genus Gopherus) in North America are mostly restricted to deserts, arid lands and southeastern coastal regions (Bury and

Germano, 1994). They attract much attention because of their distinct ecologies as arid- or xeric-adapted species and because they play ecological roles as keystone species (Bury and Germano, 1994). Species of Gopherus appear long-lived, but the actual average and maximum longevities of these species are not known (Germano, 1994). The desert tortoise (Gopherus agassizii) occupies the largest range, both geographically and ecologically, of the four Gopherus species (Germano, 1994). It occurs in valleys and bajadas of Mojave Desert scrub in the northern part of its range, on rocky hillsides of

Sonoran upland vegetation in the center of its range, and on hills covered by Sinoloan thorn scrub and deciduous woodland in the south (Germano et al., 1994).

The desert tortoise is listed as a threatened species due to a trend of declining population size in many portions of its range (Fish and Wildlife, 1994). Acquisition and analysis of demographic data in species with small or declining populations are challenging under the best of circumstances (Berry, 2002). As a group, chelonians exhibit great longevity, and few long term studies have encompassed even one generation

(Swingland and Klemens, 1989; Berry, 2002). Absolute ages are extremely difficult if not impossible to ascertain, especially as neonates and juveniles are generally difficult to sample during the first few years of life (Berry and Turner, 1986; Morafka, 1994). For rare, threatened and endangered chelonians, comprehensive demographic data bases are

2 vital for developing the life tables and population viability analyses that are an essential part of recovery programs (Berry, 2002).

Gopherus agassizii spends much of its life in underground burrows used as general shelter, as hibernacula, as thermal refugia from daily and seasonal temperature fluctuations and as nesting sites (Bulova, 1994; U.S. Fish and Wildlife Service, 1994;

Spotila et al., 1994). Desert tortoises emerge to feed and mate during the fall (following nesting) and early spring (prior to nesting) and remain active throughout the spring and sometimes after summer thunderstorms (U.S. Fish and Wildlife Service, 1994; Rostal et al., 1994). Desert tortoises eat a wide variety of herbaceous vegetation, particularly grasses and flowers of annual plants (U.S. Fish and Wildlife Service, 1994). Tortoises lay on average 1-2 clutches per year with a mean clutch size of 4.5 to 6.7 eggs, depending on the region (Turner et al., 1986; Rostal et al., 1994). Adults are, for the most part, well protected against predators and are consequently long lived (Turner et al., 1986;

Germano, 1992; 1994). Their longevity helps compensate for their variable annual reproductive success, which tends to correlate with environmental conditions (U.S. Fish and Wildlife Service, 1994). However, even when adult survivorship is “normal”

(approximately 98 % per year), populations are not capable of rapid growth and, under reasonably favorable conditions, might be able to grow at an average rate of 1 % per year

(U.S. Fish and Wildlife Service, 1994).

Longevity estimates for wild G. agassizii are 32 years (western Mojave Desert),

48-52 years (eastern Mojave Desert), 35 years (Sonoran Desert) and 40-60 years

(Sinaloa). These are, however, minimum values because of the uncertainty of the accuracy of the estimates (Germano, 1992; Germano, 1994). In general, maximum body

3 size varies geographically but does not show clinal variation. The largest tortoises seem to be found at both ends (western Mojave Desert and Sinaloa, Mexico) and the middle of the distribution (northern Sonoran Desert), with smaller tortoises distributed in between

(eastern and southern Sonoran Deserts).

Differences in geographic habitats of desert tortoises allow for the testing of how the environment affects growth and, therefore, ultimately the life-history traits of long- lived species (Germano, 1994). My goal was to determine whether bone, specifically bone from dead animals, could be used to indicate differences in longevity and growth strategies among wild desert tortoise populations. Studies on growth rates in wild desert tortoises have been evaluated using annual body size measurements, scute rings and body mass as parameters (see Medica et al., 1975; Turner et al., 1987; Germano, 1988; 1994).

The absence of data from different populations of desert tortoises prevents the examination of inter-regional variation in growth rates and maturity. Moreover, the different methodologies employed in each study confound the comparison of results. I wished to unify and interpret population data using a common methodology that would allow for comparisons about tortoise biology across a broad geographic scope. I wanted to investigate whether variation in growth rate could be determined via skeletal morphology. Ecological constraints (like diet and climate) are known to influence bone growth strategies, thus life history strategies, in a wide variety of amphibians and reptiles.

Although longevity studies involving skeletal analysis (called skeletochronology) have been conducted on sea turtles, few studies have concentrated on tortoises. Gopherus agassizzi represented a perfect species to investigate, as they are long-lived tortoises, living in a habitat where their survival is dependent on resource availability and quality,

4 which is directly or indirectly influenced by environmental factors (mainly rainfall).

This study attempts to answer questions about tortoise biology by incorporating multiple populations of wild desert tortoises and addresses these questions without harming, killing or disturbing living animals of this threatened species.

Skeletochronology

Ectothermic tetrapods display cyclical growth patterns in their skeletons. This cyclicity produces growth marks (Castanet and Smirina, 1990; Castanet et al., 1993). For poikilothermic species living in temperate climates, thermal seasonality is the main factor that directly influences the yearly cyclical growth rhythm. Growth marks (GM) are also found in homeothermic animals and in poikilotherms living in more or less constant climatic conditions throughout the year, although they are less distinct (Castanet et al.,

1993). The current opinion, therefore, is that bone growth variations and associated GM are ultimately caused by an internal (genetically based) rhythm which, under natural conditions, becomes synchronized with, and reinforced by, seasonal cycles, even if these cycles have a low amplitude ((Meunier et al, 1979; Castanet et al., 1993).

This annual cyclical growth pattern provides information that has been used in many studies to accurately age amphibians and reptiles (e.g., Castanet and Cheylan,

1979; Zug et al., 1986; Germano, 1988; Castanet and Baez, 1991; Tucker, 1997; Zug and

Glor, 1998; Coles et al., 2001). In cyclical growth, broad zones are associated with periods of rapid growth (osteogenesis), whereas narrow zones or annuli (avascular regions) are associated with periods of decreased growth rates (Castanet et al., 1993;

Wake and Castanet, 1995; Esteban et al., 1996). These latter periods can also manifest as

5 lines of arrested growth (LAGs), which are functionally defined as rest lines (Castanet et al., 1993). Zones are formed during favorable seasons, marked by comparative warmth, moisture and food availability, whereas annuli and/or LAGs are formed during periods that are comparatively stressful (Castanet and Smirina, 1990; Castanet et al., 1993). A zone plus an annulus and/or LAG equals one growth cycle (usually a one year period).

Growth rings or layers theoretically can be used to age individuals, and this technique is called skeletochronology.

Bone remodeling is fundamental to the general process of skeletal growth and is concerned with the continual reshaping of a bone during growth, as it increases in size

(Enlow, 1963). Because bone is hard, it cannot grow and increase in mass by the internal expansion of existing tissues, as do most soft tissues. It can only grow by an appositional process of surface deposition together with resorption from a contralateral surface

(Enlow, 1963). At the medullary cavity margin, endosteal resorption can also remove initial growth layers. In other words, the earliest growth layers deposited can disappear and cause an underestimated age (Castanet and Smirina, 1990). The rate of periosteal bone resorption can be evaluated by using either known-age individuals or by a back- calculation method (Castanet and Smirina, 1990).

Compared to similar aging methods such as scalimetry or otolithometry, investigations of bone tissues have some specific advantages (Castanet et al., 1993). For instance, the functional interpretation of the various histological patterns of bone tissues allows construction of detailed hypotheses about the growth dynamics of bones and whole skeletons, and these data are extrapolated to the growth sequence of an individual

(Castanet et al., 1993). By using bone growth marks as well as other progressive

6 transformations of bone structures closely related to the time flow, skeletal tissues now appear to be an essential chronological tool in many fields of research (Castanet et al.,

1993).

General methodology

I obtained the humerus, femur, ilium and scapula (or any of these bones) from each tortoise carcass. I determined that these bones showed the most growth layers from a skeletal screening process in which I sectioned all appendicular skeletal elements as well as vertebrae, gulars and marginal, neural and plaston bones. I chose to include the ilia and scapulae because in the wild, the limbs and skull of tortoise carcasses have usually been removed by predators and/or scavengers, and the pectoral and/or pelvic bones are usually the only remaining elements. Initially, I cut serial sections of the entire bone to determine which area showed the most primary periosteal bone (and thus the least amount of remodeling). I then sectioned limited areas throughout all bones used for this aging study to obtain bone tissue data, but I concentrated on the midshaft region because this area showed the greatest number of unresorbed growth marks (Fig. 1). In the midshaft area, I measured all diameters along the dorsoventral plane because, although the lateromedial plane showed the largest diameters, remodeling around the resorption core was more intense in these two areas as opposed to the dorsal and lateral areas (Fig. 1). I chose two methods of estimating age by skeletochronology, the Ranking

Protocol and Correction Factor method; I will discuss these in detail in the following sections.

Ranking protocol

Criteria for this aging method are set out in Parham and Zug (1997) and Zug (1990).

I recorded the diameters of each growth mark (in smaller individuals), resorption core and bone cross section (periosteum diameter) at 10X magnification using a stage micrometer and transmitting light microscope. I defined the resorption core as the medullary cavity (usually filled with spongy bone) plus any endosteal bone (primary and/or secondary bone deposited from the endosteum inward; Fig. 1). The steps of the protocol evaluate these diameters and make assignments to growth-cycle classes in the following manner:

1.) Rank the bone sections in order of increasing resorption core diameters.

2.) Starting with the section with the smallest core, assign the innermost (smallest)

growth layer (GL) to the earliest growth class possessing an appropriate range of the

GLs diameters.

3.) The succeeding (outer) GL diameters are placed in successive classes. No classes

are skipped.

4.) Each growth cycle presumably equals one year.

Individuals are ranked from smallest resorption core diameter to largest, using hatchling resorption cores and total bone diameters as an indicator of pre-growth expression state.

In this way, successive GL diameters are recorded into a ranking class/age class and, for at least the initial years, it is possible determine a range to assign a certain age.

I estimated ranking age in two ways. I labeled the method discussed above as the growth layer (GL) ranking protocol. As an alternate method (which I have called the periosteum diameter (PD) ranking protocol), I used the age classes and corresponding

8 diameter ranges from the GL ranking protocol but, instead of assigning visible growth layers into age classes, I used the periosteum diameter (diameter of outer cross section perimeter) to represent the last growth layer, which I then placed into the appropriate age class. This method does not take the number of visible GL into account but assumes that the periosteum diameter represents that animal’s age at death; i.e., the periosteum represents the last GL. The main difference between these two methods is that the GL ranking protocol estimates ages as resorbed GL added to the number of visible growth layers (does not include the periosteum diameter), whereas the PD ranking protocol estimates age from the periosteum diameter which in theory represents the age at death

(or last growth layer). I chose to include the PD as an age estimate because I wanted to see how this compared with the GL age estimates. If the two methods correlated well, the PD method would provide a useful alternative to obtaining age estimates without having to section and count visible growth layers (if discernable) and calculate resorbed

GLs.

Correction factor

The criteria for this method are set in Parham and Zug (1997), which relies on the presence and size (diameters) of growth layers (GL) in the smaller individuals of the sample. A growth trajectory (X = number of visible growth layers; Y = diameter of the bone cross-section or periosteum diameter) is determined for each individual by connecting the point (Xi, Yi) at smallest size to point at death (Xi+n, Yi+n). The

‘average’ slope of the trajectories of the smallest individuals is determined by regression analysis (least square) and the Y-intercept is set at the diameter of hatchling bone. The correction factor (CF) is the slope coefficient and determines the number of resorbed GL.

The CF is determined by substituting the diameter of the resorption core in the regression equation:

Diameter = hatchling diameter + [(average slope)*(number of resorbed growth layers)]

Number of resorbed GL = (resorption core diameter – hatchling diameter) / average slope

The number of resorbed GL (i.e., Correction Factor) is added to the number of visible GL observed and yields an estimate of the age or total number of GL for individuals (the number of visible GL includes the periosteum diameter as the last GL or age at death).

General problems and considerations

The major problem with all skeletochronology methods is that, unless validated, growth layers are assumed to be annual. Furthermore, even if we know that GL are deposited annually, we can still only estimate age because it is impossible to accurately determine the number of lost GL due to bone remodeling (unless the animal was followed from birth). Another area of concern when using animals in which age cannot be validated is that skeletochronology relies on the formation of visible growth marks

(LAGs and annuli) to estimate age, but growth mark (GM) appearance can vary considerably between and within individuals, especially in large long-lived reptiles. It is therefore highly possible that I have underestimated age in some individuals due to poor

GM expression.

In lizards and amphibians, the appearance of double LAGs seems to be relatively easy to distinguish, at least in the smaller species where LAG spacing is more easily distinguished; however, in tortoises, double LAGs can be difficult to distinguish from two closely-spaced single LAGs. In this study where the distinction between the two

LAG expressions was not clear, I chose to record them as two single LAGs, which yields an over-estimate of one year or more in some individuals. Where annuli were also

10 difficult to discern from bone tissue anomalies (possibly be as a result of differential staining), or in cases where I questioned the appearance of a GM, I chose to exclude the age estimate from my analysis.

One concern with the GL ranking protocol is that the periosteum diameter (PD) is not included as a GL in the age estimate because it does not theoretically represent a full year’s growth. Although technically the PD does represent the age at death, the length of that growth period remains unknown. However, the problem with excluding this GL from the age estimate is that, when we compare age and body size parameters, we are taking this last period of growth into consideration regarding the size measurement but not the age estimate. In contrast, although the PD ranking protocol does take the last growth phase into account, it does not consider growth variation within and between individuals. By not measuring GL width, yearly growth is assumed to be similar and relatively unchanging within all individuals across all years, but this is definitely not the case and, in adults especially, GL width is highly variable within and between individuals.

The CF method is problematic because yearly growth variation within individuals is also not taken into consideration following the initial GL diameters used to estimate GL resorption. This method does, however, take the periosteum diameter into account and considers this growth mark the last visible GL or age at death. This is technically more accurate when considering body size, which had continued since the last visible GL deposition. However, the problem with this assumption of a full year’s growth, as previously mentioned, is that age may be over-estimated due to the unknown time of growth between the last visible GL and death.

In the following chapters, I record and discuss age estimates, age at maturity and growth curves for desert tortoises from different populations. Chapter 2 includes a comprehensive analysis of the three skeletochronological methods, and age estimates are validated against a sample of known-aged desert tortoises from a long-term demographic study in Rock Valley, Nevada to validate whether skeletochronology can be used to age tortoises. Chapter 3 includes an analysis of age structure, age at maturity and growth strategies in desert tortoises from two genetically and geographically isolated populations, namely the western Mojave Desert, California and the Sonoran Desert,

Arizona. Chapter 4 shows similar results for eastern Mojave Desert tortoises from two areas, The Desert Tortoise Conservation Center, Clark County Nevada and Ivanpah

Valley in the Mojave National Preserve, California. Chapter 5 concludes with a comparison of growth strategies for all three populations and I discuss ecological implications and various conclusions with suggestions for further research.

Dorsal surface Growth mark Growth Layer Periosteum Area of endosteal bone remodeling

Lateral mca Medial surface surface

Resorption core diameter

Ventral surface

Figure 1. Diagrammatic representation of a cross-section through the mid-shaft area of a long bone. Arrows represent diameter measurements. The initial solid arrow represents the resorption core diameter and subsequent dashed arrows represent complete growth layer (GL) diameters. The initial two GLs are partially resorbed and diameters are this not available. All diameters were recorded along the dorsoventral plane, but only where the entire circumference of a growth layer was visible; mca = medullary cavity area

CHAPTER 2. KNOWN-AGED DESERT TORTOISES (GOPHERUS AGASSIZII) FROM ROCK VALLEY, NEVADA VALIDATE SKELETOCHRONOLOGICAL ANALYSIS

Introduction

Life history traits such as longevity, growth, sexual maturity and reproduction are all important traits that require documentation when attempting to develop population attributes, life tables and viability models. Turtles are among the longest-lived vertebrates; as such, concepts related to aging are of obvious interest because they may have substantial impact on life history evolution (Congdon and Gibbons, 1990). Age at sexual maturity and longevity are, therefore, essential baseline data that all demographic studies require to conserve extant species, especially long-lived species. This demographic data is in essence, what I have tried to determine in desert tortoises

(Gopherus agassizii), which are listed as threatened throughout the Mojave Desert portion of its range, north and west of the Colorado River (U.S. Fish and Wildlife

Service, 1994).

Previous estimates of age and growth rates in desert tortoises have used scute laminae counts, annual body size measurements and body mass (Medica et al., 1975;

Turner et al., 1987; Germano, 1988; 1994). Germano’s 1988 study claimed that the number of scute laminae more or less corresponded to the ages of captively-raised tortoises and were within one to two scute laminae less than the known-aged tortoises in

Rock Valley wild desert tortoises. He therefore proposed that scute laminae and long- bone growth layers could be used to age juveniles and subadults to about 25 years of age.

Tracy and Tracy (1995), however, claim that Germano’s study does not accurately reflect growth histories because from their study of scute laminae formation in captively-raised

14 juveniles, body size, and not age, appears to determine the number of scute laminae. A recent study by Berry (2002) on scute laminae formation in juveniles from the Mojave and Colorado Deserts found that laminae counts varied by desert region. Data from 11 localities support the Tracy and Tracy (1995) premise that scute laminae are not accurate estimators of age because the scute growth seems mainly dependent on environmental factors. Berry (2002), however, stated that “in most cases the modes and medians show

1.0 ring formed per year” recognizing that the formation of multiple annual scute laminae may be influenced by sample size and/or the duration of the study and environmental conditions. A thorough review of the literature pertaining to scute growth and aging in turtles was recently published by Wilson et al. (2003).

The important question remains: How do we obtain accurate age estimates without long-term mark-recapture studies? There are no published or validated assessments of age structure in desert tortoises, aside from the mark-recapture studies of Woodbury and

Hardy (1948) and Rock Valley desert tortoises (Medica et al., 1975; Turner et al., 1987).

Both of these studies, however, derive from populations in the northeastern Mojave

Desert. Elsewhere, age structure data for desert tortoise populations are anecdotal at best.

If we are to discern and compare life history evolution among desert tortoise populations, we need a method to estimate age without the need for long-term mark-recapture studies, although the latter provides the best data. Skeletochronology has been shown to be an accurate aging method for many reptiles and is the best method for estimating age when mark-recapture is not possible (Castanet and Smirina, 1990).

In 1963, a desert tortoise mark-recapture study was initiated in Rock Valley,

Nevada by the University of California, Laboratory of Nuclear Medicine and Radiation

Biology (Medica et al., 1975). Rock Valley is located approximately 110 km northwest of Las Vegas in Nye County, NV, and along the southern boundary of the Nevada Test

Site (NTS). The goal of this study was to document incremental growth of a cohort of hatchlings and very small tortoises identified between 1963 and 1966; these small tortoises were 49-74mm in plastron length at first capture and estimated as one to four years old (Medica et al., 1975; Turner et al., 1987). Based upon the mark-recapture data,

Medica and Turner concluded that growth is largely confined to the spring and early summer, and little to no growth occurs outside of this period (Medica et al., 1975). They also determined that juvenile growth was, on average, around 9 mm per year and was positively correlated with winter rainfall and the production of winter annual plants.

To test skeletochronological age estimates, I used a sample of these known-aged

Rock Valley desert tortoises, found dead in subsequent surveys between 1996 and 2003.

Skeletochronology estimates age by counting growth layers (zones plus annuli and/or lines of arrested growth [LAGs]) (Castanet et al., 1993). My age estimates derive from three skeletochronology protocols and test which age estimates most closely match the known ages of these tortoises. This study provides the gateway to obtain accurate age estimates, most importantly ages at death and sexual maturity for desert tortoises.

Materials and methods

A sample of nine Gopherus agassizii were obtained from Rock Valley (Nye

County, Nevada): one juvenile (USNM 560940), six adult females (USNM 560933,

560934, 560935, 560936, 560939, 560941) and two adult males (USNM 560937,

560938). An additional sample of 14 tortoises derive from Piute Valley, southern

Nevada (six of which were from the Christmas Tree Permanent Study Plot and eight from south of Cal-Nev-Ari). Eight of the nine desert tortoise carcasses from Rock Valley were part of a long-term mark-recapture study (Medica et al., 1975; Turner et al., 1987) and are known-aged individuals (+ one year), USNM 560940 had not been previously captured and its age was estimated based on carapace length (CL) size using Turner et al.

1987, Table 1.

I recorded various shell measurements (where carcasses were intact), namely carapace length (CL), carapace width (CW) and plastron length (PL), with calipers (+ 0.5 mm; Table 1). In an initial study, I sectioned all appendicular skeletal elements, together with vertebrae and gular, marginal, neural and plastron scute bones to determine which elements showed the greatest number of growth layers (GL). The humerus, femur, ilium and scapula were the long bones that I used for the skeletochronological analysis. I recorded longitudinal length (LL), proximal width (PW), midshaft width (MW) and distal width (DW) from all long bones using digital calipers (+ 0.05 mm; Table 2).

I fixed, decalcified and processed bones according to standard histological protocol. I embedded all bones in Paraplast Plus©, cut 20 µm cross sections through the midshaft area of each bone using a rotary microtome and stained sections with hematoxylin and eosin. All growth layer diameters were measured along the dorsoventral plane using a transmitting light microscope.

Skeletochronology

I compared two skeletochronological methods to assess the variation in published chelonian age estimation methodology (Ranking Protocol and the Correction-Factor method). The Ranking Protocol involves assigning ages to growth layer (GL) diameters

17 and ranking them into age classes (see Zug, 1990; 1991; Parham and Zug, 1997). I also assigned an age estimate to the periosteum diameter (total bone cross section diameter), which technically represents the last GL deposited and, therefore, the age at death (see

Chapter one). The Correction-Factor (CF) method is explained in detail in Parham and

Zug (1997). Hatchlings and small juveniles are essential for this method.

I assessed the mean difference between age estimates and known ages using a

Students’ t-test. I calculated the difference between skeletochronology age estimates and known age and determined the mean squared prediction error (MSE) and root mean squared error (RMSE) to determine which skeletochronology method provided age estimates with the least error and bias to the known ages. I calculated MSE by subtracting age estimates from actual ages and then squared the corresponding value, I then calculated the mean (+ one standard error of the mean) of the squared errors for each aging method. The RMSE is simply the square root of the MSE. I calculated RMSE because this gave the error value the same dimensionality as the actual and predicted values. I calculated the mean of the root of the squared error values for each age estimate

(+ one standard error from the mean). I assessed the relationship between known age

(independent variable) and skeletochronology age (dependent variable) and also age

(independent variable) and size (dependent variable) using the Pearson correlation matrix and least square regressions. SYSTAT for Windows, version 8 was used for all statistical analyses.

Results

Age and longevity

In Gopherus agassizii, the humeri and ilia generally displayed the greatest number of visible growth layers (GL). In general, all three skeletochronology methods produced similar age estimates in juveniles less than 95 mm CL (Table 3 and 4). Above 95 mm

CL, variation in age estimates increased, especially in adults. In the only juvenile which had been assigned a known-age (based on size; USNM 560940), the Correction Factor age estimate was most similar to the recorded age, although all age estimates exceeded the recorded age by six to ten years (Table 4; Fig. 2).

The Correction Factor (CF) method generally produced the youngest age estimates of all three methods, especially in subadult and adult tortoises (Table 4). When all three skeletochronology age estimates were compared to adult known ages, GL age estimates were the closest or equal to the known ages in six out of eight individuals

(Table 4; Fig. 2). In USNM 560933 (a subadult), the CF age estimate equaled the recorded known age, whereas in another subadult (USNM 560934), the PD age estimate most closely matched the known age (Table 4; Fig. 2). In adults, the GL age estimate was closer to the known age than either the CF or PD estimate (Table 4; Fig. 2).

I compared age using Students’ t-tests and found no significant difference between skeletochronology age estimates and known age (Table 5). The PD and GL mean were similar to the known-age mean, whereas the CF mean was noticeably lower

(Fig. 3), however, GL age estimates showed the least error amongst the three aging methods (Table 5). Using the Pearson correlation matrix, which corresponded to the r2 values from the regression analysis, GL age estimates correlated more strongly with

19 known ages than either PD or CF age estimates (GL age: 0.95; PD age: 0.55; CF age:

0.91).

To determine if age and body size were related, I regressed age versus carapace length and found a positive relationship between age and size for all methods (Fig. 4).

Known age and GL age, however, showed the strongest relationship with body size (Fig.

4).

Age at sexual maturity

Turner et al. (1987) used a size of approximately 190 mm CL to indicate a size at maturity in Rock Valley tortoises because all females above this size were shown to have eggs. The only Rock Valley female around this size (189 mm CL) had a known age of

15 years (Table 4). Skeletochronology age estimates for this female ranged from 15 to 24 years (depending on the method). In comparison, I interpolated age at maturity (~ 190 mm CL) from the known age vs size regression line, and obtained an age of around 26 years (Fig. 5).

Discussion

Age and longevity

In general, all three skeletochronology methods yielded similar age estimates for juveniles; however, there is some indication that the Correction Factor (CF) method is the better method to use for aging juvenile tortoises because it relies solely on hatchling and juvenile bone resorption rates to calculate the number of growth rings resorbed by bone remodeling (Parham and Zug, 1997). Subadult tortoises present more of a problem, however, because as tortoises approach sexual maturity and energy gets channeled into

20 maintaining more than just maintenance and growth, individual variation in remodeling rates will become more pronounced. The CF method is the most conservative of the three aging techniques and, as such, ages tend to be greatly underestimated the larger and older tortoises become (Table 3). In the smallest subadult female (USNM 560933; near the minimum size at sexual maturity), the Correction Factor age actually equaled the known age and my conclusion is that this skeletochronology protocol provides the most accurate method for aging juvenile tortoises and potentially also subadult desert tortoises.

In adults, the Correction Factor method significantly underestimated actual age.

The likely reason is that this method is based solely on hatchling and juvenile growth to determine resorbed growth rings in adults. Even though cortical remodeling is considered to some degree (within resorption core diameters), it is mainly considered in regard to juvenile growth rate (in which remodeling is usually very limited) and not the rate and extent to which it occurs in adults. Growth of juvenile and subadult desert tortoises appears to be linear at least up to ages of 25 years (Turner et al. 1987, Figure 1), but the growth of these same animals plateaus as they attain adult size (Medica, unpublished data). The growth layer (GL) Ranking Protocol produced estimates close to or equal to adult known ages (Table 4). Where ages are over- or under-estimates, it is possible that we classified a double LAG (two periods of arrested growth within one growth cycle) as two single LAGs or vise versa, or that a LAG was too indistinct and not counted. Overall, however, it appears that skeletal growth layer formation is annual and supports my hypothesis that skeletochronology can be used to provide reliable age estimates in adult desert tortoises. Therefore, I recommend using the Correction Factor

(CF) method for aging juveniles and subadults and the growth layer (GL) Ranking

Protocol for aging adults.

Age at sexual maturity

Turner et al. (1987) previously estimated age at maturity in Rock Valley tortoises to be between 17-20 years. This estimate was based on the assumption that relationships between age, size and maturity were similar between tortoises from Goffs, California and

Rock Valley, Nevada (Turner and Berry 1986; Turner et al., 1987). Of the 15 Rock

Valley tortoises recaptured in 1985 (Turner et al., 1987), five were identified as female based on secondary sexual features and x-rayed for eggs. These females were all above

200 mm CL and were 23-24 years in age. If the minimum size at sexual maturity in females is 190 mm CL, then a minimum age at maturity of 15-18 years is supported by the results of my study.

The growth of juvenile desert tortoises has been shown to be closely correlated with rainfall and annual production in the northeastern Mojave Desert (Medica et al.

1975). Similarly, Berry (2002) showed that juvenile tortoise growth (based upon scute laminae counts) is positively correlated with annual precipitation at Goffs, CA.

Environmental conditions at Rock Valley are similar to those at Goffs (Medica, pers. comm.) and, for this reason, although Rock Valley size/age at sexual maturity estimates were based on the size/age relationships of tortoises from Goffs, the age estimates reported by Turner et al. (1987) should not differ by more than one year. Comparing rainfall amounts between 1982 and 1986 in Rock Valley with those listed in Berry (2002,

Table 4:421) indicated that in eight of twelve identical seasonal comparisons, Rock

Valley received more precipitation than Goffs (Table 6). Summer rains during the same

22 period of time exceeded those at Goffs three of the four years (Table 6). Unfortunately, I do not have annual biomass data for the Rock Valley site during the identical time period for direct comparative purposes but it was evident that density of Bromus rubens in 1983-

1985 was increasing significantly and native annuals also were present in significant numbers (Hunter, 1991). At Goffs during the 1982-83 hydrologic year, with 298 mm of rainfall, annual biomass production was 42.0 g/m2. In an earlier El Nino year, 1973, with a September-August rainfall of 250 mm, net annual production of annuals amounted to 64.4 g/m2 (Turner and Randall, 1989) indicating that ephemeral production can be high at both locations as a result of winter rain. At both Goffs and Rock Valley, the predominant rainfall occurs in the fall and winter although Rock Valley has more summer rain; however, summer rains support only a few summer annual species that are rarely a prominent component of the community (Beatley, 1976). The significance of this difference may be that tortoises in Rock Valley have a greater opportunity to drink after summer rains than those at Goffs. Therefore, their consumption of dry annuals maintains a semblance of water and salt homeostasis (Nagy and Medica, 1986).

Additionally, the lack of summer rains and drought conditions may account for the subsequent high mortality observed at Goffs between 1995 and 2000 (Tracy et al., 2004).

Any size/age estimates for desert tortoises based on data coming from outside the area of study, is not reliable until validated. Mueller et al. (1998) studied desert tortoises at Yucca Mountain, Nevada, a locality within 10 km of Rock Valley, and found the smallest sexually mature tortoise to be 209 mm CL. This female, however, was the smallest tortoise with eggs x-rayed; also, the years in which this study was conducted were some of the wettest years ever recorded. They estimated age at this size as 19-20

23 years old, based on scute laminae counts. Berry (2002) documented that scute laminae formation is highly variable between desert tortoise localities. Her data shows that scute laminae expression is influenced more by environmental factors than by an internal rhythm.

Conclusions and implications for future research

This study demonstrates that it is possible to obtain relatively accurate age estimates for desert tortoises using skeletochronology where known-age mark-recapture data are unattainable. Skeletochronology is a relatively inexpensive, non-labor intensive method that can be used across populations with the added advantage of being able to provide invaluable data for desert tortoise life history studies by providing sex and size/age structure of past populations (using carcass or fossil collections).

An interesting result of this study, in contrast to most skeletochronology studies, was that ilia generally showed the greatest number of visible growth rings and provided the most accurate age estimates (when calibrated to the known-aged Rock Valley tortoises), not the femora and humeri usually used. This observation is important because the pelvic girdle tends to be the only long bone material remaining in a carcass shell after scavenging. From my preliminary skeletal screening, there is some support for the use of marginal and, to a lesser extent, vertebral shell bones but the remodeling process is more intensive in shell skeletons than in upper limb and girdle bones. I therefore do not recommend the use of plastron or carapace bones unless age estimates from these elements can be validated.

The most serious problems facing the remaining desert tortoise populations in the

Mojave Desert region is the combined impact of habitat loss, human impacts (i.e.,

24 urbanization, proliferation of roads and recreation) as well as predation and disease. A detailed discussion of impacts upon desert tortoises has been thoroughly reviewed in a recent report by Tracy et al. (2004). Drought is another problem facing the desert tortoise and years of back-to-back drought appear to result in high adult desert tortoise mortality

(Longshore et al., 2003).

All the above factors increase mortality rates in desert tortoise populations and leave many carcasses, which were of limited benefit in increasing our understanding of population dynamics and demography. Now, with skeletochronology, carcasses found in the wild or in museum collections can be used to obtain size and age data, as well as growth rates. These data provide information on microevolution within the species when compared to fossil material, all without the need to harm or disturb natural populations of this threatened species. Ecological constraints are known to influence bone growth strategies and, by extension, life history strategies in a wide variety of reptiles (Castanet et al., 1993). Consequently, the desert tortoise skeleton can provide a biological

‘calendar’ of the events that shaped life up until the time of death.

Table 1. Shell measurements for Gopherus agassizii carcasses from the East Mojave Desert, Nevada.

carcass # Sex Locality CL CW PL (mm) (mm) (mm)

14 E 4 Juvenile *Cal-Nev-Ari 58 47

R10 Juvenile *Christmas Tree Plot 54.5 49.8 51.5

19-2/2 Juvenile *Cal-Nev-Ari 54.7 49.1 52.1

19-2/1 Juvenile *Cal-Nev-Ari 56.7 47.6 54.7

19-1/1 Juvenile *Cal-Nev-Ari 60.2 50.8 60.9

R8 Juvenile *Christmas Tree Plot 61 54.9 60

49 E 4 Juvenile *Cal-Nev-Ari 61.7 52 59.3

19-1/3 Juvenile *Cal-Nev-Ari 67.3 54 60.2

R5 Juvenile *Christmas Tree Plot 67

20-3/1 Juvenile *Cal-Nev-Ari 80.4 70.2 74.2

17 E 1 Juvenile *Cal-Nev-Ari 87.4 69.6 84.4

R3-T Juvenile *Christmas Tree Plot 88.6 70.5

R19 Juvenile *Christmas Tree Plot 102.5 81.5 100.6

R17 Juvenile *Christmas Tree Plot 122.6 100.8 110

USNM 560940 Juvenile † Rock Valley 123 87 117

USNM 560935 Female † Rock Valley 141 169

USNM 560933 Female † Rock Valley 189 138 176

USNM 560934 Female † Rock Valley 220 172 208

Table 1 (continued).

USNM 560936 Female † Rock Valley 225 184 215

USNM 560937 Male † Rock Valley 226 182 223

USNM 560941 Female † Rock Valley 228 180 217

USNM 560939 Female † Rock Valley 243 181 219

USNM 560938 Male † Rock Valley 254 185 233

* Southern Piute Valley † Nye County, Nevada

Table 2. Bone measurements for Gopherus agassizii carcasses from the East Mojave Desert, Nevada; CL = carapace length, H = humerus, F = femur, I = ilium, S = scapula, J = juvenile, F = female, M = male.

Tortoise Sex CL Bone Length (mm) Proximal Width (mm) Midshaft Width (mm) Distal Width (mm) ID (mm) H F I S H F I S H F I S H F I S

19-2/2 J 54.7 13.3 5.1 1.5 4.6

19-2/1 J 56.7 14.2 5.4 1.9 4.6

19-1/1 J 60.2 13.4 11.2 4.7 4.8 1.4 1.7 4.3 5.9

14 E 4 J 11.3 4.4 1.8 6.2

R8 J 61 14.3 4.1 1.7 5.8

49 E 4 J 61.7 15.4 5.6 2 6

19-1/3 J 67.3 15.4 5.6 1.9 5.3

R5 J 16.9 13.6 6 6.3 2 2.2 6 7

20-3/1 J 80.4 10.3 3.4 1.8 7

17 E 1 J 87.4 15.3 6.3 2.1 7.4

Table 2 (continued).

R3-T J 88.6 14.6 6.3 2.3 7.8

R19 J 102.5 22 16.7 7.8 7.6 2.4 2.8 7.3 8.6

R17 J 122.6 33.5 22.5 11.3 9.2 3.3 3 10.2

USNM 560940 J 123 27 22.4 9.4 9.9 3.6 3.7 8.9 12

USNM 560935 F 4.9 4.8 15.9

USNM 560933 F 189 8.5 3.7

USNM 560934 F 220 57.4 47.7 40 27.9 19.7 17 16.6 10 6.8 6.1 6.5 7.2 19.9 15.3 22.2 22.9

USNM 560936 F 225 6.5 6

USNM 560937 M 226 41 22 10.3 6.6 7.2 23

USNM 560941 F 228 61 22 11.6 7.6 7

USNM 560939 F 243 60 22.3 7.5 22.6

USNM 560938 M 254 42.1 19.5 6.5 23.2

Table 3. Periosteum diameter (PD) and growth layer (GL) ranking protocol and Correction Factor (CF) age estimates for Gopherus agassizii from the East Mojave Desert (Nevada); J = juvenile, F = female, M = male.

Tortoise ID sex CL Humerus ilium scapula Femur

(mm) Age Age Age Age Age Age Age Age Age Age Age Age (GL) (PD) (CF) (GL) (PD) (CF) (GL) (PD) (CF) (GL) (PD) (CF) 14 E 4 J 3 5 4

20-3/1 J 80.4 6 14 7

19-1/1 J 60.2 3 2 4 3 4 4

19-1/3 J 67.3 8 9 6

19-2/1 J 56.7 3 4 4

R8 J 61 7 8 7

17 E 1 J 87.4 6 9 8

49 E 4 J 61.7 7 8 6

R3-T J 88.6 8 9 8

Table 3 (continued).

R10 J 54.5 5 6 4 4 4 5 3 4 4

R7 J 122.6 13 17 12 17 16 13

R5 J 95 6 11 10 6 12 10

R19 J 102.5 13 17 10 12 12 11

USNM 560940 J 123 18 19 15 13 18 14

USNM 560933 F 189 21 24 15

USNM 560935 F 29 23 22 34 20 23

USNM 560934 F 220 35 36 21 40 31 31 31 23 29 33 23 25

USNM 560936 F 225 34 36 23 40 27 25

USNM 560937 M 226 39 27 24 35 27 22

USNM 560941 F 228 35 49 29 29 24 20

Table 3 (continued).

USNM 560939 F 243 46 48 32

USNM 560938 M 254 47 27 35

Table 4. Maximum age estimates for Gopherus agassizii from the East Mojave Desert (Nevada) using the periosteum diameter (PD) and growth layer (GL) ranking protocol and correction factor (CF) skeletochronology methods. Age estimates in bold are equal or closest to the known age (where available). The value in brackets is the difference between the age estimate and known age; CL = carapace length.

ID maturity CL age age age Known (mm) (PD) (GL) (CF) Age

14 E 4 juvenile 5 3 4 N/A

R10 juvenile 54.5 6 5 5 N/A

19-2/1 juvenile 56.7 4 3 4 N/A

19-1/1 juvenile 60.2 4 3 4 N/A

R8 juvenile 61 8 7 7 N/A

49 E 4 juvenile 61.7 8 7 6 N/A

19-1/3 juvenile 67.3 9 8 6 N/A

20-3/1 juvenile 80.4 14 6 7 N/A

17 E 1 juvenile 87.4 9 6 8 N/A

R3-T juvenile 88.6 9 8 8 N/A

R5 juvenile 95 12 6 10 N/A

R19 juvenile 102.5 17 13 11 N/A

R17 juvenile 122.6 17 17 13 N/A

USNM 560940 juvenile 123 19 18 15 (-6) 9

USNM 560933 subadult female 189 24 21 15 15

USNM 560935 subadult female 23 34 23 34

USNM 560934 Adult female 220 36 40 31 36

Table 4 (continued).

USNM 560936 Adult female 225 36 40 25 40

USNM 560937 adult male 226 27 39 (-1) 24 40

USNM 560941 Adult female 228 49 35 (-5) 29 40

USNM 560939 Adult female 243 48 46 (+4) 32 42

USNM 560938 adult male 254 27 47 (-3) 35 50

Table 5. Student’s t-test analysis and mean difference (+ one standard error [SE] of the mean) between known age versus skeletochronology age estimates in Gopherus agassizii from Rock Valley, Nevada. Mean squared error (MSE) and root mean squared error (RMSE) are given to show which aging method provided ages with the least error and bias to known ages; PD = periosteum diameter, GL = growth layers, CF = correction factor.

Known age vs Mean MSE RMSE t p

skeletochronology difference (+ SE) (+ SE)

Estimate (+ SE)

PD age 9.44 (2.14) 128.89 (53.37) 9.44 (2.14) 0.330 0.746

GL age 4.56 (1.23) 25.67 (8.66) 4.11 (1.05) -0.298 0.770

CF age 10 (1.80) 123.22 (31.48) 9.89 (1.78) 1.701 0.108

Table 6. Rainfall comparisons between Goffs, California and Rock Valley, Nevada during identical time periods between 1982 and 1986. The (+) accentuates which location was greater during the time period measured.

Years Annual Winter Summer Precipitation Precipitation Precipitation

October-September October-March July-September

Goffs Rock Goffs Rock Goffs Rock

Valley Valley Valley

1982-3 298.0+ 278.1 118.0 152.4+ 161.9+ 35.6

1983-4 238.5+ 184.4 135.1+ 41.1 72.7 121.4+

1984-5 110.0 167.1+ 91.0 106.4+ 15.2 138.9+

1985-6 58.3 178.1+ 37.0 108.7+ 21.3 44.2+

60 known age PD age 50 GL age CF age 40

30 Age (years) 20

0 560940 560933 560935 560934 560936 560937 560941 560939 560938 Tortoise ID

Figure 2. A comparison of skeletochronology age estimates with known-ages in Gopherus agassizii from Rock Valley, Nevada; PD = periosteum diameter, GL = growth layers, CF = correction factor.

known age

CF age

GL age

PD age

0 10 20 30 40 50 60 Age (years)

Figure 3. Box plot of skeletochronology age estimates and known age of Gopherus agasizzii from Rock Valley, Nevada; PD = periosteum diameter, GL = growth layers, CF = correction factor. Vertical lines within the box plots represent the median age and whiskers the 10th and 90th percentile.

60 known age vs PD age y = 0.44x + 17.03 (r ² = 0.30) known age vs GL age y = 0.71x + 11.22 (r ² = 0.82) known age vs CF age y = 0.48x + 9. 00 (r ² = 0.82) 50

20 Skeletochronology Age (years)

10 0 10 20 30 40 50 60

Known Age (years)

Figure 4. Regression of known age versus skeletochronology age estimates in Gopherus agassizii from Rock Valley, Nevada; PD = periosteum diameter (solid line), GL = growth layers (dashed line), CF = correction factor (dotted line).

280

260 240

220

200 180 known age vs CL y = 2.72x + 121.04 (r ² = 0.86) 160 PD age vs CL y = 2.24x + 139.04 (r ² = 0.36)

Carapace Length (mm) 140 GL age vs CL y = 3.20x + 100.42 (r ² = 0.75) 120 CF age vs CL y = 4.64x + 93.92 (r ² = 0.72) 100 0 10 20 30 40 50 60

Age (years)

Figure 5. Regression of age versus carapace length (CL) in Gopherus agassizii from Rock Valley, Nevada; PD = periosteum diameter, GL = growth layers, CF = correction factor.

CHAPTER 3. AGE AND GROWTH STRATEGIES OF SONORAN DESERT AND WEST MOJAVE DESERT TORTOISES (GOPHERUS AGASSIZII).

Introduction

Individual growth is intimately associated with changes in life-history traits

(Shaffer, 1974). Growth rate affects how long juveniles are subject to high predation, when juveniles mature, how many eggs can be produced per year by a female (if clutch size is dependent on body size), the size that adults achieve, and the ability of adults to survive over long periods of time (Congdon and Gibbons, 1990; Germano, 1994).

Ecological constraints are known to influence bone growth strategies and thus life history strategies in a wide variety of amphibians and reptiles (Caetano et al., 1985; Castanet and

Baez, 1991; Collins and Rodda, 1994; Bjorndal et al., 1998; Zug and Glor, 1998; Curtin et al., 2005), however, few such studies have been done on tortoises.

Tortoises, like other ectothermic tetrapods, display a cyclical pattern of growth.

Cyclic skeletal growth produces successive bony layers in association with an internal

(genetically based) rhythm synchronized and reinforced by seasonal cycles (Meunier et al., 1979; Castanet et al., 1993). Growth layers can theoretically be used to estimate age, and this method is called skeletochronology. Bone remodeling is fundamental to the general process of skeletal growth and is concerned with the continual reshaping of a bone during growth (Enlow, 1963). Because bone is hard, it cannot grow and increase in mass by the internal expansion of existing tissues, as do most soft tissues, it can only grow by an appositional process of surface deposition together with resorption from a contralateral surface (Enlow, 1963). At the medullary cavity margin, endosteal resorption can also remove initial growth layers, in other words the earliest growth layers

41 deposited, thereby causing individual ages to be underestimated (Castanet and Smirina,

1990). The rate of periosteal bone resorption, however, can be evaluated by using either known-age individuals or by a back-calculation method (Castanet and Smirina, 1990).

Studies on growth rates in wild desert tortoises (Gopherus agassizii) have been evaluated using annual body size measurements, scute rings and body mass as parameters

(Woodbury and Hardy, 1948; Medica et al., 1975; Turner et al., 1987; Germano, 1988;

1994). These studies have mainly concentrated on tortoises from the northeastern

Mojave Desert, however, and the absence of data from different populations of desert tortoises prevents the examination of inter-regional variation in growth rates and maturity. Moreover, the different methodology employed in each study confounds the comparison of results. I wished to unify and interpret population data using a common methodology that would allow for comparisons about tortoise biology across a broad geographic scope. Desert tortoises represent a perfect species to investigate as they are long-lived, living in a habitat where their survival is extremely dependent on resource availability and quality, which are directly or indirectly influenced by environmental factors (mainly rainfall). By using carcasses collected in the wild and determining age estimates by Skeletochronology, I had the added advantage of not having to harm, kill or disturb living animals of this threatened species.

In this study I compared West Mojave and Sonoran Desert tortoises. Using restriction site analysis, Lamb et al. (1989) resolved five different mtDNA genotypes among 22 populations of desert tortoises and found three well-defined genetic assemblages: a Mojave assemblage, Sonoran assemblage and Sinaloan assemblage. The genetic distances (5.1 – 5.6 percent) observed between Mojave and Sonoran genotypes of

42 desert tortoises were significantly higher than distance values reported for any other turtles species (Walker and Avise, 1998; Lamb and McLuckie, 2002). Consequently, depending on molecular clock timing, the Mojave and Sonoran mtDNA lineages appear to have diverged some 5 or 6 million years ago (Lamb and Lydeard, 1994; Lamb and

McLuckie, 2002). Regardless of the timing of the divergence of the Mojave and Sonoran

Desert populations, there can be no denying that there is a great deal of genetic distance between tortoises from these two deserts (McCord, 2002). The Sonoran Desert formed as part of a drying trend beginning in the middle Miocene and desert tortoise habitat was in existence by the late Miocene (8 to 5 million years ago). In contrast, the Mojave Desert continued to develop well into the Pleistocene, with the modern Mojave Desert having a continuous history of only 5000 to 12 000 years (Morafka and Berry, 2002; Van

Devender, 2002). The Mojave Desert, therefore, is considered the youngest biotic province in North America (Van Devender, 2002), and it is clear that Mojave tortoises are younger on the evolutionary scale than Sonoran tortoises. Moreover, within the

Mojave Desert, West Mojave tortoises are potentially younger than their eastern counterparts (Morafka and Berry, 2002). I therefore wanted to compare age and growth strategies within West Mojave and Sonoran Desert tortoise populations because they represent the eastern and western extremes, both morphologically and behaviorally as well as climatically and ecologically, within the desert tortoise range in the United States

(Morafka and Berry, 2002; Van Devender, 2002).

West Mojave Desert tortoises

The Mojave Desert is a high desert, occurring at elevations between 600 and

1,200 m (Luckenbach, 1982; Germano et al., 1994). Creosotebush (Larrea tridentata)

43 and white bursage (Ambrosia dumosa) dominate as much as 70 % of the landscape and the desert tortoise (Gopherus agassizii) often occurs in this habitat type (Germano et al.,

1994). Where the Joshua-tree (Yucca brevifolia) and Mojave yucca (Y. schidigera) are conspicuous, the abundance of desert tortoises is usually low to moderate (Luckenbach,

1982). The Mojave Desert is especially rich in ephemeral plants, most of which are winter annuals (Turner and Brown, 1982) and these are important foods for the desert tortoise, although they also eat perennial grasses (Woodbury and Hardy, 1948; Berry,

1978; Luckenbach, 1982). Most of the Mojave Desert has been invaded by introduced winter annuals such as Bromus rubens, Schismus barbatus and Erodium cicutarium, which can dominate cover and biomass in some years (Oldemeyer, 1994).

Tortoises emerge to feed and mate during the fall (following nesting) and early spring (prior to nesting) and remain active throughout the spring and sometimes after summer thunderstorms (Rostal et al., 1994). Eggs and hatchlings are quite vulnerable with pre-reproductive adult mortality averaging 98 % (U.S. Fish and Wildlife, 1994).

Adults are, for the most part, well protected against predators and their longevity helps compensate for their variable annual reproductive success, which tends to correlate with environmental conditions (Germano et al., 1994).

In Mojave Desert populations, there are approximately equal numbers of males and females (Averill-Murray et al., 2002b). Female tortoises as small as 176 mm CL lay eggs in the West Mojave Desert, and this is the currently accepted minimum size at sexual maturity (Germano, 1994). Minimum age at first reproduction may be as low as nine years but estimates range up to 26 years (Medica et al., 1975). In addition, tortoises lay their eggs earlier in the Mojave Desert than in the Sonoran Desert, from late April

44 though mid-July (Turner et al., 1986; Wallis et al., 1999; Averill-Murray et al., 2002b).

Rainfall greatly influences tortoise reproduction, and tortoises may lay as many as three clutches in a year (but the average is 1 to 2 clutches).

Temperatures are hot in summer (mean is 36 °C) and near or below freezing in winter. Mean January temperature is around -3 °C, and there are on average 84 days when temperatures are below freezing (U.S. Fish and Wildlife, 1994). Rainfall in the

Mojave Desert is low (80 to 170 mm/yr) and lower in the western portion than in other areas of the desert tortoise range (Germano, 1988, 1994). In the western Mojave Desert, most precipitation is in winter, with less than 10 % occurring in the summer.

Sonoran Desert tortoises

The Sonoran Desert has been divided into six units of recognizable vegetational differences, and G. agassizii occur in portions of four of them: the lower Colorado River valley, Arizona uplands, plains of Sonora and central gulf coast (Germano et al., 1994).

In this study, population samples came from Colorado River valley (Mohave county) and

Arizona Upland communities. Arizona Upland is more similar to foothills thornscrub than to the arid creosotebush communities of the lower Colorado River Valley (Van

Devender, 2002) and Turner and Brown (1982) suggested that this community could be considered a drier and more temperate thornscrub rather than a subdivision of the

Sonoran Desert.

Sonoran Desert tortoises grow most rapidly early in life and reach 36-47 % of their maximum CL before growth begins to slow (Murray and Klug, 1996). Rapid early growth contributes to high juvenile survivorship, but maximum sizes differ between sexes among populations (Averill-Murray et al., 2002b). Hart (1996) found that

45 individuals in populations north of the Gila River in Arizona tend to reach larger sizes than individuals south of the river.

In the Sonoran Desert, with only a few exceptions, sex ratios are typically balanced with approximately equal numbers of males and females (Averill-Murray et al.,

2002b). Female tortoises lay an average of about five eggs, but between one and twelve eggs in a clutch have been recorded (Averill-Murray et al., 2002b). Clutch size is not related to body size, and thus far, the smallest recorded size at which Sonoran desert tortoises have laid eggs in the wild is 220 mm CL in the Mazatzal and Maricopa

Mountains (Averill-Murray et al., 2002b). Ages of these tortoises were not measured directly, but were estimated at between ten to twenty years based on growth curve comparisons. More precise estimates of age at maturity were not possible because of insufficient data on growth (which varies geographically) for these populations (Averill-

Murray et al., 2002b). Minimum size at sexual maturity, however, is currently accepted as ~180 mm CL (Germano, 1994; Averill-Murray and Averill-Murray, 2005).

Sonoran Desert females lay a maximum of one clutch in a year, but many females may not reproduce each year. Egg-laying in the Sonoran Desert generally occurs near the onset of the summer rainy season and has been observed from early June to early August, but the average egg-laying date each year does not appear to be directly related to recent rainfall (Averill-Murray et al., 2002).

Thus, given that West Mojave and Sonoran Desert tortoises represent the relative extremes of tortoise life in the desert portion of their range, I wanted to compare how life in these differing desert environments affected population age structure, age at maturity and age at death.

Materials and Methods

Materials

I obtained a sample of 72 wild desert tortoise (Gopherus agassizii) carcasses from five counties in the Sonoran Desert, Arizona (Table 7 and 8). In total there were 24 juveniles, 28 adult females, 17 adult males and 4 individuals of unknown sex. The title of juvenile, adult male, adult female and unknown sex had been assigned to the carcasses and recorded on carcass sheets by the Arizona Fish and Game collectors. The West

Mojave Desert population sample consisted of 69 wild desert tortoise carcasses; 13 adult females, 30 adult males, 22 juveniles and 4 individuals of unknown sex. West Mojave carcasses were collected along highway 58 in San Bernardino County, California

(donated for use in this study by Dr. William Boarman).

Skeletochronology

I recorded various shell measurements (where carcass shells were intact), namely carapace length (CL), carapace width (CW) and plastron length (PL), with manual calipers (+ 0.05 mm). In an initial study I sectioned all appendicular skeletal elements, together with vertebrae and gular, marginal, neural and plastron scutes to determine which elements showed the greatest number of growth marks (GMs). The humerus, femur, ilium, and scapula were the long bones that I used for the skeletochronological analysis. I recorded longitudinal length (LL), proximal width (PW), midshaft width

(MW) and distal width (DW) from all long bones using digital calipers (+ 0.05 mm).

I fixed, decalcified and processed bones according to standard histological protocol. All bones were embedded in Paraplast Plus©, cut 20 µm cross sections through the midshaft area of each bone using a rotary microtome and stained sections with

47 hematoxylin and eosin. I measured all growth layer diameters along the dorsoventral plane using a transmitting light microscope.

Comparisons of skeletochronology age estimates (determined by three methods) to a sample of known-aged desert tortoises from Rock Valley, Nevada (Chapter two) indicated that the Correction Factor method provided the most accurate juvenile age estimates and the growth layer Ranking Protocol (hence forth called Ranking Protocol) the most accurate adult age estimates. I therefore used these two skeletochronology methods, in this and the following chapter, to determine juvenile and adult age estimates.

The Ranking protocol involved assigning ages to growth layer (GL) diameters and ranking them into age classes (see Zug, 1990; 1991; Parham and Zug, 1997). The

Correction-Factor (CF) method is set out in detail in Zug and Parham (1997). Hatchlings and small juveniles are essential for this method.

Statistical analysis

Because the Arizona sample consisted of multiple populations, I compared all size measurements and age estimations to determine whether differences existed among the populations. I conducted one way analysis of variance (ANOVA) tests, which I combined with a Tukey post hoc test to ascertain statistically significant differences between populations. I conducted least squares regression analyses and ANOVAs to compare shell and bone measurements between the Sonoran and West Mojave population samples. I compared the various shell and bone measurements and age estimates from within and between both groups to ascertain any significant sexual dimorphism using

ANOVA. I conducted statistical analyses using the SYSTAT (Version 9.0) statistical package and accepted statistical significance as P < 0.05.

Results

Body size

I compared carapace length (CL), carapace width (CW) and plastron length (PL) between the various Sonoran Desert populations and found no significant difference between localities for any group (adult males, females and juveniles). There was no significant size dimorphism between adult males and females in the Sonoran sample.

When I compared CW and PL and removed the effect of CL (ANCOVA) to make the analysis more precise (in other words as a way of comparing regression slopes) I also found no statistically significant difference (CW: F = 0.229, P 0.636, r2 = 0.76; PL: F =

0.627, P = 0.435, r2 = 0.75; Fig.6 and 7A).

The West Mojave population, showed a strong sexual dimorphism in all shell measurements (Fig. 7B). When the effect of CL was removed (ANCOVA), however there was no significant difference between the sexes in CW (F = 0.564, P = 0.460, r2 =

0.90) or PL (F = 0.096, P = 0.760, r2 = 0.92).

Sonoran females were significantly larger than West Mojave females in CL (F1,33

= 6.462, p = 0.016) and approached having statistically significantly larger PL than West

Mojave females (F1,30 = 3.687, P = 0.064; Fig. 8A). When the effect of CL was removed

(ANCOVA), West Mojave females were significantly wider than Sonoran females (F =

6.045, P = 0.021, r2 = 0.73). The adjusted least squares means from this analysis can be regarded as a comparison of the regression lines because these values represent the predicted CW (170.3 + 2.0 SE in Sonoran females and 179.9 + 3.3 SE in West Mojave females) at the predicted average value of the independent variable (carapace length).

There was no difference between Sonoran and West Mojave females in PL when the effect of CL was removed (F = 0.023, P = 0.881, r2 = 0.79; Fig. 8A).

West Mojave and Sonoran males were generally similar in body length (CL: F1,33

= 0.013, P = 0.192; PL: F1,31 = 3.459, P = 0.072), but West Mojave males were significantly wider than Sonoran males (F1,30 = 7.157, P = 0.012; Fig 8B). When the effect of CL was removed in an ANCOVA analysis, the difference in CW between males was still statistically significant, with West Mojave males being wider than Sonoran males (F = 11.259, P = 0.002, r2 = 0.89). The adjusted least squares mean for predicted

CW at the average CL was 184.0 + 3.5 SE in Sonoran males and 198.8 + 2.7 SE in West

Mojave males. There was no difference in PL between SD and WM males when the effect of CL was removed (F = 3.640, P = 0.068, r2 = 0.88; Fig. 8B).

Juveniles were similar in shell size and showed no statistically significant difference between the Sonoran and West Mojave Deserts (Fig. 9).

Bone morphometry

There was no statistically significant difference in bone length in adult male or female long bones between the various Sonoran Desert populations. There was no statistically significant difference in bone length between males and females in the

Sonoran Desert (Fig. 10A). To remove the effect of body size, I conducted an analysis of covariance (ANCOVA) with CL as the covariate and again found no sexual dimorphism in the length of all bones. In the West Mojave Desert, male tortoises had statistically significantly longer humeri than females (F1,8 = 6.412, P = 0.035; Fig. 10B). Ilia were also longer in males but the sexual dimorphism was just below statistical significance

(F1,30 = 3.878, P = 0.058; Fig. 10B). There was no sexual dimorphism in scapula length

50 in West Mojave adults. When I removed the effect of body size on bone length

(ANCOVA), however, I found no statistically significant difference in length for any long bone between West Mojave males and females.

Humeri and ilia were statistically significantly longer in Sonoran females than in

West Mojave females (Fig. 10; Table 9). When the effect of body size (carapace length) was removed by analysis of covariance, however, then only ilial length was still

2 statistically significant between females of both groups (F1,23 = 7.382, P = 0.012, r =

0.60). There was no statistically significant difference in the bone length of humeri, ilia and scapulae in males or juveniles between the two deserts (Fig. 10; Table 9).

To examine population trends, I grouped adults and juveniles together and regressed TBL against body size (Table 10). In Sonoran tortoises, there was a statistically significant relationship between bone length and CL for all bones (humeri:

F1,21 = 497.069, P < 0.001; ilia: F1,39 = 972.252, P < 0.001; femora: F1,15 = 656.360, P <

0.001; scapulae: F1,23 = 202.568, P < 0.001). West Mojave tortoises had similar relationships (humeri: F1,17 = 359.328, P < 0.001; ilia: F1,39 = 1686.098, p < 0.001; scapulae: F1,18 = 718.408, P < 0.001).

The widths of long bones, proximal width (PW), midshaft width (MW) and distal width (DW), were generally similar between adult males and females in both populations and there was no statistically significant sexual dimorphism in bone width. West Mojave females had statistically significantly narrower ilia (PW: F1,30 = 4.341, P = 0.05; MW:

F1,29 = 9.002, P = 0.005; DW: F1,28 = 5.020, P = 0.033) and humeri (MW: F1,19 = 8.058, P

= 0.010; DW: F1,15 = 8.828, P = 0.010) than Sonoran females (Fig. 11). There was no statistically significant difference between populations in male long bone widths (Fig.

12). In juveniles, the only statistically significant difference between West Mojave and

Sonoran tortoises was in humerus PW (F1,22 = 4.461, P = 0.046), with Sonoran juveniles having wider humeri than West Mojave juveniles.

Resorption core diameters

Resorption core diameters (RCDs) were crucial in calculating age estimates. In

Sonoran tortoises, there was no statistically significant difference in RCDs between localities for adult males, females or juveniles. Sonoran females had statistically significantly larger ilial RCDs than males (F1,31 = 5.164, P = 0.03). When the effect of size was removed (ANCOVA), however, there was no sexual dimorphism in ilial RCDs

2 (F1,24 = 1.063. P = 0.313, r = 0.05). There was no sexual dimorphism in RCDs between male and female tortoises from the West Mojave Desert.

Sonoran females had significantly larger RCDs in all bones than West Mojave females (humeri: F1,18 = 6.180, P = 0.023; ilia: F1,24 = 31.017, P < 0.001; scapulae: F1,13

= 6.365, P = 0.025; Fig. 13A). Sonoran males had significantly larger humeri (F1,9 =

6.619, P = 0.03) and ilia RCDs (F1,32 = 4.786, P = 0.036) than West Mojave males (Fig.

13B). I compared juveniles from both groups and found no significant difference in

RCDs between the Sonoran and West Mojave populations.

I grouped adults and juveniles to gain insight into population trends and found a close relationship between body size and RCDs for all bones in both groups (Table 11,

Fig. 14).

Age and longevity

The ilium and humerus consistently yielded the greatest number of periosteal growth layers and for the most part, produced similar age estimates (Table 12 and 13). In

52 juveniles, the Ranking Protocol (RP) and CF method produced similar ages (+ 1 year) between 30 - 120 mm CL in Sonoran juveniles and 40 - 110 mm CL in West Mojave juveniles (Table 14 and 15). Beyond this body size, variation between the two methods increased significantly, especially beyond the reported minimum size at sexual maturity

(around 180 mm CL in both Sonoran and West Mojave tortoises). From this carapace length onward, based on the known-age validation study, I took the RP age to represent the most accurate age estimate for adults (Table 14 and 15).

The individual variation between age and size was considerable (Table 14 and

15). The oldest Sonoran adult males were 47 to 54 years old (unknown CL; 241 mm CL;

266 mm CL). The oldest Sonoran females were 42 to 43 years old (unknown CL; 223 mm CL; 239 mm CL). The oldest West Mojave male (262 mm CL) was 56 years old.

The second oldest male was significantly larger (280 mm CL) but was only 36 years old

(Table 15). The oldest West Mojave female (235 mm CL) was only 27 years old, followed by a much smaller female (198 mm CL) estimated as 26 years old (Table 15).

In the Sonoran Desert sample, I found no statistically significant difference in population age estimates between localities for adult males, females or juveniles. I therefore grouped adult males together and adult females together to ascertain sexual dimorphism. There was no significant difference in age estimates between males and females (Fig. 15B). There was no sexual dimorphism in age estimates between West

Mojave adults either (Fig 15B).

Sonoran adult tortoises reached significantly older ages than West Mojave adult tortoises (Fig. 15B). Sonoran female tortoises reached statistically significantly older ages than West Mojave female tortoises (F1,31 = 66.198, P < 0.001; Fig. 15B). Similarly,

Sonoran males tortoises reached statistically significantly older ages than West Mojave male tortoises (F1,33 = 25.323, P < 0.001; Fig. 15B).

West Mojave tortoises grew faster than Sonoran tortoises and reached adult sizes at younger ages (Fig. 16). In general, Sonoran tortoises of similar sizes to West Mojave tortoises were older in age and were also older at death (Fig. 16). I compared size/age relationships between Sonoran and West Mojave populations and removed the effect of age (covariate) by ANCOVA. West Mojave tortoises had a significantly faster growth than Sonoran tortoises (F = 14.640, P < 0.001, r2 = 0.71). The adjusted least squares mean for West Mojave tortoises was 200.6 + 6.0 SE and for Sonoran tortoises 168.2 +

5.8 SE at the mean age of 21.8 years.

In general, males not only lived longer, but also were larger at similar ages than females (Fig. 17). West Mojave males showed the only statistically significant relationship between age and size (F1,19 = 8.143, P = 0.007). When I compared CL between Sonoran Desert adults but added age as a covariate (ANCOVA), I found no statistically significant difference between the sexes for either Sonoran or West Mojave tortoises (SD: F = 0.867, P = 0.359, r2 = 0.32; WM: F = 1.807, P = 0.191, r2 = 0.26;

Fig.12). The adjusted least squares mean of Sonoran females was 221.8 + 10.7 SE and was 239.1 + 7.1 SE in Sonoran males at the mean age of 23.0 years. The adjusted least squares means in West Mojave females was 221.8 + 10.7 SE and was 239.1 + 7.1 SE in

West Mojave males at the mean age of 29.2 years.

I arranged Sonoran and West Mojave tortoises into size classes to obtain a better idea of size/age relationships and compared variance in age between the two groups using

ANOVA (Fig. 17). I excluded the 40 year old West Mojave juvenile (125 mm CL) from

54 the analysis because it represented an extreme outlier in its size class (101 – 150 mm

CL). Sonoran tortoises were statistically significantly older than West Mojave tortoises within the 51-100, 201-250 and 251-300 mm CL size classes (Table 16, Fig. 17).

Age at sexual maturity

I obtained an estimate of age at sexual maturity by looking at the age estimates of tortoises with carapace lengths similar to published minimum sizes at sexual maturity

(176 mm CL for West Mojave tortoises and around 180 mm CL for Sonoran tortoises).

In the Sonoran sample, three individuals of 175, 180 and 195.5 mm CL, showed ages of

26, 27 and 25 years respectively (mean age was 26 years; Table 14). In comparison, I extrapolated age at maturity (~ 180 mm CL) from the age versus size regression line, and obtained an estimate of around 22 years (Fig. 16), similar to that obtained from the actual data.

In the West Mojave (WM) sample, four individuals of 168, 176, 177 and 178 mm

CL showed ages of 17, 22, 19 and 11 years (Table 15). Mean age was 17 years. When I extrapolated age at maturity from the age/size regression line, I obtained an estimate of around 15 years. Even at similar sizes to Sonoran tortoises, estimated ages were on average lower than those of Sonoran individuals (mean age of West Mojave individuals between 176 to 195 mm CL is 18.7 years; Table 15).

Discussion

Body size and bone morphometry

I found no significant difference in adult or juvenile body or bone sizes between the various Sonoran Desert localities, but this lack of size differentiation could be due to

55 the small population sample sizes that would make any trends difficult to discern. In previous studies at two of the localities from which I received samples, namely Little

Shipp Wash (LSW, Yavapai) and Eagletail Mountains (EM, Maricopa), a distinct sexual dimorphism in body size was found. Murray and Klug (1996) and Averill-Murray (2002) noted that males reached larger average maximum sizes than females (LSW: males = 299 mm, females = 267 mm CL; EM: males = 288, females = 268 mm CL). On the Granite

Hill plots (in Pinal County), they found that sexes attained a similar maximum size

(around 244 mm CL) - significantly smaller than the previous two populations (Murray and Klug, 1996). In contrast, Averill-Murray (2002) found that females reached the same or larger average sizes at three plots south of the Gila River (of which I had carcasses from San Pedro Valley and West Silver Bell Mountains; Table 7). The underlying reasons for these sexual dimorphisms are currently unknown.

In the Sonoran Desert, there was no significant sexual dimorphism in body size or bone length. In the West Mojave Desert, however, males were larger than females

(longer carapace lengths; Fig. 1 and 2). In both groups, CL and bone length were highly related. Mean CL for Sonoran females was 232.2 mm CL, whereas West Mojave females were significantly smaller at 214.5 mm CL. When I compared bone length between adults, humeri and ilia were significantly longer in Sonoran females than in

West Mojave females (Fig. 10). In males, mean CL was similar in both deserts populations (250.9 mm for Sonoran males and 244.7 mm for West Mojave males) as was total length in all bones measured.

The significant sexual dimorphism in body size in West Mojave adults (due to the noticeably smaller sizes of West Mojave females), probably resulted from the energy

56 females channeled into reproduction, a physiological trait directly influenced by resource availability (Peterson, 1996). Mean annual rainfall in the Arizona Upland subdivision of the Sonoran Desert (where most of my sample derived; Table 8) is much more reliable than rainfall in the Mojave Desert (Averill-Murray, 2002). It is about 85 to 300 mm greater than annual rainfall in the Mojave Desert, which receives even less annual rain in the western portion (around 50 to 75 mm per year) than in the eastern portion

(Wallis et al., 1999).

Geographic variation in rainfall is known to correlate with variation in G. agassizii reproductive traits (Wallis et al., 1999). East Mojave females lay eggs at smaller body sizes, lay proportionally smaller eggs, and lay more eggs than West Mojave females (Wallis et al., 1999). Sonoran Desert tortoises may be investing their entire reproductive output in a single clutch laid prior to more predictable rainfall (Averill-

Murray et al., 2002). Rainfall seems to influence mean clutch size and the proportion of females reproducing each year (Averill-Murray et al., 2002b). In dry years, smaller tortoises are less likely to lay eggs than larger ones. Sonoran females lay on average only one clutch per year (Averill-Murray et al., 2002b) and, in drought periods, may not reproduce at all. Wirt and Holm (1997) reported that only two of the six females they studied in the Maricopa Mountains in 1994 laid eggs after almost ten years of drought, whereas all seven female tortoises under observation laid eggs at a nearby site, which apparently was less influenced by drought.

The investment in a single clutch of eggs allows Sonoran females to invest more energy into growth than West Mojave females, especially as annual resource availability appears to be more consistent in the Sonoran Desert. In contrast, West Mojave females

57 generally produce more than one clutch each year, even during periods of drought

(Turner et al., 1986; Wallis et al., 1999). The physiological stress of low food resources would seriously constrain the amount of energy available for growth. In addition, resources available during egg development would generally affect the health and energy input available to female desert tortoises. Annual field metabolic rate (FMR) of female desert tortoises has been positively correlated with the number of eggs laid (Henen,

1997), which indicates that energy expenditures associated with reproduction (e.g. digging nests, producing eggs and increasing foraging effort to gather extra nutrients for producing eggs) are substantial (Henen et al., 1998). Droughts of eighteen months or longer occur regularly in the Mojave Desert (Oftedal, 2002). Absence of rain precludes germination of annuals or re-growth of perennials. At these times, the desert is nearly devoid of food for tortoises, except for some of the smaller, less armored cacti that tortoises can eat and whatever non-woody senescent material that has not disintegrated or blown away, like dried grasses (Oftedal, 2002). In years of low winter rainfall, foraging choices are limited by the small number of plant species that germinate and grow, not to mention that many of them are introduced weedy species (Oftedal, 2002). Alien annual grasses are widespread and abundant in the Mojave Desert, and negative correlations between alien and native annual plants suggest that competition may occur between them

(Brooks, 2000). At two sites in the northeastern Mojave Desert, introduced annual grasses and the introduced filaree accounted for about 80 % of all tortoise feeding bites in a year (1990) with very low biomass production of annuals, but as biomass production increased in subsequent years, these species accounted for only 34-40 % (1991) and 34-

64 % (Esque, 1994). There has been concern that tortoise preference for introduced

58 exotic annuals, such as Mediterranean grass in the Mojave Desert, might lead to nutrient deficiencies and health problems (Avery, 1998). Introduced species have been found in all diet studies in the Sonoran Desert; however, these species accounted for only eight of the 222 food plants identified at Sonoran Desert and Sonoran sites (Van Devender et al.,

2002). Introduced species, therefore, do not seem to impact Sonoran tortoise diets as much as they impact Mojave tortoise diets and this is possibly because alien plant species tend to be prominent in washes (both in species richness and biomass; Brooks, 1999), which are areas generally inhabited by Mojave tortoises and not Sonoran tortoises

(Averill-Murray and Averill-Murray, 2005).

During dry periods between rains, desert tortoises retain urine while suffering chronic negative water balance (Nagy and Medica, 1986, Peterson, 1996) and, if they are simultaneously starving, with concomitant catabolism of protein, urea levels can build up to extraordinarily high levels (Peterson, 1996). According to Peterson (2002), very high blood urea levels in tortoises are as indicative of dehydration as of poor nutrition, if not more so. Except in the driest years, Sonoran tortoises are less likely to be critically stressed by drought because they rehydrate in spring and/or summer, and dried or fresh grasses and dead annuals are generally available (Van Devender et al., 2002). Well- hydrated tortoises achieve positive energy balances and are able to store lipids as potential energy reserves (Peterson, 1996; 2002). Critical physiological stress is much more likely in the uniseasonal Mojave Desert, where the winter-only rainfall presents greater difficulty in eating dried plants in the summer and fall due to longer periods of dehydration (Van Devender et al., 2002). As males do not experience the physiological constraints of egg production and development, they are able conserve more energy

59 and/or focus more energy into growth during stressful conditions (like regular droughts), something practically impossible in West Mojave females if they are gravid.

Resorption core diameters

Resorption core diameter (RCD) gives an indication of bone remodeling, which involves the removal of primary bone (first bone deposited) and thus early growth layers.

There was no significant sexual dimorphism in RCD in either population. When I compared adults between the two groups, Sonoran females showed larger RCDs than

West Mojave females (Fig. 13A) and Sonoran males had larger humeral and ilial RCDs than West Mojave males (Fig. 13B). There was an association between body size and

RCD for all bones in both groups. Sonoran tortoises displayed larger RCDs at similar carapace lengths than West Mojave tortoises (Fig. 14). Resorption core diameters include not only the medullary cavity area, but also any secondary bone deposited, in other words, endosteal bone that was deposited after the primary bone (incorporating initial growth layers) was resorbed. Larger RCDs imply either larger medullary cavity areas and thus higher rates of bone removal, and/or higher rates of secondary bone deposition, usually associated with intracortical remodeling (Castanet and Smirina,

1990). Either way Sonoran tortoises generally had a greater rate of endosteal remodeling than West Mojave tortoises, especially since juveniles start off with similar RCDs at similar sizes (Fig. 14).

Bone was originally seen as a temporary store of calcium and phosphates that remodeling then released back into circulation. However, as Currey (2003) states, bone is being resorbed and deposited at the same time, almost contiguously, so any advantage to the body as a whole must be small. It is now apparent that other processes, like

60 mechanical competence, changing the grain of bone as an adaptation to muscle insertion, or the replacement of dead cells, are more important (Currey, 2003). Non-mechanical factors also need to be taken into account; for example, in a study conducted on Nile monitors, both males and females started off with bones having the same mechanical properties and density, but in females, the endosteal cavity increased in size as they underwent more egg laying cycles (de Buffrénil and Francillon-Viellot, 2001). Females lost bone while producing eggs and did not fully regain it before the next clutch developed.

Clutch production and development could certainly explain the sexual dimorphism in RCDs in Sonoran tortoises due to a need for extra calcium and phosphates. If this were the case, however, I would have expected there to be a sexual dimorphism in West Mojave adults, which there was not. The need for calcium and phosphates also does not explain the tendency toward larger resorption cores in Sonoran females than in West Mojave females. I would have expected, due to the higher number of clutches produced on average in West Mojave populations, that if RCDs were mainly related to the release of ions for egg shelling, West Mojave females would show larger resorption cores. In addition, if increased remodeling promoted ion release due to nutrient deficiencies (resulting from a diet rich in less nutritious exotic species), I would again have expected the West Mojave population to have larger RCDs than Sonoran tortoises. Although there was a trend for larger RCDs in West Mojave males, results were not statistically significant, so I am hesitant to form conclusions based on this explanation.

I propose that larger resorption cores in Sonoran tortoises are potentially a biomechanical adaptation. Most high tortoise densities observed in the Mojave Desert have been within intermountain valleys and flat open land, where friable soils allow for the construction of deep burrows (Germano et al., 1994; McLuckie et al., 1999).

However, in the Sonoran Desert, tortoises are found at the highest densities on steep, rocky hills and desert mountain slopes and are generally absent from or occur at low densities in intermountain valleys and washes (Averill-Murray et al., 2002a; Riedle et al.,

2002; Averill-Murray and Averill-Murray, 2005). The biomechanical constraints of locomotion on mountain slopes and rocky hills may demand greater remodeling in

Sonoran tortoise bone, thus producing larger RCDs than in West Mojave tortoises. In addition, the potential dual foraging period and frequent lack of hibernation in Sonoran tortoises (Averill-Murray et al., 2002; Van Devender, 2002) would cause them to be more active throughout the year than West Mojave tortoises, where drought and winter freezes generally promote hibernation. Burrows serve as thermal refugia (Spotila et al., 1994) and during these stressful periods, Mojave tortoises sustain a much reduced metabolic rate, rarely emerging to feed (Nagy and Medica, 1986; Peterson, 1996).

Longevity and age at maturity

When I compared the number of visible growth layers within the various limb bones samples, I found that the bone with the highest number of growth layers varied

(Tables 12 and 13). From the preliminary skeletal screening I found that the tibiae, fibulae, ulnae and radii showed fewer visible growth layers (i.e. thinner cortical bone) than the bones that I chose for this study. I, therefore, suggest that future skeletochronology studies should involve a skeletal screening where possible, either by x-

62 ray, as in de Buffrénil and Castanet (2000), or preferably by histological sectioning, because my results show that assuming that the humerus or femur shows the greatest number of visible growth layers may be misguided.

Past studies reported that male and female desert tortoises generally reached similar ages (Turner et al., 1987; Germano, 1994; Germano et al., 2002), and this study showed no statistically significant sexual dimorphism in age (Fig. 15). When I compared the two groups, Sonoran tortoises reached significantly older ages than West Mojave tortoises and in general, were older at similar sizes than West Mojave tortoises (Fig. 16 and 17). It is interesting to note, however, that even though Sonoran males were significantly older than West Mojave males, there was no significant difference in body size.

The greatest overlap in age range within size classes (between both groups) occurred before 200 mm CL, or more or less before the onset of sexual maturity in both groups. There was a high degree of variation, however, between ages and size, with some smaller individuals showing much older ages than larger individuals. West Mojave tortoises reached sexual maturity at earlier ages than Sonoran tortoises (around 17 years as opposed to 26 years in Sonoran tortoises).

Growth strategies, ecological implications and conclusions

The most surprising conclusion from this study was that West Mojave tortoises seem to grow faster and reach sexual maturity at earlier ages than Sonoran tortoises (Fig.

16). Favorable conditions support the development of a rapid growth rate and earlier ages of sexual maturity and this has been previously demonstrated in captive desert tortoises (Jackson et al., 1976; 1978). For this reason, I expected Sonoran tortoises to

63 display the faster growth rates and earlier ages at sexual maturity of the two desert populations, mainly because the Sonoran Desert seems to have an environment more favorable to tortoise growth. The longer life-spans of Sonoran tortoises supported this hypothesis, but their delayed sexual maturity and slower growth rates, in comparison to

West Mojave tortoises (supposedly the harsher of the two environments), does not. If in fact the West Mojave is the more productive of the two deserts, why were West Mojave females significantly smaller in body size than Sonoran females and why did West

Mojave tortoises have shorter life-spans than Sonoran tortoises? Lagarde et al. (2001) noted that in steppe tortoises, which live in a similar habitat and climate to West Mojave

Desert tortoises, animals matured earlier if they grew faster, but at smaller body sizes.

One noticeable difference, however, is that female steppe tortoises reached significantly larger body sizes than male steppe tortoises (Lagarde et al., 2001). Wikelski et al. (1997) proposed that rapid growth and early maturity at smaller body sizes may be of benefit due to the low maintenance requirements of small body size. Such energy savings can then be invested into maintenance, promoting reproduction and survival (Wikelski and Thom,

2000). Lagarde et al. (2001) suggested that the earlier maturity and smaller body sizes were especially beneficial to male steppe tortoises because annual activity (3.5 months) was strongly constrained by the harsh environment. Alm (1959) proposed that the early maturity noticed in stunted perch resulted from either a genetically fixed maturation age, or a change in environmental conditions (i.e., from good opportunities for growth during early life to poor conditions after reaching maturity). Jansen (1996) argued that the considerable range in age of sexual maturity seen within and between various populations of perch species, together with the known response of age at first reproduction to

64 environmental factors like temperature and nutrient levels, made a strictly genetic basis for the onset of maturity seem unlikely. Optimal life history strategies allocate resources to maintenance, growth, and reproduction in a way that maximizes individual fitness, i.e., age specific fecundity and survivorship (Congdon and Gibbons, 1990). Stunted perch reproduce earlier during their life in order to decrease risk of mortality before reaching maturity (Jansen, 1996). As a consequence of this strategy of early and high reproductive effort under poor growing conditions, stunted perch experienced poor somatic condition and a reduced life expectancy (Jansen, 1996).

West Mojave females could be optimizing their life history strategies by reproducing at earlier ages and producing more clutches than Sonoran females to compensate for the low juvenile survivorship and comparatively shorter life-spans. The benefit for maturing early is usually manifested through an increased survivorship (by diminishing mortality before maturity) and by augmentation of the number of reproductive episodes, at least in iteroparous species (Hamilton, 1966; Congdon et al.,

1982; Lagarde et al., 2001). The trade-off, however, is that (possibly in response to low juvenile survivorship and maybe even hatching success), females produce at least one, but usually more, clutches per year, and they do this consistently even during the chronic and frequent droughts characteristic of the West Mojave Desert. These frequent droughts, coupled with the West Mojave Desert having the lowest annual rainfall of the entire desert tortoise range, even if it may be more productive following winter rains than either the Sonoran or East Mojave Desert (Wallis et al., 1999), could cause chronic physiological stress, especially in West Mojave females, and thus the resultant life- history strategies that they have developed. Peterson (1996) concluded from his studies

65 in the West and East Mojave Desert of California that the desert tortoise is not physiologically adapted to live in the desert, but is a tenuous relic of a less rigorous climate. This conclusion is consistent with the hypothesis proposed by Van Devender

(2002) that the Mojave tortoises evolved from Sonoran or Sinaloan tortoises in summer rainfall environments.

Age at sexual maturity determined from this study (15 to 17) supports the published data of 9 to 26 years in Mojave tortoises (Medica et al., 1975), but was around or slightly higher than published estimates of 10 to 20 years for Sonoran tortoises. I determined age at maturity to be significantly older in this group, with a conservative estimate of 22 to 26 years. The delayed sexual maturity in Sonoran tortoises could be related to the more extreme summer temperatures (regularly over 38 °C; Van Devender et al., 2002) experienced in this desert. Berrigan and Charnov (1994) suggested that low food quality and/or availability could result in delayed maturity and small body size at maturity, but that a reduction or increase of environmental temperatures (causing a reduction in annual activity) entailed a delayed maturity but a larger body size at maturity. For example, in populations of a perch species faced with elevated water temperatures during times of unfavorable feeding conditions, the fish were observed as adopting an energy-saving strategy. They reduced fecundity or even restrained from reproducing in the year(s) following the year of first reproduction, in order to reduce the high mortality risk associated with the considerable energy investment into gonadal production (Sandström et al., 1995; Jansen, 1996). Delayed sexual maturity could be an energy-saving strategy in Sonoran females, together with the tendency to produce only

66 one clutch or even no clutches at all, possibly as a response to unfavorable environmental conditions (Averill-Murray et al., 2002b).

The estimated longevity in both groups were higher (noticeably so in Sonoran tortoises) than those proposed by Germano (1992; 1994) for West Mojave (32 years) and

Sonoran Desert (35 years) populations. Maximum age estimates were 47 to 54 years in

Sonoran males and 40 to 43 years in Sonoran females. In West Mojave males the oldest age estimate was 56 years. The oldest males, however, were typically 27 to 37 years old.

In West Mojave females, maximum age estimates were only 24 to 27 years. It is significant to note that, irrespective of desert, females had shorter life-spans than males, especially West Mojave females, and this should be considered in future conservation strategies.

Table 7. Population samples of desert tortoises (Gopherus agassizii) from the Sonoran Desert, Arizona.

County Location Sample size Maricopa Eagletail Mountains 7 juveniles

1 adult male

Mohave Beavertail Slope 1 juvenile 7 juveniles East Bajada plot 1 female

1 unknown

1 juvenile Hualapai foothills 3 adult

females 1 adult male 2 juveniles 1 adult Virgin Slope females 2 adult males Yavapai Bonanza Wash 1 adult male 1 juvenile 5 adult Harcuvar females 1 adult males 4 juveniles 5 adult Little Shipp females 2 adult males

Table 7 (continued).

1 juvenile 3 adult Pinal Granite Hills females 1 adult male

4 adult females

San Pedro 4 adult males 1 unknown

sex

4 adult

Tortilla Mountains females

2 adult males

3 adult

Pima West Silverbells females 2 adult males

Table 8. Location, elevation and habitat type for populations of Gopherus agassizii from Arizona. NSD = northern Sonoran Desert; SSD = southern Sonoran Desert.

County Site Elevation Vegetation

Mohave Beavertail Slope 150-900 m Colorado River (NSD) Valley East Bajada plot

Hualapai foothills

Virgin Slope Maricopa Eagletail Mnts. 460-698 m Arizona Upland (NSD) Desert scrub Pinal Granite Hills 600-700 m Arizona Upland (SSD) granite boulder San Pedro 656-853 m habitat

Tortilla Mnts. 760-855 m Yavapai Bonanza Wash 960-1080 m Arizona Upland (SSD) granite boulder Little Shipp Wash 788-975 m habitat Pima West Silverbells 760-855 m Arizona Upland (SSD) vegetation

Table 9. Analysis of variance in bone length between adult Gopherus agassizii from the Sonoran Desert, Arizona and West Mojave Desert, California. df = degrees of freedom.

F P df

Females: Humeri 10.052 0.005 1,18 Ilia 11.700 0.002 1,28 Scapulae 0.880 0.364 1,14

Males: Humeri 0.385 0.550 1,9 Ilia 0.015 0.902 1,34 Scapulae 0.579 0.462 1,12

Table 10. Details of regression lines for carapace length (CL) versus total bone length (TBL) in Gopherus agassizii from the Sonoran Desert, Arizona and the West Mojave Desert, California.

slope intercept r2 CL vs humerus TBL: Sonoran Desert -2.454 0.283 0.96 West Mojave Desert -1.487 0.292 0.95

CL vs ilium TBL: Sonoran Desert -1.021 0.191 0.96 West Mojave Desert -0.957 0.190 0.98

CL vs scapula TBL: Sonoran Desert -9.614e-3 0.140 0.90 West Mojave Desert -0.691 0.147 0.98

Table 11. Regression statistics of carapace length (CL) versus resorption core diameter (RCD) for Sonoran Desert and West Mojave Desert tortoises

CL vs RCD F P r2 Humerus: Sonoran Desert 92.553 <0.001 0.77 West Mojave Desert 50.503 <0.001 0.71 Ilium: Sonoran Desert 135.045 <0.001 0.77 West Mojave Desert 10.682 0.003 0.27 Femur: Sonoran Desert 128.824 <0.001 0.90 West Mojave Desert - - - Scapula: Sonoran Desert 34.421 <0.001 0.62 West Mojave Desert 27.848 <0.001 0.70

Table 12. Age estimation for Sonoran Desert Gopherus agassizii using the Correction Factor (CF) method and Ranking Protocol (RP) method. J = juvenile; F = female; M = male.

Tortoise sex CL Humerus Femur Ilium Scapula ID (mm) age age age age age age age age (RP) (CF) (RP) (CF) (RP) (CF) (RP) (CF) 93-12 J 0 0 93-17 J 35.3 0 0 0 0 0 0 93-15 J 42.2 0 0 0 1 0 1 0 1 94-17 J 0 3 96-24 J 48.4 0 0 91-11 J 50.6 0 0 1 1 0 0 1 2 94-2 J 55.6 2 5 4 3 4 3 2 4 94-25 J 67.3 5 8 6 6 6 5 90-3 J 70.7 5 5 4 6 93-10 J 4 10 7 11 92-1 J 87.9 8 11 10 10 7 7 93-20 J 11 13 91-12 J 93.1 10 11 11 10 1 8 93-9 J 93.5 1 6 6 7 97-18 J 107.4 9 9 7 8 97-2 J 115.8 12 13 96-14j J 13 11 13 93-6 J 124 11 10 11 10 91-2 J 13 94-24 J 133.8 12 14 16 17 12 11

Table 12 (continued).

97-19 J 149 16 10 91-9a J 150 31 20 uk AZ J 154 18 14 22 11 17 15 90-4 J 161.5 30 22 93-17 J 166 17 14 16 10 16 7 91-4 J 23 11 95-15 J 175 22 17 26 13 92-01 M 180 27 16 97-5 M 195.5 19 14 25 16 20 17 91-10 F 202 32 30 26 17 31 16 95-6 F 213 35 25 36 28 91-05 M 218.5 42 21 92-12 M 221 29 16 95-12 F 37 28 95-13 F 222 30 16 96-18 F 223 31 20 39 23 43 23 37 16 90-7 M 39 29 96-21 M 223 42 31 22 91-1b M 224 33 24 90-4 F 225 29 20 27 91-3 F 226 35 28 29 19 31 18 97-15b F 227 30 21 34 17 91-09 F 229 30 17 35 18 31 16 96-13 F 233 33 23 33 20 97-18 F 233 33 24 38 19

Table 12 (continued).

93-21 F 236 30 17 22 11 96-13b F 237 31 27 96-14 F 238.5 29 16 96-19 F 239 31 22 31 21 24 14 97-14 F 239 34 23 42 19 91-11 M 240 33 27 34 21 34 20 91-4 M 241 47 32 95-1 M 252 37 15 30 18 91-1 F 252.5 32 22 24 14 95-16 F 31 21 29 12 32 12 93-10 F 37 24 34 22 97-16b F 257 33 25 39 23 35 18 95-15 F 259 21 28 37 27 91-10 F 262 28 18 54 F 262.5 40 24 33 16 97-17 M 266 47 22 41 20 97-4 M 280 39 26 33 33 96-17 M 280 34 28 36 28 22 13 97-14b M 287 42 29 34 23 33 19 90-5 M 54 31 32 34 90-9 F 43 20

Table 13. Age estimation for West Mojave Gopherus agassizii using the Ranking Protocol (RP) and Correction Factor (CF) method.

Tortoise ID sex CL (mm) Humerus Ilium Scapula age age age age age age (RP) (CF) (RP) (CF) (RP) (CF) O2245 juv 41 0 1 O2272 juv 41 1 2 O2294 juv 44.2 1 2 O2260 juv 49.5 0 1 0 1 O2284 juv 54 1 2 94-1 juv 54 1 2 1 2 91-15 juv 58.9 2 3 95-4 juv 86 4 4 91-53 juv 87.3 6 7 5 6 91-35 juv 95 6 7 91-42 juv 107.5 9 8 7 8 93-N-17 juv 117 17 12 93-1 juv 125 39 40 13 14 RKP-115 juv 151 12 14 AJC 1 juv 154 16 16 RKP-100 juv 163 17 12 A-94-3 juv 167 21 21 93-N-47 juv 167 21 21 93-N-28 male 168 22 17 RKP 102 male 176 22 21 15 13 94-6 juv 177 19 10 16 12

Table 13 (continued).

91-8 male 178 11 13 RKP-109 unk 184 19 21 A-94-2 juv 194 19 11 10 9 RKP302 female 198 26 28 19 20 22 24 RKP11 male 202 24 19 22 17 22 16 91-50 female 205 15 16 93-N-21 female 208 13 13 94-7 male 208 21 16 18 14 95-N-5 male 214 20 15 93-N-59 female 223 19 12 11 13 91-26 female 224 20 14 93-N-30 male 232 15 13 91-36 male 232 26 21 21 23 26 22 RKP2 male 233 22 20 91-37 female 235 27 24 23 17 93-N-29 male 239 17 10 91-34 female 240 24 22 93-N-58 female 245 18 15 20 13 93-N-42 male 248.5 21 21 93-N-7 male 256 16 13 93-N-10 male 256 25 17 91-31 male 262 56 24 93-N-46 male 277 28 24 93-N-44 male 280 32 27 36 27 91-48 male 282 27 22

Table 13 (continued).

93-N-45 male 286 26 21 93-N-35 male 288 23 25 93-N-13 male 290 26 20 CA1 unk 24 18 RKP-110 juv 25 27 23 16 93-N-31 male 24 18

Table 14. Size and age estimates of Gopherus agassizii from the Sonoran Desert, Arizona determined by the Correction Factor (CF) method and Ranking Protocol (RP). Age in bold represents the more accurate age estimate; CL = carapace length; CW = carapace width; PL = plastron length; J = juvenile, M = male; F = female; Unk = unknown sex.

Tortoise Sex Location CL CW PL Age Age ID (mm) (mm) (mm) (GR) (CF) 93-17 J Eagletail Mountains 35.3 30.5 36.1 0 1 93-12 J East Bajada plot 0 0 94-17 J Granite Hills 34.1 42 0 3 93-15 J Eagletail Mountains 42.2 36.1 37.2 0 1 96-24 J Beaver Dam Slope 48.4 48.6 0 0 91-11 J Eagletail Mountains 50.6 45.7 48.4 1 2 94-2 J Eagletail Mountains 55.6 50.7 56.6 4 5 94-25 J Little Shipp 67.3 59.1 62 6 8 90-3 J Little Shipp 70.7 53.3 64.7 5 6 92-1 J Eagletail Mountains 87.9 71.1 84.1 10 11 91-12 J Eagletail Mountains 93.1 74 11 11 93-20 J Eagletail Mountains 11 13 93-9 J East Bajada plot 93.5 73.4 6 7 97-18 J East Bajada plot 107.4 87.6 102.9 9 9 93-10 J East Bajada plot 84.5 104 7 11 97-2 J Virgin Slope 115.8 93.5 117.8 12 13 91-2 J San Pedro 119 13 14 96-14juv J Hualapai foothills 120.3 11 13 93-6 J East Bajada plot 124 104 122 11 10 94-24 J Little Shipp 133.8 100 128 16 17

Table 14 (continued).

97-19 J East Bajada plot 149 16 10 91-9a J San Pedro 150 175 130 31 20 uk AZ J Eagletail Mountains 154 121 145.5 22 15 91-4 J San Pedro 141 23 11 90-4 J East Bajada plot 161.5 131.5 30 22 93-17 J Little Shipp 166 108 17 14 95-15 J San Pedro 175 130 165 26 17 92-01 F Little Shipp 180 137 176.5 27 16 97-5 M Virgin Slope 195.5 145 184.5 25 17 91-10 F Granite Hills 202 151 181 32 30 95-6 F San Pedro 213 157 204 36 28 91-05 M West Silverbells 218.5 157 194.5 42 21 95-12 F West Silverbells 162 195 37 28 92-12 M San Pedro 221 160.5 217 29 16 90-7 M Granite Hills 163 217 39 29 95-13 F West Silverbells 222 165 196 30 16 96-18 F Tortilla Mountains 223 177 219 43 23 96-21 M Tortilla Mountains 223 164 220 42 31 91-1b M San Pedro 224 208 33 24 90-4 F Granite Hills 225 169 210 31 20 91-3 F Hualapai foothills 226 179 203 35 28 97-15b F Harcuvar 227 186 228 34 21 91-09 F West Silverbells 229 177 198 35 18 96-13 F Tortilla Mountains 233 184.5 204 33 23

Table 14 (continued).

97-18 F Harcuvar 233 174 210 38 22 93-21 F Little Shipp 236 183 226 30 19 96-13b F East Bajada plot 237 182 225 31 27 96-14 F Tortilla Mountains 238.5 175 211.5 29 16 96-19 F Tortilla Mountains 239 160 212 31 22 97-14 F Harcuvar 239 178 213 42 23 91-11 M San Pedro 240 182 224 34 27 91-4 M Hualapai foothills 241 175 227 47 32 95-1 M San Pedro 252 179 226 37 18 91-1 F Hualapai foothills 252.5 198 224 32 22 95-16 F San Pedro 230 32 21 93-10 F Harcuvar 193 236 37 24 97-16b F Harcuvar 257 240 39 25 95-15 M West Silverbells 259 184 246 37 28 91-10 F Little Shipp 262 210 237 29 27 54 F Little Shipp 262.5 186 224 40 24 97-17 M Harcuvar 266 191 240 47 22 97-4 M Virgin Slope 280 202.5 255 39 33 96-17 M Tortilla Mountains 280 36 28 97-14b M Bonanza Wash 287 42 29 90-9 F Little Shipp 242 43 20 90-5 M Little Shipp 265 54 34

Table 15. Age estimates of Gopherus agassizii from West Mojave Desert, California determined by the Correction Factor (CF) method and Ranking Protocol (RP). Age in bold represents the more accurate age estimate; CL = carapace length, PL = plastron length.

Tortoise Sex CL CW PL Age Age ID (mm) (mm) (mm) (RP) (CF) O2245 Juvenile 41 28 30 0 1 O2272 Juvenile 41 28.6 1 2 O2294 Juvenile 44.2 28.1 33.9 1 2 O2260 Juvenile 49.5 37 46.2 0 1 O2284 Juvenile 54 42 50 1 2 94-1 Juvenile 54 45 54 1 2 91-15 Juvenile 58.9 51.3 61.5 2 3 95-4 Juvenile 86 71 82 4 4 91-53 Juvenile 87.3 75.4 81.1 6 7 91-35 Juvenile 95 72.9 86.1 6 7 91-42 Juvenile 107.5 88.9 104 9 8 93-N-17 Juvenile 117 97 110 19 12 93-1 Juvenile 125 103 121 39 40 RKP-115 Juvenile 151 12 14 AJC 1 Juvenile 154 16 16 RKP-100 Juvenile 163 17 12 A-94-3 Juvenile 167 154 21 21 93-N-47 Juvenile 167 133 161 21 21 93-N-28 Male 168 22 17 RKP 102 Male 176 22 21 94-6 Juvenile 177 139 165 19 12

Table 15 (continued).

91-8 Male 178 140 163 11 13 RKP-109 Unknown 184 19 21 A-94-2 Juvenile 194 19 11 RKP302 Female 198 150 186 26 28 RKP11 Male 202 147 182 24 19 91-50 Female 205 156.5 180 15 16 93-N-21 Female 208 162 188 13 13 94-7 Male 208 166 198 21 16 95-N-5 Male 214 20 15 93-N-59 Female 223 183 208 19 13 91-26 Female 224 190 214 20 14 93-N-30 Male 232 180 217 15 13 91-36 Male 232 179 218 26 23 RKP2 Male 233 22 20 91-37 Female 235 193 225 27 24 93-N-29 Male 239 17 10 91-34 Female 240 197 226 24 22 93-N-58 Female 245 20 15 93-N-42 Male 248.5 208.5 245.5 21 20 93-N-7 Male 256 208 235 16 13 93-N-10 Male 256 226 264 25 17 91-31 Male 262 210 231 56 24 93-N-46 Male 277 216 250 28 24 93-N-44 Male 280 230 271 36 27 91-48 Male 282 213 252 27 22

Table 15 (continued).

93-N-45 Male 286 228.5 269.5 26 21 93-N-35 Male 288 235 275 23 25 93-N-13 Male 290 275 26 20 CA1 Female 24 18 RKP-110 Juvenile 25 27 93-N-31 Male 24 18

Table 16. Analysis of variance in age within size classes between Gopherus agassizii from the Sonoran Desert, Arizona and West Mojave Desert, California; CL = carapace length. Values in bold represent a statistically significant difference between size classes from the two localities.

Size class (mm CL) F P df r2

0 – 50 0.272 0.616 1,8 0.03 51 – 100 16.676 0.001 1,15 0.53 101 – 150 1.574 0.236 1,11 0.13 151 – 200 2.763 0.116 1,16 0.15 201 – 250 90.627 <0.001 1,35 0.72 251 – 300 7.401 0.012 1,22 0.25

300 female CL vs CW y = 0.74x + 4.79 (r ² = 0.74) A 280 female CL vs PL y = 0.71x + 47.58 (r ² = 0.72) male CL vs CWy = 0.66x + 14.90 (r² = 0.97) 260 male CL vs PLy = 0.80x + 30.75 (r ² = 0.90) 240

220 200

180 160 Shell measurements (mm)

140

120300 female CL vs CW y = 1.19x - 85.23 (r ² = 0.96) B 280 female CL vs PLy = 1.17x - 52.91 (r ² = 0.94) male CL vs CW y = 0.96x - 36.94 (r ² = 0.87) 260 male CL vs PLy = 0.96 - 4.15 (r ² = 0.91) 240

220 200

180 160 Shell measurements (mm)

140 120 160 180 200 220 240 260 280 300

Carapace Length (mm)

Figure 6. Regression of carapace length (CL) versus carapace width (CW) and plastron length (PL) in adult male and female Gopherus agassizii from A) the Sonoran Desert, Arizona and B) the West Mojave Desert, California. Solid lines = adult females, broken lines = adult males.

350 A 300

250

200

150

100 Shell measurements (mm) 50

3500 B 300

250

200

150

100 Shell measurements (mm) 50 F M F M F M 0

Carapace Length Carapace Width Plastron Length

Figure 7. Shell measurements of adult male and female Gopherus agassizii from A) the Sonoran Desert, Arizona and B) the West Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers.

280 CL vs CW (SD) A 260 CL vs PL (SD) CL vs CW (WM) 240 CL vs PL (WM)

220

200

180

160 Shell measurement (mm) 140

120 160 180 200 220 240 260 280 300 CL vs CW (SD) B 280 CL vs PL (SD) CL vs CW (WM) 260 CL vs PL (WM) 240 220

200 180

Shell measurement (mm) 160

140 120 160 180 200 220 240 260 280 300

Carapace length (mm)

Figure 8. Regression of carapace length (CL) versus carapace width (CW) and plastron length (PL) in A) adult female and B) adult male Gopherus agassizii from the Sonoran Desert (SD), Arizona and the West Mojave (WM) Desert, California. Solid lines represent Sonoran Desert tortoises, broken lines represent West Mojave tortoises.

180 CLvsCW (SD)y = 0.69x + 9.65 (r ² = 0.97) 160 CL vs PL (SD)y = 0.91x + 3.79 (r ² = 0.99) CL vs CW (WM)y = 0.86x - 5.65 (r ² = 0.99) 140 CL vs PL (WM)y = 0.98x - 3.54 (r ² = 0.99)

120

100

40 Carapace Width/Plastron Length (mm) 20 20 40 60 80 100 120 140 160 180 200

Carapace Length (mm)

Figure 9. Regression of carapace length versus carapace width (CW) and plastron length (PL) in juvenile Gopherus agassizii from the Sonoran Desert (SD), Arizona and the West Mojave (WM) Desert, California. Solid lines represents SD, broken lines represent WM adults.

100 A

40 Bone Length (mm) 20

1000 B

40 Bone Length (mm) 20

F M F M F M 0

Humeri Ilia Scapulae

Figure 10. Bone lengths for adult male (M) and female (F) Gopherus agassizii from A) the Sonoran Desert, Arizona and B) the West Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers. Numbers above box plots represent sample sizes.

10 Proximal width (mm) 5

14140

4 Midshaft width (mm) 2 SD WM SD WM SD WM 400

Humeri Ilia Scapulae

Figure 11. Bone widths for adult female Gopherus agassizii from the Sonoran Desert (SD), Arizona and West Mojave Desert (WM), California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers.

400

Distal width (mm) 10

SD WM SD WM SD WM 0 Humeri Ilia Scapulae

Figure 11 (continued).

10 Proximal width (mm) 5

14120

Midshaft width (mm) 2 SD WM SD WM SD WM 400 Humeri Ilia Scapulae

Figure 12. Bone widths for adult male Gopherus agassizii from the Sonoran Desert, Arizona and West Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers.

400

Distal width (mm) 10

SD WM SD WM SD WM 0 Humeri Ilia Scapulae

Figure 12 (continued).

6 A 5

1 Resorption core diameter (um)

06 B 5

1 Resorption core diameter (um)

SD WM SD WM SD WM 0 Humeri Ilia Scapulae

Figure 13. Resorption core diameters for adult A) female and B) male Gopherus agassizii from the Sonoran Desert, Arizona and the West Mojave Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers.

6 CL vs RCD (SD) y = 0.014 + 0.167 (r ² = 0.77) A CL vs RCD (WM) y = 0.010 + 0.136 (r ² = 0.71) 5

Humerus Resorption Core DIameter (um) 0 0 50 100 150 200 250 300 Carapace Length (mm) 6 CL vs RCD (SD) y = 0.014x + 0.036 (r ² = 0.77) CL vs RCD (WM) y = 0.001x + 0.725 (r ² = 0.27) 5 B 4

1 Ilium Resorption Core Diameter (um) 0 0 50 100 150 200 250 300 350

Carapace Length (mm)

Figure 14. Regression of carapace length versus resorption core diameters in A) humeri and B) ilia for Gopherus agassizii from the Sonoran Desert, Arizona (AZ) and the West Mojave (WM) Desert, California. Solid lines represent the SD sample and dashed lines WM sample.

WM juveniles

SD juveniles

WM males

WM females

SD males

SD females

0 10 20 30 40 50 60

Age (years)

Figure 15. Correction Factor age estimates for A) juvenile and Ranking Protocol age estimates for B) adult Gopherus agassizii from the Sonoran Desert (SD), Arizona and the West Mojave (WM) Desert, California. Horizontal lines within each box represent the median value and whiskers represent the 10th and 90th percentile. Dots represent outliers.

350 Sonoran Desert y = -0.07x2 + 8.55x + 29.199 (r ² = 0.92) 2 300 West Mojave y = -0.16x + 12.18x + 32.48 (r ² = 0.75)

250

200

150

100 Carapace length (mm)

0 0 10 20 30 40 50 60

Age (years)

Figure 16. Regression of skeletochronology age estimates versus carapace length for Gopherus agasiizii from the Sonoran Desert, Arizona (solid line) and the West Mojave Desert, California (dashed line).

300 females y = 1.574x + 178.938 (r ² = 0.22) A 280 malesy = 1.890x + 175.075 (r ² = 0.29)

260

240

220

200 Carapace length (mm)

180

160 0 10 20 30 40 50 60 320 females y = 1.479x + 187.146 (r ² = 0.10) B 300 males y = 1.753x + 198.853 (r ² = 0.18)

280

260

240

220

Carapace length (mm) 200

180

160 0 10 20 30 40 50 60

Age (years)

Figure 17. Regression of Ranking Protocol age estimates versus carapace length for adult female and adult male Gopherus agasiizii from the A) Sonoran Desert (SD), Arizona and the B) West Mojave (WM) Desert, California.

100

WM w 251-300 SD s

201WM-250 w SD s

151WM-200 w SD s w 101WM-150 SD s

WM w

Size Classes (mm CL) 51-100 SD s WM w 0-50 SD s

0 10 20 30 40 50 60

Age (years)

Figure 18. Box plots of age range within size classes for Gopherus agassizii from the Sonoran Desert (SD), Arizona and the West Mojave (WM) Desert, California. Horizontal lines within the box plots represent the median age and whiskers the 10th and 90th percentile. Outliers are represented by black dots; w = West Mojave, s = Sonoran Desert.

101

CHAPTER 4. AGE ESTIMATES AND GROWTH STRATEGIES IN EAST MOJAVE DESERT TORTOISES (GOPHERUS AGASSIZII).

Introduction

Turtles are among the longest-lived vertebrates and concepts related to aging are of obvious interest because they have a substantial impact on life history evolution.

Accurate assignment of age in individuals is important for determining age at first reproduction, age specific survivorship and fecundity, generation time, and other life history attributes essential to understanding the potential for a population to respond when its numbers are low (Congdon et al., 1982; Frazer et al., 1990; Berry, 2002). Desert tortoises (Gopherus agassizii) are listed as threatened in the Mojave Desert portion of its range, due to declines in population sizes (U.S. Fish and Wildlife Service, 1994).

Juvenile mortality is high in the Mojave (averaging 98 %) even though adult females lay one to three clutches per year with 4.5 to 6.7 eggs per clutch (Turner et al., 1986;

Germano, 1994; Rostal et al., 1994). Female tortoises as small as 176 mm CL are sexually mature and minimum age at first reproduction may be as low as 9 years, with estimates of up to 26 years (Medica et al., 1975; Germano, 1994). Germano (1992) proposed that East Mojave tortoises reached ages of 48 to 52 years but also stated that these estimates, although based on wild animals, were only minimum values because of the uncertainty of the accuracy of the estimates (Germano, 1994). East Mojave tortoises have also been estimated to reach sexual maturity at an average minimum age of 15.4 years (range of 12 – 19 years; Germano, 1994) or 17.5 years (range of 9 – 26 years;

Turner et al., 1987). My goal was to try and obtain more accurate estimates of age at sexual maturity and age at death within East Mojave Desert tortoise populations. Using

102 known-aged tortoises, I demonstrated that bone growth layers are produced annually in desert tortoise skeletons, thereby producing growth marks that can be used to estimate individual ages (Chapter two). I conducted a skeletochronological analysis of East

Mojave Desert tortoises using carcasses found in the wild to estimate age at sexual maturity and age at death, two life history traits that are essential baseline data required for the conservation of this threatened species.

Materials and methods

The East Mojave Gopherus agassizii population sample consisted of 60 tortoises.

I collected 25 tortoise carcasses from the Desert Tortoise Conservation Center (DTCC) in

Clark County, Nevada; 7 juveniles, 5 adult females, 3 adult males and 9 adults of unknown sex. These were wild tortoises that had all originally come from localities within Clark County, Nevada. The remaining 35 tortoise carcasses were collected from

Ivanpah Valley in the Mojave National Preserve, California; 12 juveniles, 11 adult females, 8 adult males and 4 adults of unknown sex. Therefore, in total, the East Mojave

Desert tortoise sample consisted of 18 juveniles, 16 adult females, 11 adult males and 15 adults of unknown sex (these skeletal samples either had no shells to determine sex or the shells were partially disintegrated).

I recorded carapace length (CL), carapace width (CW) and plastron length (PL) where carcass shells were intact, using manual Vernier calipers (+ 0.5 mm, Table 17). In an initial study, I sectioned all appendicular skeletal elements, together with vertebrae, gular, marginal, neural and plastron bones to determine which elements showed the greatest number of growth marks (GMs). The humerus, femur, ilium and scapula were

103 the long bones that I used for the skeletochronological analysis. I recorded longitudinal length (LL), proximal width (PW), midshaft width (MW) and distal width (DW) from all long bones using digital calipers (+ 0.05 mm; Table 18).

I measured resorption core diameters in the midshaft region of all long bone cross sections at 10X magnification using a stage micrometer. I defined the resorption core as the medullary cavity (usually filled with spongy bone) plus any endosteal bone (primary and/or secondary bone deposited from the endosteum inward (see Chapter one, Fig. 1).

In chapter two I compared skeletochronology age estimates (determined by three methods) to the known ages of a group of desert tortoises. From these data, I concluded that the Correction Factor method provided the most accurate juvenile age estimates and the growth layer Ranking Protocol (henceforth called Ranking Protocol) the most accurate adult age estimates. I applied these two skeletochronology methods to determine juvenile and adult age estimates for East Mojave tortoises.

The Ranking protocol involves assigning ages to growth mark diameters and ranking them into age classes (see Zug, 1990 and Zug and Parham, 1997). The

Correction-Factor (CF) method is described by Zug and Parham (1997). Hatchlings and small juveniles are essential for both methods.

I compared shell size of adults and juveniles from the Desert Tortoise

Conservation Center (DTCC) to adults and juveniles from Ivanpah Valley (Mojave

National Preserve) via analysis of variance (ANOVA) to ascertain whether the two localities could be grouped to represent a single population sample. I compared the various shell and bone measurements and age estimates between adult males and females to ascertain any significant sexual dimorphism using ANOVA. Statistical analyses were

104 conducted using the SYSTAT (Version 9.0) statistical package. Probability values less than 0.05 were considered statistically significant.

Results

Body size

All juveniles from Ivanpah were smaller than 100 mm CL. One large juvenile

(133 mm CL) from the DTCC caused the group comparison to be significantly different for all shell measurements. When I only compared juveniles less than 100 mm CL between the two groups, results were not statistically significantly different (CL: F1,11 =

0.080, P = 0.782; CW: F1,11 = 0.570, P = 0.466; PL: F1,11 = 0.078, P = 0.785). I also found no statistically significant difference in body size between females (CL: F1,9 =

0.256, P = 0.635; CW: F1,12 = 0.194, P = 0.667; PL: F1,13 = 0.880, P = 0.365) and between males from the two localities (CL: F1,12 = 0.146, P = 0.710; CW: F1,11 = 0.501, P

= 0.246; PL: F1,9 = 3.916, P = 0.079; Fig. 19).

I compared body size between the sexes (Fig. 20 and 21) and found that adult males reached significantly larger sizes than adult females (CL: F1,23 = 5.905, p = 0.023;

CW: F1,25 = 8.912, p = 0.006; PL: F1,24 = 15.829, p = 0.001). When I removed the effect of CL on CW and PL by analysis of covariance (ANCOVA), I found that PL showed the

2 only statistically significant difference between the sexes (F1,19 = 7.952, P = 0.01, r =

0.86). The adjusted least squares mean for PL at the mean CL (247.9 mm) was 208.8 +

2.9 for females and 220.9 + 2.9 for males.

Mean juvenile carapace length (CL) was 64.6 mm (+ 7.3 S.E.), carapace width (CW) was

58.7 mm (+ 6.0 S.E.) and plastron length 70.6 mm (+ 8.3 S.E.).

105

Bone morphometry

In East Mojave adults, males had significantly longer bones than females (Fig. 22; humeri: F1,10 = 11.076, P = 0.008; ilia: F1,24 = 14.833, P = 0.001; scapulae: F1,9 = 8.309,

P = 0.018). When I removed the effect of body size on bone length (ANCOVA), males

2 still showed significantly longer bones than females (humeri: F1,6 = 11.243, P = 0.015, r

2 2 = 0.91; ilia: F1,21 = 7.461, P = 0.013, r = 0.86; scapulae: F1,6 = 15.400, P = 0.008, r =

0.92). Femora were excluded from the analysis due to a small sample size (Fig. 23). I grouped adults and juveniles together and compared bone length to body size. There was a strong relationship between bone length and CL for all bones (Fig. 23).

I compared the various bone width parameters, namely proximal width (PW), midshaft width (MW), and distal width (DW) between adult males and females. Males had larger bones than females and all bone widths (except scapula midshaft width) showed a significant sexual dimorphism (Table 19, Fig. 24). When I removed the effect of body size on bone width by analysis of covariance, males still had significantly wider

2 bones that females except for ilial MW (F1,22 = 3.338, P = 0.081, r = 0.78), scapular MW

2 2 (F1,6 = 0.007, P = 0.935, r = 0.07) and ilial DW (F1,22 = 2.880, P = 0.104, r = 0.68).

Resorption core diameters

Adult tortoises had larger resorption cores than juveniles. In adults, humeri showed the only significant sexual dimorphism, with males having larger humeral RCDs than females (F1,11 = 4.849, P = 0.05; Fig. 25). I regressed resorption core diameters

(RCDs) against carapace length and generally found a positive relationship between body size and RCDs for most bones. The exceptions were female humeri and scapulae, where

RCDs seemed to decrease with increasing size (Fig. 26). Males generally reach larger

106 sizes with larger corresponding RCDs than females (Fig. 26). The only statistically significant relationship between body size and RCDs, however, was in juvenile and male ilia (Table 20; Fig. 26).

Age and longevity

The humeri and ilia in East Mojave tortoises generally displayed the greatest number of visible growth layers. In general, Ranking Protocol (RP) and Correction

Factor (CF) age estimates were similar in juveniles, but at the onset of sexual maturity (~

180 mm CL), RP estimates were higher than CF estimates (Table 21 and 22). At 180 mm

CL and greater, the RP age was the most accurate age estimate for adults, whereas CF age was most accurate for juveniles (Table 22). I compared age estimates between adults and found that males reached significantly older ages than females (F1,25 = 4.741, P =

0.039). When I removed the effect of size (carapace length) on age by analysis of

2 covariance, I found no sexual dimorphism in age (F1,20 = 1.035, P = 0.321, r = 0.38)

The oldest adult male (59 years old) was also one of the largest tortoises at 270 mm CL. He was followed in age by a 46 and 45 year old male, only one year apart, but

244 and 285 mm CL respectively. The oldest adult female was estimated as 39 years

(258 mm CL), followed by a 36 and 35 year old female (unknown CLs).

I compared age to body size (carapace length) to view growth variation (Fig. 28).

There was a positive relationship between body size and age, so generally as size increased age also increased (Fig. 10). Juveniles and adult females showed the only statistically significant correlation between age and size (Table 23; Fig. 29).

To obtain a better idea of size/age relationships, I arranged individuals into size classes and determined the range in age within each class (Fig. 30). There is some

107 overlap in age estimates within the size classes, but this mainly occurs in tortoises > 150 mm CL (Fig. 30).

Age at sexual maturity

To obtain an age estimate for the onset of sexual maturity, I looked at the age estimates of those tortoises with carapace lengths similar to published minimum sizes at sexual maturity (around 180 mm CL in females). Two females of 178 and 182 mm CL respectively, showed ages estimates of 19 and 26 years respectively (Table 22). This produced an average of 22.5 years. For comparison, I interpolated age at sexual maturity from the age/size regression line (Fig. 28) and obtained an estimate of around 19 years, similar to that obtained from the data. There is no published age or size at sexual maturity for East Mojave males.

Discussion

Body size and bone morphometry

Previous studies (e.g. Woodbury and Hardy, 1948) have shown a sexual dimorphism in body size within Mojave Desert tortoise populations, and this study provides further support for those data. Adult males reached significantly larger body sizes (both in shell and limb length) than adult females (Fig. 19 to 21). Mean female body size was 223.3 mm CL (+ 8.0 SE) and mean male body size was 246.7 mm CL (+

5.6 SE). Males also had longer and thicker bones than females and most bone widths showed a significant sexual dimorphism (Fig. 24).

Males had significantly larger humeral resorption core diameters (RCDs) than females and also showed a significantly positive relationship between ilia RCDs and

108 body size. The larger RCDs exhibited by males and the general increase in RCDs with increasing size implies a greater bone remodeling rate in males than in females. Henen et al. (1998) recorded higher field metabolic rates (FMR) in male desert tortoises from three different sites in the Mojave Desert than in corresponding females (especially throughout most of the warm season when growth was most pronounced), which they expected since male home ranges are much larger than those of females (Berry, 1986; O’Connor et al.,

1994). Males tend to court and attempt to mate with females at any opportunity (Rostal et al., 1994), and thus spend more energy actively seeking out mates than do females

(Henen et al., 1998). The larger body sizes and higher FMRs, coupled with the higher activity and larger home ranges (therefore more movement) in males, could have resulted in increased bone remodeling as a biomechanical adaptation (Currey, 2003), producing larger RCDs in males than in females. Bone remodeling has been linked to physiological and biomechanical effects, both of which are mainly influenced by environmental constraints (Castanet et al., 1993; Currey, 2003). For example, in the agile, long-limbed lizard Pseudocordylus capensis and heavily-armored lizard Cordylus cataphractus, the bone remodeling process seemed to correlate with the locomotor performance abilities of each species (Curtin et al., 2005). In P. capensis, intensive localized endosteal resorption, together with limited endosteal and periosteal bone deposition, created a relatively thin-boned, light skeleton, contributing to the speed and agility required by this active predator. In contrast, in C. cataphractus, the genetically determined basal pattern of remodeling was obviously masked by epigenetic processes, in that areas of intense resorption were also areas of intense deposition (Curtin et al., 2005). The regular cycles of overlapping resorption and deposition may have been related to cycles of food scarcity

109 and abundance and the concomitant mobilization of and storage of minerals in bone

(Currey, 2003; Curtin et al., 2005). The overall pattern of growth in C. cataphractus resulted in relatively thick bone walls and small medullary cavities, creating a heavier skeleton to support body armor (thick osteoderms and spinose scales). It also tied in with the relatively shorter limbs and low agility of C. cataphractus, which interestingly also had the lowest basal metabolic rate of any cordylid species studied to date (Curtin et al.,

2005). Sexual dimorphism in endosteal resorption was also observed in C. cataphractus, in which the relative medullary cavity area was significantly larger in females than in males (Curtin et al., 2005). In desert tortoises, RCD measurements incorporated both endosteal resorption and endosteal bone deposition, and my initial observations seemed to indicate more intensive resorption and endosteal bone deposition in tortoise males than in females. This phenomenon will be addressed quantitatively in a future study.

Growth strategies, ecological implications and conclusions

There was no sexual dimorphism in age estimates once the effects of size were removed. Males appeared to live longer than females. The oldest males were 10 to 20 years older than the oldest females (Fig. 27). To make the size/age comparisons between males and females more informative and to increase sample size, I added size (plastron length) and age data obtained from Turner et al. (Table 1; 1987) - a cohort of known-aged

East Mojave Desert tortoises - to the size (plastron length) and age data of males and female tortoises obtained from this study (Fig. 31). Males and females display similar growth patterns although older males seem to have a slightly higher growth rate than older females. Sexual dimorphism in size and age at maturity in many turtle species suggests the existence of a dual adaptive strategy of the sexes (Gibbons et al., 1981).

110

Unfortunately there are no published estimates of age at sexual maturity in East Mojave male tortoises, but both sexes are assumed to be sexually mature at 180 mm CL (Duda et al., 1999). To obtain an age estimate for the onset of sexual maturity, I looked at the age estimates of those tortoises with carapace lengths similar to published minimum sizes at sexual maturity (around 180 mm CL in females). Three females of 178, 182 and 205 mm

CL respectively, showed ages estimates of 19, 26 and 23 years respectively (Table 22).

This produced an average of 22.7 years. For comparison, I determined age at sexual maturity from the age/size regression line (Fig. 27) and obtained an estimate of around 19 years, similar to that obtained from the data. A Rock Valley adult female of 189 mm CL, considered to be part of the East Mojave Desert tortoise assemblage, had a known age of

15 years (Chapter two, Table 4), and with this female included in my analysis, the average age at sexual maturity for females became 20.8 years at an average size of 188.5 mm CL. The average age at sexual maturity was higher than those proposed by Turner et al. (1987) and Germano (1994), but it is within the ranges proposed by both studies.

The potential for different adaptive strategies in East Mojave male and female tortoises may exist, especially considering the higher field metabolic rates (FMR) found in males as opposed to females throughout their active season (Henen et al., 1998).

Annual FMR of females has been positively correlated with reproductive output which suggests that females expend a substantial amount of energy during the spring and summer to produce clutches (Henen, 1997). This energy expenditure in the face of short term droughts, and the resultant scarcity of food and water, promotes poor body condition

(Peterson, 1996, Henen et al., 1998), which may cause the younger ages at death seen in

East Mojave females as opposed to males (Avery, in prep.).

111

The estimated adult survivorship of G. agassizii from the California portion of its range is estimated to be high (Berry, 1986; U.S. Fish and Wildlife Service, 1994).

Estimates were obtained during three to nine year studies of 14 sites (Berry, 1986) and indicated that mortality was low in the eastern Mojave Desert and moderate to high in the western Mojave Desert (Berry, 1986; Corn, 1994). The abilities of desert tortoises to cope with the exigencies of their environment are occasionally pushed to the limit however. Peterson (1996) found that several tortoises died at Ivanpah Valley during the summer of 1990 apparently from drought-related dehydration and starvation. Avery (in. prep.) also documented a high degree of mortality in Ivanpah Valley, which appeared to corresponded to an elevation and rainfall gradient. There was significantly higher adult mortality in the lower elevation site, most likely due to significantly less annual rain

(both in wet and drought years; Fig. 32) and thus fewer resources, than in the higher elevation site. Most of the carcasses that we collected in Ivanpah Valley for this study came from the lower elevation site, including all the juveniles and 16 adults. We were only able to find 3 carcasses in the higher elevation site (all adults; Table 22). There was no distinction in size class or age group in the carcasses found, with the exception of juveniles, which were all a year or less in age. Peterson (1994) concluded that Mojave

Desert tortoise populations are under continuing selection pressure for conservation of water and tolerance of anhomeostasis.

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Table 17. Shell measurements of Gopherus agassizii from two localities in the East Mojave Desert: the Desert Tortoise Conservation Center (DTCC) in Nevada and Ivanpah Valley, Mojave National Preserve (MNP) in California; CL = carapace length, CW = carapace width, PL = plastron length.

ID Locality sex CL (mm) CW (mm) PL (mm) DTCC 19 DTCC juvenile 65.6 56.4 65.2 DTCC 7 DTCC juvenile 133 101 125 DTCC 8 DTCC juvenile 117 DTCC 10 DTCC juvenile 109 137 DTCC 18 DTCC juvenile 62.6 59 57.4 LC04 DTCC female 182 157 DTCC 11 DTCC female 251 186 231 DTCC 13 DTCC female 259 191 228 DTCC 15 DTCC female 205 DTCC 17 DTCC male 220 174 LC03 DTCC male 223 175 202 DTCC 6 DTCC male 226 180 213 DTCC 12 DTCC male 270 193 230 LC02 DTCC male 285 220 IVLP 16 Ivanpah juvenile 42 37.2 41.6 IVLP 17 Ivanpah juvenile 42.2 38.8 40.7 IVLP 14 Ivanpah juvenile 44.4 41 45.5 IVLP13 Ivanpah juvenile 45 40.7 43.5 IV09 Ivanpah juvenile 47 47 48.4 IVLP 15 Ivanpah juvenile 51.2 43.6 50.4 IV07 Ivanpah juvenile 64.3 55.7 61.4 IVLP 12 Ivanpah juvenile 65.2 55 62.5

113

Table 17 (continued).

IV06 Ivanpah juvenile 79.5 63.8 72.1 IVLP3 Ivanpah juvenile 97.9 73.5 91.6 AK1 Ivanpah female 178 138 168 R-11 Ivanpah female 205 150 189 IVLP11 Ivanpah female 205 153.5 189 R-07 Ivanpah female 160 190 IV02 Ivanpah female 171 196 IVLP 7 Ivanpah female 211 165 202 R-03 Ivanpah female 228 172 207 R-76 Ivanpah female 238 188 217 IVPH5 Ivanpah female 181 212 ABFX Ivanpah female 240 181 215 CIMA 5 Ivanpah female 258 226.5 197 IVPH3 Ivanpah male 230 179 224 IVPH3 Ivanpah male 230 179 224 IVA3 Ivanpah male 232 208 224 IVPH 2 Ivanpah male 242 200 233 IVLP 10 Ivanpah male 243 190 227 IV05 Ivanpah male 244 194 225 CIMA 1 Ivanpah male 244 198 226 IVPH 1 Ivanpah male 257 IVPH 4 Ivanpah male 270 205 248 IV08 Ivanpah male 285 215 261

114

Table 18. Bone length and width measurements for Gopherus agassizii from the East Mojave Desert (CA and NV); J = juvenile, F = female, M = male; Unk = unknown sex; H = humerus, I = ilium, F = femur, S = scapula.

Tortoise ID sex CL Total Bone Length Proximal Width Midshaft Width Distal Width (mm) (mm) (mm) (mm) (mm) H I F S H I F S H I F S H I F S IVLP 16 J 42 7.4 9.2 3.4 3.1 1.3 0.9 3.9 3 IVLP 17 J 42.2 6.4 8.8 3.1 3.1 1.3 0.9 3.7 2.7 IVLP 14 J 44.4 1.4 1.4 3.9 3.5 IVLP 15 J 51.2 9.5 11.5 3.9 4 1.4 1.1 4.8 3.2 DTCC 18 J 62.6 10 8 4.2 2.4 1.2 1.1 5.1 6.5 IVLP 12 J 65.2 1.7 4.2 DTCC 19 J 65.6 10.2 13.6 4.5 4.3 1.5 1.2 4.9 4.3 IV06 J 79.5 11.5 5.8 1.5 IVLP 3 J 97.9 2.3 9.6 DTCC 7 J 133 34.5 21.8 17 11.3 9.9 6.8 4 3.9 3.7 11.4 12.3 13.1 DTCC 8 J 18.5 8.1 2.8 10.6 DTCC 10 J 35.6 21.2 17.4 12.8 9.5 6.2 3.7 3.5 3.7 12.6 13 16.7

114

115

Table 18 (continued).

DTCC 1 J 20.9 29.2 9.3 10.5 4 3.4 11.2 11.1 DTCC 16 J 32 16.2 8.9 11.2 9 4.6 2.7 2.7 2.5 11.2 8.4 10.1 DTCC 6 M 226 68.5 43.3 32.8 25.1 20.2 12.2 7.4 6.9 6.7 24.3 23.3 25.4 IVPH 3 M 230 42.4 19.3 5.6 21.5 IVA3 M 232 48.3 20.9 7.1 27.7 IVPH 2 M 242 41.2 22 6.8 29.4 IVLP 10 M 243 45.5 19.5 6.5 24.7 IV05 M 244 67.8 44.1 33.4 26.7 17.1 13.7 7.6 6.9 7.8 24.8 25.5 25.3 CIMA 1 M 244 43.9 22.5 7.2 26.6 IVPH 1 M 257 44.4 21.2 6.6 24.6 DTCC 12 M 270 76.7 48.2 39.2 26.6 19.2 13.5 7.8 7.7 7.8 25.9 28.5 33.8 IVPH 4 M 270 48.5 28 8.6 34.4 IV08 M 285 50.7 22.8 8.1 27 LC02 M 285 85.3 50.9 28.6 23.9 7.9 8.6 29.3 31.9 A-01 Unk 35.7 15 9.4 31.2 CIMA 6 Unk 66.5 6.7 21.1 11

5 115

116

Table 18 (continued).

DTCC 2 Unk 57.4 39.4 29.6 21.9 17.4 10.3 6.6 5.7 6.8 21.3 24.1 21.8 DTCC 20 Unk 43.9 27.3 21.1 17.1 12.8 9.1 4.5 4.7 5.1 15.6 15.1 16.6 DTCC 21 Unk 82.9 51.8 39.8 30.3 25.7 15 9.4 9 12 30.5 32.8 28.6 DTCC 3 Unk 41.3 25.9 20 14.5 11.6 8 4.3 4 5.7 13.9 15.3 16.8 DTCC 4 Unk 84 25.5 6.8 23.5 DTCC 5 Unk 57.1 36 26.5 20.2 16.5 8.9 6.3 6.5 5.9 18.6 20.5 19.5 IVLP 1 Unk 1.8 6.2 IVLP 2 Unk 47.5 22 7.5 24.3 IVLP 4 Unk 43.3 19 5.8 21.5 IVLP 9 Unk 77.5 7.8 20 LC01 Unk 60.2 36.4 29.5 20.5 17.9 10.2 6.7 5.7 8.3 19.6 22.3 23.8

116

117

Table 19. Analysis of variance (ANOVA) in proximal, midshaft and distal bone width between adult male and female Gopherus agassizii from the East Mojave Desert, CA and NV.

Bone measurement F P df

Proximal Width: Humerus 38.951 < 0.001 1,10 Ilium 9.383 0.005 1,25 Scapula 42.434 < 0.001 1,9 Midshaft Width: Humerus 22.392 0.001 1,10 Ilium 4.767 0.039 1,25 Scapula 0.620 0.451 1.9 Distal Width: Humerus 20.327 0.001 1,9 Ilium 4.699 0.040 1,25 Scapula 9.853 0.012 1.9

118

Table 20. Regression statistics of carapace length (CL) versus resorption core diameter (RCD) in juvenile and adult Gopherus agassizii from the East Mojave Desert (CA and NV).

CL vs RCD r2 F P

Humerus: Juveniles 0.66 7.860 0.049 Females 0.18 1.100 0.342 Males 0.49 2.854 0.190 Ilium: Juveniles 0.92 99.615 < 0.001 Females 0.31 4.066 0.075 Males 0.35 5.856 0.034 Scapula: Juveniles 0.002 0.003 0.959 Females 0.002 0.009 0.931 Males 0.44 1.559 0.338

119

Table 21. Skeletochronological age estimates for Gopherus agassizii from the East Mojave Desert (CA and NV) using the Correction Factor (CF) and Ranking Protocol (RP) methods; CL = carapace length; J = juvenile, F = female, M = male; Unk = unknown sex.

Tortoise sex CL humerus ilium scapula Femur ID (mm) Age Age Age Age Age Age Age Age (RP) (CF) (RP) (CF) (RP) (CF) (RP) (CF) IV09 J 0 1 IVLP 16 J 42 0 1 0 1 IVLP 17 J 42.2 0 1 0 1 IVLP 14 J 44.4 0 1 0 1 IVLP 15 J 51.2 0 1 0 1 DTCC 18 J 62.6 0 1 4 1 IVLP 12 J 65.2 0 1 DTCC 19 J 65.6 1 2 2 3 DTCC 8 J 7 8 DTCC 1 J 9 7 9 8 DTCC 16 J 11 10 5 4 DTCC 7 J 133 12 12 10 12 8 10 DTCC 10 J 14 14 12 15 11 13 AK1 F 178 19 20 LC04 F 182 26 24 21 17 19 10 R-11 F 205 23 21 21 15 IVLP 7 F 211 17 18 20 18 R-03 F 228 21 22 R-76 F 238 30 26 28 23

120

Table 21 (continued).

ABFX F 240 29 21 23 20 17 15 DTCC 11 F 251 31 24 28 24 21 19 CIMA 5 F 39 29 DTCC 13 F 259 31 26 31 25 22 17 R-07 F 35 29 35 29 30 24 IV02 F 32 25 DTCC 15 F 26 22 15 11 21 11 LC01 F 29 22 21 14 20 12 DTCC 17 M 220 42 28 31 26 LC03 M 223 23 22 DTCC 6 M 226 32 21 24 17 25 18 IVA3 M 232 22 24 IVPH 2 M 242 23 21 IVLP 10 M 243 34 33 IV05 M 244 46 33 24 21 25 21 CIMA 1 M 244 26 23 IVPH 1 M 257 37 26 DTCC 12 M 270 59 31 42 26 36 18 IVPH 4 M 270 39 30 IV08 M 285 30 26 LC02 M 285 45 32 40 22 DTCC 3 unk 23 24 15 16 14 15 DTCC 20 unk 15 10 11 11 11 9 DTCC 5 unk 26 17 12 13 16 13

121

Table 21 (continued).

DTCC 2 unk 29 14 26 28 22 17 A-01 unk 42 32 28 17 IVLP 9 unk 36 24 DTCC 4 unk 44 32 DTCC 21 unk 43 27 50 25 39 31

122

Table 22 Maximum skeletochronological age estimates for Gopherus agassizii from the East Mojave Desert (CA and NV) determined by the Correction Factor (CF) and Ranking Protocol (RP) methods; CL = carapace length (mm); unk = unknown sex.

Tortoise sex Locality CL PL RP age CF age ID (mm) (mm) IVLP 16 juvenile *Lower plot 42 41.6 0 1 IVLP 17 juvenile *Lower plot 42.2 40.7 0 1 IVLP 14 juvenile *Lower plot 44.4 45.5 0 1 IV09 juvenile *Lower plot 47 48.4 0 1 IVLP 15 juvenile *Lower plot 51.2 50.4 0 1 DTCC 18 juvenile DTCC 62.6 57.4 4 1 IVLP 12 juvenile *Lower plot 65.2 62.5 0 1 DTCC 19 juvenile DTCC 65.6 65.2 2 3 DTCC 8 juvenile DTCC 123 117 7 8 DTCC 1 juvenile DTCC 9 8 DTCC 16 juvenile DTCC 11 10 DTCC 7 juvenile DTCC 133 125 12 12 DTCC 10 juvenile DTCC 144 137 14 15 AK1 female *Lower plot 178 168 19 20 LC04 female DTCC 182 26 24 R-11 female *Lower plot 205 189 23 21 IVLP 7 female *Lower plot 211 202 20 18 DTCC 17 male DTCC 220 42 28 LC03 male DTCC 223 202 23 22 DTCC 6 male DTCC 226 213 32 21 R-03 female *Lower plot 228 207 21 22

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Table 22 (continued).

IVA3 male *Lower plot 232 224 22 24 R-76 female *Upper plot 238 217 30 26 ABFX female *Lower plot 240 215 29 20 IVPH 2 male *Lower plot 242 233 23 21 IVLP 10 male *Lower plot 243 227 34 33 CIMA 1 male *Upper plot 244 226 26 23 IV05 male *Lower plot 244 225 46 33 DTCC 11 female DTCC 251 231 31 24 IVPH 1 male *Lower plot 257 37 26 CIMA 5 female *Upper plot 258 197 39 29 DTCC 13 female DTCC 259 228 31 26 IVPH 4 male *Lower plot 270 248 39 30 DTCC 12 male DTCC 270 230 59 31 IV08 male *Lower plot 285 261 30 26 LC02 male DTCC 285 45 32 DTCC 9 female(pet) DTCC 325 300 39 13 DTCC 14 female(pet) DTCC 310 292 36 25 DTCC 3 unk DTCC 23 24 DTCC 15 female DTCC 205 26 22 DTCC 5 unk DTCC 26 17 LC01 female DTCC 29 22 DTCC 2 unk DTCC 29 28 IV02 female *Lower plot 196 32 25 R-07 female *Lower plot 190 35 29

124

Table 22 (continued).

IVLP 9 unk *Lower plot 36 24 A-01 unk *Lower plot 42 32 DTCC 4 unk DTCC 44 32 DTCC 21 unk DTCC 50 27

* Ivanpah, Mojave National Preserve

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Table 23. Regression statistics for age versus carapace length (CL) in juvenile and adult Gopherus agassizii from the East Mojave Desert (CA and NV); CL = carapace length. Values in bold are statistically significant.

Age vs CL r2 F P

Juveniles 0.77 37.074 > 0.001 Females 0.56 10.353 0.012 Males 0.18 2.486 0.143

126

350 (DTCC) CL vs CW y = 0.708x + 13.303 (r ² = 0.99) (DTCC) CL vs PL y = 0.878x + 7.823 (r ² = 0.99) 300 (MNP) CL vs CWy = 0.768x + 4.951 (r ² = 0.99) (MNP) CL vs PLy = 0.902x + 4.372 (r ² = 0.99) 250

200

150

100 Shell measurement (mm) 50

0 0 50 100 150 200 250 300 350

Carapace length (mm)

Figure 19. Regression of carapace length (CL) vs carapace width (CW) and plastron length (PL) for Gopherus agassizii from the Desert Tortoise Conservation Center (DTCC; solid lines), Nevada (NV) and the Mojave National Preserve (MNP; dashed lines), California (CA).

127

300 CL vs CW (female)y = 0.757x + 3.981 (r ² = 0.79) 280 CL vs PL (female)y = 0.623x + 62.752 (r ² = 0.76) CL vs CW (male)y = 0.546x + 59.137 (r ² = 0.67) 260 CL vs PL (male)y = 0.669x + 64.515 (r ² = 0.79) 240 220

200 180

Shell measurement (mm) 160 140 120 160 180 200 220 240 260 280 300 Carapace length (mm)

Figure 20. Carapace length (CL) vs carapace width (CW) and plastron length (PL) in adult male (dashed lines) and female (solid lines) Gopherus agassizii from the East Mojave Desert (CA and NV).

300

250

200

150

Shell size (mm) 100

F M F M F M 0

Carapace length Carapace width Plastron length

Figure 21. Shell size of A) adult female and B) male Gopherus agassizii from the East Mojave Desert (CA and NV). Males are significantly larger than females for all shell measurements (p < 0.05). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers. 129

100

40 Bone Length (mm) 20

F M F M F M 0

Humerus Ilium Scapula

Figure 22. Bone length for adult male and female Gopherus agassizii from the East Mojave Desert (CA and NV). In all cases males had significantly longer bones than females (p < 0.05). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers.

130

100 humerus y = 0.291x - 3.334 (r ² = 0.88) iliumy = 0.178x - 0.203 (r ² = 0.97) femur y = 0.207x + 0.370 (r ² = 1.00) 80 scapula y = 0.136x - 0.376 (r ² = 0.95)

40 Bone Length (mm) 20

0 0 50 100 150 200 250 300 350 Carapace Length (mm)

Figure 23. Regression of carapace length versus bone length in Gopherus agassizii from the East Mojave Desert (CA and NV).

131

30 25

20 15 10

Proximal Width (mm) 5 100

Midshaft Width (mm) 2

F M F M F M 400 Humeri Ilia Scapulae

Figure 24. Bone widths of adult male and female Gopherus agassizii from the East Mojave Desert (NV and CA). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers.

132

400

Distal Width (mm) 10

F M F M F M 0 Humeri Ilia Scapulae

Figure 24 (continued).

133

Resorption Core Diameter (um) 1 F M F M F M 0

Humerus Ilium Scapula

Figure 25. Resorption core diameters for adult Gopherus agassizii from the East Mojave Desert (CA and NV). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers.

134

7 juveniles y = 0.004x + 0.733 (r ² = 0.66) 6 females y = -0.011x + 5.214 (r ² = 0.18) males y = 0.029x - 3.104 (r ² = 0.49) 5

3 Humerus

1 resorption core diameter (um)

05 juveniles y = -0.013x + 0.016 (r ² = 0.91) females y = 0.012x - 0.373 (r ² = 0.31) 4 males y = 0.021x - 2.353 (r ² = 0.35)

Ilium 2

1 resorption core diameter (um)

50 0 50 100 150 200 250 300

Figure 26. Regression of carapace length versus resorption core diameter for Gopherus agassizii from the East Mojave Desert (CA and NV).

135

50 juveniles y = 0.0002x + 0.854 (r ² = 0.002) females y = -0.0006x + 2.164 (r ² = 0.002) 4 males y = 0.016x - 0.880 (r ² = 0.44)

Scapula 2

1 resorption core diameter (um)

0 0 50 100 150 200 250 300 Carapace length (mm)

Figure 26 (continued).

136

males

females

0 10 20 30 40 50 60 70

Age (years)

Figure 27. Age estimates adult male and female Gopherus agassizii from the East Mojave Desert (CA and NV). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers.

137

300

250

200

150

100 Carapace length (mm) 50

y = -0.102x2 + 9.907x + 30.891 (r ² = 0.91) 0 0 10 20 30 40 50 60 70 Age (years)

Figure 28. Regression of age versus body size (carapace length) in Gopherus agassizii from the East Mojave Desert (CA and NV).

138

320 females y = -0.053x + 6.621x2 + 86.360 (r ² = 0.57) 300 males y = -0.011x + 1.698x2 + 203.945 (r ² = 0.19)

280

260

240

220

Carapace Length (mm) 200

180

160 10 20 30 40 50 60 70

Age (years)

Figure 29. Regression of age versus size in adult male and female Gopherus agassizii from the East Mojave Desert (CA and NV).

139

251-300

201-250

151-200

102-150

Size Classes (mm) 51-100

0-50

0 10 20 30 40 50 60 70 Age (years)

Figure 30. Age range within the size classes of Gopherus agassizii from the East Mojave Desert (CA and NV). Horizontal lines within each box represent the median age and whiskers represent the 10th and 90th percentile. Dots represent outliers.

140

350 females y = -0.17x + 11.40x2 + 32.17 (r ² = 0.89) 2 300 males y = -0.16x + 11.59x + 29.98 (r ² = 0.96)

250

200

150

100 Plastron Length (mm)

0 0 10 20 30 40 50 60 Age (years)

Figure 31. Regression of size (plastron length) versus age for East Mojave Desert tortoises (Gopherus agassizii) from Rock Valley, Nevada (data from Table 1; Turner et al., 1987), Clark County, Nevada and Ivanpah Valley, Mojave National Preserve, California.

141

1997 1998 1999 2000 2001 2003

Figure 32. Average annual rainfall for the lower and higher elevation study plots (800 – 1100 m) in Ivanpah Valley, Mojave National Preserve, California (Avery, unpubl. data) between 1997 and 2003.

142

CHAPTER 5. GROWTH STRATEGIES IN DESERT TORTOISE POPULATIONS: ECOLOGICAL IMPLICATIONS AND FUTURE TRENDS.

Introduction

The desert tortoise, Gopherus agassizii, inhabits the broadest range of habitats and latitudes of the four species of North American tortoises (Berry et al., 2002).

Considerable variation in ecology (Luckenbach, 1982, Averill-Murray et al., 2002b), behavior (Zimmerman et al., 1994; Averill-Murray et al., 2002a), morphology (Germano,

1993; Berry et al., 2002), and DNA (Lamb et al., 1989; Glenn et al., 1990; Lamb and

McLuckie, 2002) have been noted in different populations of desert tortoises and three genetically distinctive populations have since been defined in the Mojave Desert,

Sonoran Desert and tropical Sonora and Sinaloa (Lamb et al., 1989; Lamb and McLuckie,

2002).

The fossil record suggests that G. agassizii evolved in a more mesic climate and the subsequent formation of the Sonoran and Mojave deserts during Miocene to

Pleistocene glacial climates left tortoises in an increasingly dry and unpredictable environment (Van Devender, 2002). Mean winter rainfall values broadly overlap between Mojave and Sonoran deserts, but summer rainfall decreases from the Sonoran

Desert through the East Mojave to the extremely dry West Mojave Desert (Turner and

Brown, 1982; Henen et al., 1998; Wallis et al., 1999; Averill-Murray, 2002). If we assume, as Van Devender (2002) suggested, that Sonoran tortoises are most similar to the ancestral G. agassizii stock because they appear more adapted to a mesic environment

(Van Devender, 2002), we can form hypotheses for the evolution of derived life-history traits in Mojave tortoises.

143

If Sonoran tortoises are relics of tropical deciduous forest-evolved populations

(Lowe, 1990), then they should be viewed as adapted to thornscrub, not desert – an observation that has been reinforced behaviorally (Averill-Murray et al., 2002a) and physiologically (at least in Mojave Tortoises; Peterson, 1996). Thornscrub tortoises from the more tropical Sonora and Sinaloa in Mexico do not inhabit valley bottoms due to heavy flooding during summer rains and also live in a largely frost-free environment, having no need to construct extensive burrows (Van Devender, 2002, McCord, 2002).

Sonoran Desert tortoises are therefore hypothesized to continue a thornscrub lifestyle because they can (McCord, 2002). During the formation of the Mojave Desert in the

Pleistocene (2.4 m.y.a.), there was a shift to a uniseasonal winter rainfall pattern and consequent evolution of diverse spring annual flora from temperate herbaceous perennial ancestors (Van Devender, 2002). During this time, most animal species simply retracted their ranges to the southeast (where tortoises with characteristics derived from thornscrub ancestors still exist), but those tortoises which persisted through the formation of the

Mojave Desert, were hypothesized to have adopted and established the juvenile traits of their tropical ancestors, thereby evolving into the modern day Mojave tortoise.

According to Van Devender (2002), in Mojave Desert tortoises, the activity period shifted to spring and aestivation began in early summer and extended through the warm season to merge with fall-winter hibernation. The excavation of extensive burrows in valleys and washes (areas not typically inhabited by Sonoran or Sinaloan tortoises) allowed Mojave tortoises to survive the extended inactivity periods, including colder winter temperatures than tortoises experience in the Sonoran Desert. The diet shifted to primarily fresh spring annuals with fewer dry grasses and mallows, and survivorship of

144 juveniles was enhanced by laying eggs in several clutches in May, late June and August, although the number of eggs per year was essentially the same as in the Sonoran tortoises

(Van Devender, 2002). Van Devender, therefore suggested that the more thermally stressed, relatively flood-free Mojave tortoise adopted and evolved a more juvenile- derived ecology in response to the evolution of the Mojave Desert. Morafka and Berry

(2002), however, claim that the desert tortoise achieved much of its morphological distinctness 12 million years prior to the formation of North American desert landscapes.

Based on their estimated origins of shared primitive and shared derived body features and behavior, they suggested that most desert tortoise differentiation and functional adaptations preceded the appearance of all North American deserts and occurred instead in response to lowland microhabitats or edaphic patches with sandy or friable soils.

Morafka and Berry (2002) therefore, suggest that G. agassizii is an opportunistic generalist rather than a desert specialist, which survives in desert environments by demonstrating morphological, physiological and behavioral plasticity allowing it to shuttle across temporal (annual) and spatial mosaics to achieve net stability.

I wanted to investigate these hypotheses by comparing growth strategies across the xeric extremes of the desert tortoise’s range and assess the potential effects of increased desertification on tortoise habitat. The Sonoran Desert is considered to be the most tropical of the four North American deserts, both because of its climate, with milder virtually frost-free winters and summer monsoonal rainfall from the tropical oceans, and its physical connection with more tropical communities to the south in Baja California and Sonora, Mexico (Van Devender, 2002). Summer temperatures are mostly hot throughout the range of G. agassizii, but winter temperatures are mild in the south and

145 become increasingly colder toward the north, where subfreezing temperatures are known to occur in the Mojave Desert (Germano et al., 1994). Mojave tortoises prefer flat, open land with plant associations of creosote (Larrea tridentata) and white bursage (Ambrosia dumosa), whereas Sonoran tortoises mainly inhabit rocky slopes and dry washes in hilly or mountainous regions populated by paloverde, mixed cacti and scrub (Averill-Murray et al., 2002b, McLuckie et al., 1999).

Studies on growth rates in wild desert tortoises have been evaluated using age, annual body size measurements, scute laminae, and body weight as parameters

(Woodbury and Hardy, 1948; Medica et al., 1975; Turner et al., 1987; Germano, 1988;

1994; Germano et al., 2002); however, these studies have been conducted mainly on desert tortoises from the northeastern Mojave Desert. Furthermore, growth variation across populations is limited and difficult to compare. Mojave tortoises are hypothesized to mature at earlier ages than Sonoran tortoises as a way of balancing the higher juvenile mortality in these populations compared to Sonoran tortoises (Iverson, 1992; Mueller et al., 1998; Averill-Murray et al., 2002). Adult survivorship is hypothesized to be similar between the two deserts (Germano, 1994; U.S. Fish and Wildlife Service, 1994; Averill-

Murray et al., 2002). The goal of my study was to evaluate growth rates from different desert tortoise populations using uniform methodology. These growth rates, along with various life-history traits, could then be used to compare and construct a more accurate picture of G. agassizii growth and future survival across the desert portion of their range.

146

Materials and methods

The Sonoran Desert sample of Gopherus agassizii came from various localities throughout Arizona (see Chapter 3) and consisted of 52 carcasses: 20 juveniles, 19 adult females, and 13 adult males. The West Mojave Desert sample from San Bernardino

County, California (see Chapter three), consisted of 48 carcasses: 19 juveniles, 9 adult females, and 20 adult males. The East Mojave Desert sample from The Mojave National

Preserve (California) and Clark and Nye Counties, Nevada, consisted of 59 carcasses: 29 juveniles, 15 adult females, and 15 adult males.

I measured carapace length, carapace width and plastron length for all intact shells. I fixed, decalcified and processed the long bones collected from carcasses according to standard histological protocol. All bones were embedded in Paraplast

Plus©. I cut 20 µm cross sections were through the midshaft area of each bone using a rotary microtome and stained sections with hematoxylin and eosin. All sectioned were observed under a transmitting light microscope. See Parham and Zug (1997) and Zug

(1990) for the skeletochronology protocols used to determine age estimates in this study.

I compared size and age estimates between the populations by analysis of variance (ANOVA). I estimated annual growth rate in each tortoise according to the

Medica et al. method (1975) whereby the total growth measured (carapace or plastron length at death) was divided by the estimated number of years over which this growth had occurred. This enabled me to compare annual growth rates between the three desert populations of Gopherus agassizii and also to previously published values. In estimating individual growth rates, I did not include juveniles less than 2 years old because annual growth was highly influenced by the size at hatching and produced grossly overestimated

147 growth rates. Standard errors represent one standard error of the mean. Statistical analyses were conducted with the SYSTAT (Version 9.0) statistical package and I accepted statistical significance at P < 0.05.

Results

West Mojave females had the smallest mean shell sizes and Sonoran females the largest of the three populations, but in general females were similar in size (Table 1, Fig.

1A). Analysis of variance with Tukey post-hoc tests, however, showed no statistically significant difference in shell size between females from the three populations (CL: F2,53

2 2 = 2.724, P = 0.076; r = 10; CW: F2,45 = 0.137, P = 0.872, r = 0.006; PL: F2,48 = 3.012, P

= 0.056, r2 = 0.11). In contrast, when I conducted an ANCOVA test with carapace length

(CL) as the covariate, I found a statistically significant difference in carapace width (CW) between females of the three populations (F = 3.374, P = 0.045, r2 = 0.77), with Mojave females being wider than Sonoran females. Adjusted least squares means were 173.7 +

2.4 SE for East Mojave females, 169.2 + 2.1 SE for Sonoran females and 179.4 + 3.4 SE for West Mojave females at the predicted CL of 224.3 mm. The ANCOVA results for PL with CL as the covariate showed no statistically significant difference between females from the three populations (F = 1.135, P = 0.332, r2 = 0.77).

Males were generally similar in size (Table 1, Fig. 1B), although there was a statistically significant difference in CW between the three populations (F2,44 = 4.049, P =

0.023, r2 = 0.16). A Tukey post-hoc test showed that Sonoran males were significantly narrower than West Mojave males (P = 0.017).

148

In age/size regressions, Sonoran and East Mojave tortoises had a similar growth pattern

(Fig. 2). West Mojave tortoises, on the other hand, displayed the fastest growth of the three populations and seemed to reach similar sizes at earlier ages than either the East

Mojave or Sonoran populations (Fig. 2). I compared juvenile age/size regressions by analysis of covariance with age as the covariate and found no statistically significant difference between juveniles from the three populations (F = 0.234, P = 0.792, r2 = 0.69;

Fig. 3). Adjusted least squares means were 93.74 + 5.64 SE for West Mojave juveniles,

91.08 + 5.11 SE for East Mojave juveniles and 96.28 + 5.49 SE for Sonoran juveniles at the predicted age of 8.68 years.

Analysis of variance showed a statistically significant difference in annual growth

2 rates between juveniles of all three populations (F2,53 = 4.502, P = 0.016, r = 0.15; Fig.

4). A Tukey post-hoc test showed that East Mojave juveniles grew significantly faster than both West Mojave (P = 0.044) and Sonoran juveniles (P = 0.029). Sonoran juveniles had a mean annual growth rate of 9.67 + 0.58 mm/yr (range: 14.90 - 6.20 mm/yr). West Mojave juveniles had a mean of 9.87 + 0.71 mm/yr (range: 13.58 – 3.13 mm/yr) and East Mojave juveniles a mean of 11.94 + 0.49 mm/yr (range: 15.38 – 8.71 mm/yr).

In chapters three and four, I reported no sexual dimorphism in body size between

Sonoran males and females but did find a significant sexual dimorphism in Mojave adults. Mojave males reach significantly larger sizes that Mojave females, in terms of carapace length and plastron length (only East Mojave males). There was no sexual dimorphism in annual growth rates between males and females from the three populations (Table 1, Fig. 4).

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Within females, analysis of covariance of size with age as the covariate showed no statistically significant difference in growth between the three populations (F = 0.616,

P = 0.545, r2 = 0.23). Adjusted least squares means were 231.9 + 8.7 SE for West

Mojave females, 220.7 + 5.4 SE for East Mojave females and 224.8 + 5.4 SE for Sonoran females at the predicted mean age of 29.7 years. West Mojave females had the shortest life-spans of the three populations, whereas Sonoran and East Mojave females showed similar longer life-spans (Table 1, Fig. 5A). West Mojave females had the highest annual growth rates at 11.17 + 0.87 mm/yr (range: 16.00 – 7.62 mm/yr), whereas Sonoran females had the lowest at 6.90 + 0.22 mm/yr (range: 9.04 – 5.19 mm/yr). East Mojave females had intermediate annual growth rates at 7.91 + 0.53 mm/yr (range: 12.60 – 5.35 mm/yr). Analysis of variance showed a statistically significant difference in annual growth between females (F2,41 = 16.703, P < 0.001; Fig. 4). A Tukey post-hoc test showed that West Mojave females had a significantly higher annual growth rate than both

East Mojave and Sonoran females (P < 0.001).

In males, all three populations reached similar sizes and, although age/size regressions seemed to show faster growth in West Mojave males (Fig. 5B), analysis of covariance removing the effect of age showed no statistically significant difference in growth between the three populations (F = 0.753, P = 0.477, r2 = 0.17). Adjusted least squares means were 251.7 + 7.3 SE for West Mojave males, 242.8 + 7.5 SE for East

Mojave, and 237.1 + 8.4 SE for Sonoran males at the predicted age of 31.8 years. West

Mojave males had the youngest mean age of the three populations even though having a similar longevity with East Mojave and Sonoran males (Table 1). West Mojave males had the highest mean annual growth at 10.83 + 0.65 mm/yr (range: 16.18 – 4.68 mm/yr),

150 whereas Sonoran males had the lowest at 6.63 + 0.27 mm/yr (range: 7.82 – 5.13 mm/yr).

East Mojave males were intermediate at 7.30 + 0.50 mm/yr (range: 10.55 – 4.58 mm/yr).

As in females, analysis of variance showed a statistically significant difference in annual

2 growth between males from the three populations (F2,46 = 17.319, P < 0.001, r = 0.43).

A Tukey post-hoc test showed that West Mojave males had significantly higher annual growth rates than both Sonoran and West Mojave males (P < 0.001). There was no significant difference in annual growth between Sonoran and East Mojave males.

In terms of age at sexual maturity, Sonoran females had the latest maturity at 22 to 27 years, and West Mojave females had earliest maturity, 15 to 19 years. East Mojave females were intermediate with maturity estimates of 19 to 25 years. All age estimates assumed sexual maturity at ~180 mm CL.

Discussion

Size, growth and productivity

Annual growth rates of East Mojave tortoises in this study were similar to those determined from previous studies. Turner et al. (1987) and Germano (1988) determined growth rates in wild, known-aged northeastern Mojave Desert tortoises. Mean annual growth for these tortoises over a 17 year period was 7.6 mm + 0.83 SE (Turner et al.,

1987). This growth rate does not differ significantly from their previous estimate of 9 mm/yr (range was 2 to 12 mm/yr) for the same cohort (Medica et al., 1975). Germano

(1988, 1994) also estimated growth for the same cohort using scute ring counts and diameters as the age/size variables; he reported growth rates of 10.9 mm/yr (scute

151 laminae counts) and 7.6 mm/yr (GL diameters). Medica et al. (1975) observed that growth occurred mainly in the spring and summer, generally between April and July.

Murray and Klug (1996), conducted growth analyses of three desert tortoise populations in Arizona. They obtained similar estimates to those of Germano et al.

(2002). Germano used CL and the Richard’s growth model to construct growth curves for Sonoran Desert tortoises. The Richards model predicted that in the Sonoran Desert, both sexes would attain a maximum CL of about 270 mm (Germano et al., 2002), which was similar to the largest mean size of desert tortoises from the West Mojave Desert

(Germano, 1994). This maximum size for Sonoran females was confirmed by the present study. The largest females were between 265 to 270 mm CL, however my data indicated that Sonoran males reached larger sizes (285 to 290 mm CL).

Data from this study suggest that growth rates are substantially lower than those estimated by the Richard’s growth model. Germano et al. (2002) predicted that both sexes would require almost 7 years to reach 100 mm CL, which equated to an annual growth rate of 14.29 mm/yr. Females would require nearly 17 years (11.77 mm/yr) and males almost 18 years (11.11 mm/yr) to reach 200 mm (Germano et al., 2002). In this study, juveniles between 88 and 107 mm CL were estimated as 7 to 11 years old (~ 10.89 mm/yr), one juvenile of 133 mm CL was estimated as 17 years old (7.82 mm/yr) and a subadult of 175 mm CL was estimated as 26 years old (6.73 mm/yr). The smallest adult female (180 mm CL) was estimated to be 27 years old (6.67 mm/yr), and the smallest adult male (195 mm CL) 25 years old (7.80 mm/yr). All adults above 200 mm CL were above 30 years in age (Chapter three, Table 8).

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Germano et al. (2002) determined minimum age at sexual maturity using a published value of 190 mm CL because a minimum size of maturity was unknown for

Sonoran tortoises. They determined age at maturity from scute annuli for those females >

190 mm CL. Age at maturity was estimated to be 15.7 years by using the mean of measurements of scute annuli for each individual. At 190 mm CL, the Richards’ growth equation yielded 15.3 years (Germano et al., 2002). I set minimum size at maturity as

180 mm CL (Averill-Murray and Averill-Murray, 2005), yielding an age estimate of 22 to 27 years old in Sonoran females, with a mean of 24.5 years, considerably older than the estimates of Germano et al. (2002).

Adult females from all three populations were generally similar in size although

West Mojave females had the smallest mean shell sizes of the three populations (Table

1). Maternal body size strongly influences reproductive output (Wallis et al., 1999).

Turner et al. (1986) reported that female body size affected clutch frequency and clutch size, whereas Mueller et al. (1998) found that female size affected clutch size and annual egg production, but not clutch frequency. Wallis et al. (1999) found that body size affected annual egg production primarily through effects on clutch frequency and the size of the first clutch, egg size and the volume of egg clutches. Their results were consistent with the use of both body reserves (water and protein; Henen, 1997) and food intake to support reproductive allocations, suggesting that large females may be able to develop more follicles and store more nutrients than smaller females (Wallis et al., 1999).

Annual rainfall decreases significantly and becomes more unpredictable from east to west in the desert tortoise’s range (Averill-Murray et al., 2002a). The sexual size dimorphism in Mojave tortoises but not Sonoran tortoises, and the smaller sizes of West

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Mojave females, could be a response to resource constraints imposed by varying rainfall patterns. In the bi-seasonal Sonoran Desert and more tropical areas to the south, tortoises are active primarily during the summer rainy season (annual rainfall between 140 to >

300 mm/yr) and the majority of the annual dietary intake occurs from July to September

(Van Devender et al., 2002). In contrast, in the winter-rainfall climate of the Mojave

Desert (annual rainfall in the West Mojave being 80 - 170 mm/yr, and in the East Mojave between 75 – 200 mm/yr), tortoises are primarily active during the spring, passing the remainder of the year in burrows (Morafka and Berry, 2002). The smaller West Mojave females may have smaller body nutrient reserves and a larger portion of the nutrients for their eggs may need to come from winter annuals consumed after emergence from hibernation (Wallis et al., 1999). Winter annuals, however, senesce quickly and desiccate by June (the peak laying period in the Mojave Desert) and females emerge from hibernation in March or April, so there may be little time for females to acquire nutrients prior to ovulation, which peaks in May (Rostal et al., 1994).

Henen (1997) suggested that tortoises drank rainwater and obtained water from forage in the spring to achieve positive energy balance (Henen, 1997), but other studies have shown that Mojave tortoises are unable to obtain enough energy to balance expenditures (Nagy and Medica, 1986; Peterson, 1996). Nagy and Medica (1986) proposed that negative energy balance, even while feeding during spring, could be due to high water content in the forage outweighing dry matter (energy) intake. Tortoises might also consume food at less than their maximum rate because of harmful components in the diet, such as excess potassium, which tortoises cannot excrete without losing water (Nagy and Medica, 1986; Peterson, 1996). Drought conditions generally tend to prevail during

154 the dry Mojave summers, and tortoises endure negative water balance and generally cease feeding, spending more time inactive in their burrows, where excess dietary salts and metabolic wastes become concentrated in their large bladders (Nagy and Medica,

1986; Peterson, 1996). The arrival of summer rainfall is important because drinking rainwater allows tortoises to flush their systems of salts and rehydrate their bladders and they can then obtain energy for growth and maintenance from still-dry forage and fresh vegetation as it becomes available (Nagy and Medica, 1996; Peterson, 1996; Nagy et al.,

1997). Summer rainfall, however, is nearly nonexistent in the West Mojave Desert (< 10

% of annual rainfall). Summer storms are more common, but erratic in the East Mojave and western Sonoran Deserts, and comprise half or more of the annual precipitation in the eastern Sonoran Desert (Turner and Brown, 1982; Averill-Murray et al., 2002a, Van

Devender et al., 2002). The growth and reproductive output of Mojave tortoises, especially West Mojave females, will be more seriously affected by increased temperature and droughts during the growing season, as opposed to those tortoises living in the Sonoran Desert with more consistent and predictable rainfall.

If tortoise growth is mostly impacted by environmental constraints then I expected Sonoran tortoises to display the highest growth rates of the three desert populations as the Sonoran Desert appears to be the most climatically favorable of the three habitats (at least in terms of annual rainfall). This hypothesis was not supported by the data. Sonoran tortoises showed the lowest growth rates of the three populations whereas West Mojave tortoises showed the highest (Table 1). In his study on Mojave tortoise growth, Germano (1994) also noted that West Mojave tortoises grew faster than

East Mojave tortoises. The explanation for the results of this study and those of Germano

155

(1994) may lie in differences in desert productivity. The West Mojave Desert, even though it receives less annual rainfall than the East Mojave or Sonoran Desert, seems to produce a greater biomass when it does rain than the other two deserts. For example,

Brooks (1995) studied productivity in the West Mojave Desert over a three year period

(1990 to 1992), and found that during a low rainfall year (1989/1990), biomass was between 4.74 – 12.33 kg/h, whereas the following year (1990/1991), after a season of high rainfall, biomass increased to an astounding 57.77 – 199.46 kg/ha. This highly productive year was followed by a year with intermediate rainfall and still substantially high biomass (39.61 – 94.92 kg/ha; Brooks, 1995). In contrast, Avery (1998) recorded much lower biomass values in the East Mojave Desert of between 2.82 – 8.25 kg/ha

(spring of 1992) and 1.18 – 3.33 kg/ha (spring of 1993). Desert tortoises certainly seem to display phenotypic plasticity in growth, with the capacity to increase growth rates in the presence of consistent and abundant food and water (Patterson and Brattstrom, 1972;

Jackson et al., 1976; Jackson et al., 1978; Turner et al., 1987).

Ecological implications and conclusions

West Mojave tortoises had the smallest females, highest growth rates, youngest age at sexual maturity but shortest longevity of the three populations. In contrast,

Sonoran tortoises had the largest females, lowest growth rates, oldest age at sexual maturity and greatest longevity, whereas East Mojave tortoises showed intermediate traits between the West Mojave and Sonoran populations (Table. 1). Sonoran tortoises invest their entire reproductive output in a single clutch during the relatively predictable summer rainy season and typically produced few eggs overall than in the Mojave Desert, except under extreme drought conditions (Averill-Murray, 2002). These differences may

156 be an evolutionary product of greater hatchling survival in the Sonoran Desert than in the

Mojave Desert. Averill-Murray (2002) speculates that drier summer conditions in the

Mojave Desert, especially in the West Mojave Desert (Peterson, 1996; Henen et al.,

1998; Wallis et al., 1999) may have resulted in the tortoises adaptively producing a second and sometimes third clutch, thus maximizing the chance that at least some hatchlings will emerge in favorable conditions. Hatchlings emerging in the Sonoran

Desert have a relatively predictable supply of forage from which they can supplement their nutrient reserves to survive their first winter. The earlier maturation and production of offspring in Mojave tortoises are suggested to have developed as a way of balancing the higher juvenile mortality in these populations compared to Sonoran tortoises (Iverson,

1992; Mueller et al., 1998; Averill-Murray et al., 2002).

Annual growth rates in West Mojave adults were similar to those of juveniles from all three populations, whereas East Mojave, and especially Sonoran adults, had significantly reduced growth rates after the attainment of sexual maturity (Fig.5). West

Mojave tortoises may be able to sustain higher overall growth rates than their eastern counterparts by taking advantage of higher productivity and more abundant forage after wet years. This lends credence to the hypothesis proposed by Morafka and Berry (2002) that desert tortoises are exaptive opportunists able to shuttle across temporal and spatial nutritional mosaics, in modes which broadly parallel those of behavioral thermo regulators as they shuttle across a thermal mosaic to achieve physiological stability. In the tortoise, net stability is achieved over an annual rather than diel cycle of behaviors and physiological changes (Morafka and Berry, 2002).

157

Certainly the innate morphological and behavioral flexibility of the desert tortoise may be viewed as substantial exaptations with which it could withstand changes in climates, habitats and forage. But the effects of anthropogenic desertification on desert tortoise habitats continue to promote a reduction in seasonal, spatial, and nutritional accessibility and availability of tortoise food supplies (Morafka and Berry, 2002), especially in the thermally stressful, uniseasonal, and already limited, rainfall area of the

Mojave Desert. The consequences may be manifest not only by the direct effects of starvation, as my data seem to suggest, but more subtly, through the spread of epidemic diseases like upper respiratory tract disease (already abundant in West Mojave populations), especially in tortoises rendered immuno-compromised by malnutrition

(Morafka and Berry, 2002). The cost to sustained high annual growth for tortoises living in the West Mojave Desert, without dependable and predictable annual forage and water for health and growth maintenance, is a higher mortality and much reduced life expectancy which females have tried to compensate for by evolving a younger age at sexual maturity (Table 1).

All of the options critical to tortoise survival are being narrowed, in many cases simulating the consequences of natural climate desiccation. The rapid rates at which these deleterious processes move forward are historically unprecedented and the potential for continued reduction in quantity and quality of tortoise forage plants is high (Morafka and Berry, 2002).

Increased thermal stress may also explain the delayed maturity and low annual growth rates of Sonoran as opposed to Mojave tortoises. In some areas of Sonoran tortoise habitat, summer highs may exceed 48.5 °C, with surface temperatures

158 approaching 82 °C (Van Devender et al. 2002). Summer temperatures in the majority of

Sonoran tortoise habitat (which is also the main growing season for many of their food plants and when the most rain falls) regularly exceed 38 °C, with humidity often in single digits (Van Devender et al. 2002).

Peterson (1996) concluded from his studies in the West and East Mojave Desert of California that the desert tortoise is not physiologically adapted to live in the desert, but is a tenuous relic of a less rigorous climate. In the absence of severe habitat deterioration or human exploitation, most turtle species apparently have the potential for extremely high levels of annual survival among adults (Frazer et al., 1990).

Anthropogenic stressors can cause smaller sizes at maturity and can result in reduced allocation to growth by juveniles, decreased age at maturity, or both (Congdon et al.,

2001). The boom-bust cycle of high productivity but lowest annual rainfall in the West

Mojave Desert probably causes short-term droughts to have a more deleterious effect

(physiological stress and younger ages at death) on West Mojave tortoises because they rely almost solely on winter rains for producing their main food sources (Peterson, 1996;

Henen et al., 1998). But inadequate and increased loss of resources, whether by habitat destruction and/or droughts, and the promotion of chronic disease are all factors that have resulted directly or indirectly in a shorter longevity than their eastern and southern counterparts.

Desert tortoise populations are potential models for the consequences of increased desertification on desert fauna, especially in those species that have not evolved as desert specialists but rather have learned to adapt and persevere in these extreme environments.

If global warming continues, the rate of short and long term droughts may also increase,

159 along with latitudinal and longitudinal drying trends (Le Houerou, 1996). The consequences of habitat desertification are issues that many species will face in the future as drying trends shift and continue to expand (exacerbated by anthropogenic causes) and global temperatures continue to increase.

160

Table 24. Summary of life-history traits and annual growth rates for Gopherus agassizii from the West and East Mojave Desert in California and southern Nevada and from the Sonoran Desert in Arizona.

West Mojave East Mojave Sonoran Desert

Desert Desert

Mean body size (mm CL) Females 214.50 (+ 5.64) 222.33 (+ 5.00) 232.16 (+ 4.26) Males 244.69 (+ 8.09) 245.94 (+ 3.59) 243.43 (+ 7.56) Maximum body size (mm CL) Females 245 259 266 Males 290 285 287 Mean age (years) Females 20.73 (+ 1.34) 30.22 (+ 1.78) 34.44 (+ 2.63) Males 24.05 (+ 1.88) 36.44 (+ 2.63) 38.38 (+ 1.84) Maximum age at death (years) Females 27 42 43 Males 56 59 54 Age at maturity (years) 15-19 19-25 22-27 Mean annual growth (mm/yr) 10.67 (+ 0.46) 8.83 (+ 0.36) 7.88 (+ 0.32) Juveniles 9.99 (+ 0.98) 10.51 (+ 0.48) 9.85 (+ 0.61) Females 11.17 (+ 0.87) 7.89 (+ 0.56) 6.90 (+ 0.22) Males 10.83 (+ 0.65) 7.33 (+ 0.54) 6.63 (+ 0.27) Clutch frequency 1 – 2c 1 – 2a 0 - 1b Clutch size 2 – 9c 2 – 8a,e 2 – 12b Mean 3.0 – 7.1c,d 4.9 – 8.4a,c 3.3 – 5.7b Mean egg width (mm) 35.81 (+ 0.66)c 37.2 (+ 0.26)e 35.70 (+ 1.08)b

161

Table 24 (continued). aRostal et al., 1994 eMcLuckie and Fridell, 2002 bAverill-Murray, 2002 cWallis et al., 1999 dHenen, 1997

162

300 A 250

200

150

Shell size (mm) 100

3500 B 300

250

200

150 Shell size (mm) 100

50 SD WM EM SD WM EM SD WM EM 0

Carapace length Carapace width Plastron length

Figure 33. Carapace length (CL), carapace width (CW) and plastron length (PL) of A) adult female and B) adult male Gopherus agassizii from the West and East Mojave Desert (California and Nevada) and Sonoran Desert (SD; Arizona); WM = West Mojave, EM = East Mojave.

163

2 350 West Mojave y = -0.159x + 12.176 + 32.481 (r ² = 0.75) East Mojave y = -0.106x2 + 9.994 + 24.052 (r ² = 0.91) Sonoran Desert y = -0.109x2 + 9.931 + 25.224 (r ² = 0.92) 300

250

200

150

100 Carapace length (mm)

0 0 10 20 30 40 50 60 70 Age (years)

Figure 34. Age versus body size in Gopherus agassizii from the West Mojave Desert, California (long dashed line), East Mojave Desert, California and Nevada (short dashed line) and Sonoran Desert, Arizona (solid line).

164

West Mojave Deserty = -0.25x + 12.04x2 + 28.29 (r² = 0.96) 200 East Mojave Deserty = 0.23x + 3.47x2 + 45.55 (r² = 0.83) Sonoran Deserty = -0.13x + 8.79x2 + 32.41 (r² = 0.86)

150

100

Carapace length (mm) 50

0 0 5 10 15 20 25 30 Age (years)

Figure 35. Age versus body size in juvenile Gopherus agassizii from the West Mojave Desert, California (long dashed line), East Mojave Desert, California and Nevada (short dashed line) and Sonoran Desert, Arizona (solid line).

165

300 Sonoran Desert y = -0.012x2 + 1.698x + 203.945 (r ² = 0.19) West Mojave y = -0.112x2 + 6.031x + 143.219 (r ² = 0.11) A 280 East Mojave y = -0.075x2 + 5.864x + 119.458 (r ² = 0.27)

260

240

220

200 Carapace length (mm)

180

160 10 15 20 25 30 35 40 45 Age (years) 320 Sonoran Desert y = -0.092x2 + 7.958x + 82.263 (r ² = 0.37) 2 300 West Mojave y = -0.091x + 7.809x + 112.833 (r ² = 0.31) East Mojave y = 2.599x2 + 0.553x + 224.095 (r ² = 0.14) B 280

260

240

220

Carapace length (mm) 200

180

160 0 10 20 30 40 50 60 70

Age (years)

Figure 36. Age versus body size in A) adult female and b) adult male Gopherus agassizii from the West Mojave Desert, California (long dashed line), East Mojave Desert, California and Nevada (short dashed line) and Sonoran Desert, Arizona (solid line).

166

4 Annual growth rate (mm/yr) 2

0 WM EM SD WM EM SD WM EM SD Juveniles Females Males

Figure 37. Box plot of annual growth rates of juvenile and adult Gopherus agassizii from the Sonoran Desert (SD) in Arizona and West (WM) and East Mojave (EM) Desert in California and Nevada. Horizontal lines represent median values, whiskers represent the 5th and 95th percentile and dots represent outliers.

167

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Vita

Amanda Jane Curtin Citizenship: South Africa Born 15 February, 1972

EDUCATION:

2006 - Ph.D. in Biological Sciences, Drexel University, Philadelphia USA 2000 – M.Sc. in Zoology, University of Stellenbosch, South Africa 1996 – B.Sc. Honors in Paleontology, University of Stellenbosch, South Africa 1995 – B.Sc., University of Stellenbosch, South Africa

PUBLICATIONS AND PRESENTATIONS:

Curtin, A.J., P.le F.N. Mouton and A. Chinsamy. 2005. Bone growth patterns in two cordylid lizards, Cordylus cataphractus and Pseudocordylus capensis. African Zoology 40(1):1-7.

Curtin, A.J., G.R. Zug, H.W. Avery and J.R. Spotila. 2005. Bone growth patterns as indicators of life history parameters in desert tortoise (Gopherus agassizii) populations. 5th World Congress of Herpetology. Stellenbosch, South Africa.

Curtin, A.J., G.R. Zug, H.W. Avery and J.R. Spotila. 2005. Bone growth patterns and indicators of life history parameters in desert tortoise (Gopherus agassizii) populations. Annual meeting of the Society of Comparative and Integrative Biology. San Diego, California.

Curtin, A.J. 2004. Bone growth patterns as indicators of life history parameters in Gopherus agassizii populations from arid environments. Joint Meeting Ichthyologists and Herpetologists. Norman, Oklahoma.

SCHOLARSHIPS AND FELLOWSHIPS:

1995: Leclair scholarship, R3 500 1996 - 1998: Boonstra Scholarship, total R15 000 1997 - 1998: National Research Foundation Masters Scholarship, total R28 000 1999: Stellenbosch 2000 scholarship, R9 300 2001 – 2004: Panaphil Foundation Fellowship, $100 000

AWARDS AND CERTIFICATES:

1999 - Best student presentation: Annual Symposium of the Zoological Society of Southern Africa. 2006 - Certificate in Teaching College Biology, Duke University, Durham USA