Thesis (Ph. D.)--University of Chicago, Committee on Evolutionary Biology, March 2001. / Includes bibliographical references. Also available on the Internet.

Ph.D. (Education) / This research focused on two short learning programmes in evolution education offered by the Department of Science and Technology Education in the Faculty of Education of the University of Johannesburg. The broad purpose of the study was to develop design principles for a series of short learning programmes on evolution offered by the University. In achieving these aims, the study looked at teachers’ conceptual understanding of evolution, which included a number of common myths and misconceptions; folk or naive evolutionary explanations; and affective and sociocultural factors that influence cognition. Results were obtained from questionnaires, narratives, observations, discussion, and pre-and post-tests. A fruitful analytic tool was developed – that of converting questionnaire responses into narratives in order to obtain a better within participant view of coherent versus fragmented thinking. Some the themes that emerged included the cognitive bias of essentialism and how this relates to the notion of phenomenological primitives or p-prims, and the mechanism of resubsumption. "Hot" and "cold" learning and the critical issues of both religious and racial objections to evolution emerged as a fairly dominant theme. Data also supported a view of knowledge-as-pieces rather than theory-like in this knowledge domain. Poor teacher knowledge and the need for longer learning programmes, are also discussed in the results. To mitigate these conceptual and affective barriers, a number of design principles are proposed and discussed for teacher professional development in this area including the importance of communities of practice to augment short learning programmes and support on-going professional development.

Stress is a ubiquitous aspect of everyday life. As such, there exists a great deal of variability in the individual response to stress, particularly as a functional cause of depression. The aim of this dissertation is to investigate the mechanisms behind individual differences in response to stressful events in the attempt to explain differing levels of vulnerability to depression and drug addiction following exposure to stress. To accomplish this goal, we examine variations in the stress response using a rodent model of individual differences based on novelty-seeking behavior. Outbred rats can be classified as either High Responders (HR) or Low Responders (LR) depending on their locomotor activity in a novel environment. Previous studies have demonstrated that HR and LR rats differ in key components of the stress response pathway and would thus make a good model of individual differences in response to stress. Of the many types of stressors that one might encounter in their daily activities, the most commonly experienced is social stress. We therefore utilized a rodent model of social stress termed social defeat to investigate whether exposure to social stress might induce depressive-like behaviors. We then examined histone modifications as a potential mechanism behind such behavioral alterations. Our results found that repeated social defeat induces a number of depressive-like behaviors in Sprague- Dawley rats that are correlated with short-term changes in histone acetylation in the hippocampus and the amygdala. We then focused on individual differences in response to social defeat and in histone modifications. We found that HR rats are more susceptible to the effects of social stress, as evidenced by the expression of depressive-like behaviors following exposure to social defeat. Additionally, HR rats differ from LR rats in the levels of hippocampal histone acetylation in both basal conditions and following exposure to social defeat. We investigated potential genes that may be responsible for our observed changes in acetylation. We found basal changes in cyclic-AMP response element binding (CREB) Binding Protein (CBP) mRNA between HR and LR rats. These results indicate a role for epigenetic mechanisms as a potential mechanism for individual differences in responses to stress. We then explored individual susceptibilities in acute versus repeated social defeat exposure. We found that while both HR and LR rats exhibit long-term memories to repeated social defeat, only HR rats display long-term memories of an acute social defeat exposure. We examined histone acetylation levels following an acute exposure to social defeat, and found changes in the timing of acetylation patterns between HR and LR rats in the hippocampus and amygdala. These findings again highlight the importance of considering individual differences in stress responses and uncover the HR/LR model as a potential model for posttraumatic stress disorder. Finally, in a collaborative study, we investigated the role of methionine in depression and drug addiction. We found that systemic injection of methionine, a methyl donor, blocked behavioral sensitization to cocaine and resulted in decreased cocaine self-administration in male Sprague-Dawley rats. We propose several follow-up studies for investigating the role of DNA methylation further, including investigation of specific DNA methyltransferases, such as DNMT3A, and their target genes. This particular enzyme has also been implicated in the pro-depressant behaviors following exposure to social defeat, providing a common mechanism for the fields of depression and addiction. / A Dissertation submitted to the Department of Biomedical Sciences in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Spring Semester, 2011. / November 8, 2010. / neurobiology, hippocampus, depression, stress, Novelty-seeking / Includes bibliographical references. / Approved: Mohamed Kabbaj, Professor Directing Dissertation; Zuoxin Wang, University Representative; Akash Gunjan, Committee Member; Carlos Bolanos, Committee Member; James Olcese, Committee Member.

Spiny lobsters (Family Palinuridae) are large, diverse, and abundant marine crustaceans, which have conquered tropical, subtropical, and temperate coastal waters around the globe despite strong predation pressure. The mechanisms and function of antipredation strategies for most species in this highly successful taxon, encompassing behavior, morphology, and life-history characteristics, are poorly understood, particularly against natural predators. I investigate mechanisms of antipredation in spiny lobster Panulirus argus in the open during the day, at night, and while sheltering diurnally in natural dens. I also examine the function of putatively defensive acoustic signals produced by P. argus during diurnal attacks by piscine predators and while escaping octopuses at night. I also compare and contrast the mechanism and survival value of antipredator behavior and morphology between sympatric Panulirus argus and P. guttatus. Finally, I investigate ontogenetic changes in defensive behavior by diurnally sheltered P. argus to chemically-mediated predator cues. Nearly 40 species of spiny lobsters produce a characteristic sound (termed stridulation), speculated to deter predation. The occurrence and efficacy of stridulation has not been documented quantitatively during encounters with natural predators. I examined sound production in the sympatric spiny lobsters Panulirus argus and P. guttatus during attacks by their common predator, gray triggerfish Balistes capriscus, to determine if lobsters produce sound during defense, how stridulation integrates with behavioral and morphological defenses, and how interspecific differences in sound production relate to efficacy in repelling predators. Both lobster species stridulated coincident with specific defensive actions during triggerfish attack. In P. argus, stridulation occurred both during antennal lunging and during escape attempts (rapid retreat by tailflips). Panulirus guttatus stridulated only coincident with tailflips and did not lunge. Same-sized individuals of P. guttatus were subdued ~3 times more quickly on average than P. argus. The two species differed also in the relative size of the primary defensive weapons, the spinose 2nd antennae, which were far more robust in P. argus, particularly at larger body sizes. These results suggest that stridulation is an integral component of aggressive defense and escape behavior in spiny lobsters. The timing of sound production during aggressive, retaliatory defensive behavior (lunging) by P. argus suggests an aposematic role for stridulation against triggerfish. Using staged encounters of P. argus with B. capriscus, I examined whether stridulation, coincident with thrusting spines during aggressive defense, functions aposematically or simply renders a defending lobster more difficult to subdue without playing an aposematic role. I demonstrate, by disabling the stridulating organ in some lobsters (muting), that sound plays a vital role in defense against inexperienced (naïve) triggerfish, resulting in fewer successful attacks in subsequent encounters. Choice experiments with triggerfish that previously bypassed defenses and consumed lobsters show that experienced attackers do not choose muted lobsters over stridulating individuals. I propose that stridulation by P. argus against triggerfish is aposematic, as part of a multi-modal display, advertising the lobster's spiny defenses to predators. It is widely documented that sound production in P. argus and other spiny lobsters accompanies grasping of the carapace or other disturbance by human captors. Additionally, stridulation accompanies tailflip escape attempts during attacks by triggerfish. Although sound production during daytime attacks does not appear to increase survival against triggerfish, stridulating during escape may be more effective against grasping predators like octopus. Here, I investigate P. argus defensive behavior during nighttime encounters with Caribbean reef octopus Octopus briareus to determine whether P. argus stridulate during octopus attacks, how stridulation is used along with other defensive behavior (e.g. tailflips), and whether sound production improves survival in stridulating individuals. Lobsters stridulate both during grasping attacks by octopus and during escape attempts after being captured. Stridulating lobsters are also more likely to escape from attacking octopuses and remain uncaptured longer during encounters. I suggest that improving the efficacy of tailflip escapes against octopus may have been the function for which the stridulating organ initially evolved in the Stridentes clade of the Palinuridae. Benthic stages of P. argus reside in shelters during the day as a primary means of antipredation. However, when an active predator approaches and/or successfully attacks a nearby conspecific, these individuals must decide whether to emigrate quickly from the area or remain in shelter (dens or macroalgae) and rely on crypticity, defensive behavior, or the presence conspecifics to avoid attack, injury, or death. In this study, I examine how the three benthic juvenile phases of Caribbean spiny lobster Panulirus argus respond to exposure to fresh conspecific body fluid and how antipredator behavior, particularly the decision to stay or leave the area, changes during ontogeny. Additionally, I examined how the presence of conspecifics affects the decision to stay or leave in gregarious juvenile stages of P. argus. Although all size classes of P. argus respond to alarm odor, the decision to stay or leave dens changes unexpectedly with increasing body size and in the presence of conspecifics. Once shelters were abandoned, body size was a strong indicator of distance traveled in response to alarm odor. This indicates that Panulirus argus undergo an ontogenetic shift in defensive behavior, more frequently leaving dens in response to alarm odor and traveling across open substrate during the day, but only after reaching a body size at which they can effectively defend against predators. / A Dissertation submitted to the Department of Biological Science in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Fall Semester, 2006. / October 4, 2006. / Marine Biology, Panulirus argus, Lobster, Triggerfish, Octopus, Aposematism, Bioacoustics, Behavioral Ecology / Includes bibliographical references. / William F. Herrnkind, Professor Directing Dissertation; David E. Thistle, Outside Committee Member; Joseph Travis, Committee Member; Walter R. Tschinkel, Committee Member; Thomas A. Houpt, Committee Member.

Gag grouper, Mycteroperca microlepis, compose an important recreational and commercial fishery along the Gulf coast and Eastern Seaboard of the United States (Coleman et al. 2000, Morris et al. 2000, and Musick et al. 2000). Like many Serranids, gag have a complex life cycle in which juveniles spend each summer developing within inshore structured habitats before joining the adult population offshore (Heemstra and Randall 1993, Koenig et al. 1996, Grover et al. 1998). As is the case with most fisheries, heavy fishing pressures have led to a decline in gag abundance (Renan et al. 1996). The most efficient policies to mitigate for these population have as of yet focused on adult gag preservation; this study, however, attempt to use select juvenile gag population metrics (abundance and date of fertilization) to provide crucial data that may one day assist in providing a juvenile-centric population model. Over the course of a three-year study, juveniles were captured in two regions; near the Florida State University Coastal and Marine Laboratory (Turkey Point Shoal and Lanark Reef sites) and inside of Saint Andrew Bay, Panama City. Otolith analysis was performed to determine age and fertilization date of the juveniles, and abundance statistics were determined for each year. It was found that, consistent with previous literature, average fertilization date varied significantly across years and sampling sites (range = 41.9 to 91.8). The 2009 field season exhibited a significantly earlier fertilization date for both regions, and only one specimen was captured in Saint Andrew Bay. The 2009 Turkey Point Shoal field season was the only occurrence where abundance significantly declined during the course of the field season. Various environmental and physical factors may have been responsible for the enormous interannual variation seen in both abundance and fertilization date, such as large-scale current patterns and predator densities in the two regions. While no concrete conclusions may be drawn from the data, this study was essential in expanding on background knowledge of early juvenile gag population dynamics. / A Thesis submitted to the Department of Oceanography in partial fulfillment of the requirements for the degree of Master of Science. / Summer Semester, 2011. / May 20, 2011. / Juvenile Grouper, Seagrass, Nursery Habitat, Abundance / Includes bibliographical references. / David Thistle, Professor Directing Thesis; Chris Koenig, Professor Co-Directing Thesis; Markus Huettel, Committee Member; Jeff Chanton, Committee Member.

The traditional method for producing taxonomic illustrations requires the preparation of inked copies of pencil originals. These individual figures are then grouped into plates for publication. I describe an alternative, computer-based approach. In a preliminary step, a pencil drawing is digitized and imported into an illustration program. The program's tools are then used to trace the image. Each figure is stored as a separate computer file. To make a plate for publication or a slide for presentation, several figures can be combined. The approach has several advantages. (1) It is easier to master than the pen-and-ink method. (2) Figures can be manipulated electronically, so the size and orientation of a figure is infinitely adjustable, providing great flexibility in the preparation of plates. (3) Multiple "originals" can be printed. (4) Mistakes can be corrected easily. I describe two new species of Diosaccidae (Harpacticoida, Copepoda) from an unvegetated sand at 18 m depth in the northern Gulf of Mexico. One is assigned to the genus Protopsammotopa; the other is assigned to Actopsyllus, and traits in common with Eoschizopera Wells & Rao (1976) are noted. I used the computer-based techniques to produce the figures for these descriptions. The effects of winter storms on the two new diosaccids described here were studied through a reanalysis of data from Thistle et al. (1995a). I found that Actopsyllus sp. nov. did not migrate downward in response to erosive flow. Protopsammotopa sp. nov. males, but not females, did. Protopsammotopa sp. nov. is found at shallower depths in the sediment than Actopsyllus sp. nov., which may explain the difference in response to erosion by two morphologically similar and phylogenetically related species. / A Dissertation submitted to the Department of Oceanography in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Fall Semester, 2003. / September 3, 2003. / Taxonomy, Benthos, Harpacticoids, Meiofauna, Storms / Includes bibliographical references. / David Thistle, Professor Directing Dissertation; William Herrnkind, Outside Committee Member; Richard Iverson, Committee Member; William Landing, Committee Member; Nancy Marcus, Committee Member.

The study of adaptive molecular evolution in natural populations has been severely limited by the difficulty of linking genetic variation to phenotypic variation to fitness effects. Most studies connecting genotype, phenotype, and fitness have used reverse genetic approaches to measure the functional effects of specific mutations in the laboratory because this relationship is difficult to measure in natural populations, particularly for complex traits because of the "many-to-one" mapping of genotype to phenotype. Many of the fundamental features of evolving systems, such as evolvability, epistasis, and pleiotropy, however, may be stronger determinants of evolutionary outcomes in natural populations than in the laboratory because artificial selection and breeding schemes are generally more simplistic relative to selection and demographic effects in natural settings. Snake venoms have emerged as a system for the study of the genetics of adaptation in complex, polygenic traits because of their genetic tractability and role in feeding, digestion, and defense, all of which are directly relevant to fitness. Because venom gene expression is tissue-specific (i.e., no pleiotropic constraints) and toxin protein abundance directly influences venom efficacy, venoms are not inherently biased toward a particular mutational pathway, enabling a systematic comparison of the molecular mechanisms underlying adaptive evolution. Venom phenotypes are manifest only upon injection into another animal, and venom functions are directly measurable through various assays, allowing direct tests of adaptive hypotheses in natural prey populations. In this work, we sought to create a genotype-phenotype-fitness map for the venom system of the eastern diamondback rattlesnake (Crotalus adamanteus) and, for the first time, identify the genetic basis of adaptation for a complex, polygenic trait in natural populations. Crotalus adamanteus is the largest species of rattlesnake and exclusively consumes endotherms. Crotalus adamanteus is historically native to seven states in the southeastern United States but has recently been extirpated from Louisiana, is endangered in North Carolina, and is currently under consideration for listing as threatened under the Endangered Species Act. In Chapter 1, we sequenced the venom-gland transcriptome and integrated mass spectrometry data to construct a transcriptome-proteome map for the venom system. We then used this map to identify significant toxin-gene expression differentiation across the range of C. adamanteus, providing candidate-genes for which to test the functional and evolutionary significance of the identified variation. In Chapter 2, we used a similar approach and identified significant ontogenetic differentiation in toxin gene expression; further analyses determined that ontogenetic effects explained more variation in toxin expression than geographic effects, although both juvenile and adult expression patterns varied geographically, and time-series experiments in lab-raised individuals demonstrated that geographic and ontogenetic expression differentiation were not environmentally induced but rather under genetic control. In Chapter 3, we used in vitro functional assays to verify that the expression differences found in the previous two chapters corresponded to differences in venom function. We found that, overall, the statistical differences in toxin expression outlined in the first two chapters equated to functional differences in toxic activities in a predictable, tractable manner, suggesting that the differences identified in the first two chapters were, in fact, biologically relevant. In Chapter 4, we used a target-enrichment approach to sequence the exons of all identified toxins in the venom-gland transcriptome as well as several thousand neutral loci to ascertain the relative roles of expression versus coding-sequence changes in a trait not inherently biased towards either mutational pathway. We found evidence for adaptive changes at both the expression and sequence levels across the entire range, although expression differentiation did appear to be the more frequent molecular mechanism. But, without functional characterizations of the identified sequence and expression evolution, it was difficult to characterize the relative roles demography, selection, and drift played in generating the identified sequence and expression divergence. Although Chapter 3 did link expression variation to functional variation, these assays were not conducted in the actual target of venoms, natural prey. To address these issues, we examined toxin sequence and expression evolution and estimated venom toxicity (i.e., fitness) in sympatric and allopatric natural prey across an island-mainland population pair in Chapter 5 to, for the first time, construct a genotype-phenotype-fitness map for a complex trait in natural populations. We found that expression differentiation was predominantly, or exclusively, the genetic basis of polygenic adaptation, suggesting that over ecological timescales complex traits may preferentially evolve through mutations affecting expression because more molecular mechanisms exist for altering the amount of protein produced than for altering their functions through their primary sequences. In Chapter 1, we found significant expression differentiation in both high- and low-abundance proteins across the range and over 1 million years of divergence, and in Chapter 4, we found both sequence and expression differentiation across the same temporal and spatial scales. In Chapter 5, however, we only identified expression differentiation, and found that this expression differentiation was restricted to low-expression proteins because of physiological and selective constraints on high-expression proteins. These differences in the molecular mechanism underlying adaptive evolution were most likely the result of temporal constraints on generating beneficial variation; because more molecular mechanisms exist for altering protein amounts than protein function, the probability of generating a beneficial expression variant is greater than the probability of generating a beneficial point mutation in the coding-region of a specific protein, and these differences in probability would be most pronounced over extremely short timescales. Given enough time, however, both mutational pathways and proteins expressed at all levels can generate beneficial variation, and these results provide qualitative predictions regarding the process of adaptation for a complex trait. / A Dissertation submitted to the Department of Biological Science in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Fall Semester 2016. / November 8, 2016. / adaptation, gene flow, protein expression / Includes bibliographical references. / Darin Rokyta, Professor Directing Dissertation; Peter Beerli, University Representative; Greg Erickson, Committee Member; Joseph Travis, Committee Member; Alice Winn, Committee Member.

The Pine Barrens treefrog (Hyla andersonii) is restricted to three isolated (disjunct) regions in the eastern United States: New Jersey, North and South Carolina, and the Florida panhandle and southern Alabama. It is a seepage bog specialist and a species of conservation concern in each state in which it occurs. Previous work showed some differentiation among the three regions; however, these studies had small sample sizes, measured few variables, and/or did not include all three regions. Moreover, the only genetic study of H. andersonii, using allozyme data, did not resolve clear relationships among the three regions. Conservation management of this species requires a clearer understanding of how populations in these three regions may differ from one another and how populations within regions are structured. To extend previous work on this species, I measured differentiation among regions using morphometric, acoustic signal, ecological, and genetic data. I developed 15 microsatellite markers and used targeted sequence capture to collect large-scale nuclear and mitochondrial genomic data to test models of its evolutionary history. It is hypothesized that the species shifted southward during the last glacial maximum (LGM) into one or more refugia, then expanded northward as the glaciers receded. Overall I find a strikingly concordant pattern in which the first axis of variation for each of the data types distinguishes populations along a latitudinal and longitudinal gradient and the second axis distinguishes the set of populations occurring in the Carolinas (CL) from those occurring in the New Jersey (NJ) and Alabama/Florida (AF) regions. I know of no comparable data set that displays such concordance among different types of data across so large a geographic range. The overlap in trait values (i.e. exchangeability) between neighboring regions, however, is substantial in all types of data, except genetic, which supports continued consideration of this taxon as a single species. Using a phylogenetic framework with large-scale genomic data, I found that AF forms a single clade in both the nuclear and mitochondrial trees and that AF is sister to the rest of the Atlantic clade (CL, NJ). Climate models suggest that the distribution of the species has been repeatedly disjunct since at least the last interglacial, but probably even earlier given the genetic divergence time estimates. All three regions also showed little overlap in broad-scale (climate) environmental data, although niche modeling using climate data alone closely matched the known distribution. Fine-scale environmental data (abiotic and biotic) show greater overlap between CL and AF, with some divergence from NJ. I found some association of genetic and morphometric measures with ecological values, mostly for the broad-scale data. Projections for 2050 suggest habitat suitability will be greatly reduced in CL and AF, with only a small area of NJ available. Finally, I found very different patterns of population genetic structure within each region. Little evidence for isolation by distance was found for all regions, suggesting isolation via environment or other factors may be important for connectivity between populations. The results of my research can be applied to the conservation and management of H. andersonii because it is the most comprehensive study of the species using both an ecological and evolutionary perspective across different temporal and spatial scales. / A Dissertation submitted to the Department of Biological Science in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Summer Semester 2016. / June 10, 2016. / conservation, disjunct distribution, exchangeability, Hyla andersonii, phylogeography, Pine Barrens Treefrog / Includes bibliographical references. / Emily Lemmon, Professor Co-Directing Dissertation; Joseph Travis, Professor Co-Directing Dissertation; Peter Beerli, University Representative; Scott Steppan, Committee Member; D. Bruce Means, Committee Member.

Differences in behavior and physiology can arise and change because of variation in life history strategies or differences in discrete polymorphisms. These differences can result from the indirect effects of predation, or responses to erratic, repeated exposures to predators in which the prey individual is not consumed (non-consumptive predation). Behavioral responses in the presence of predators, like changes in mating or antipredator behavior may differ because of life history variation or discrete morphs within populations. There are also physiological responses that occur with behavioral responses. These physiological responses occur through the animals’ stress response. A stress response is an accumulation of endocrine and neuroendocrine responses that return the body back to its normal metabolic and hormonal levels. The least killifish, Heterandria formosa is an excellent system to understand how life history variation influences responses to predation, while, melanic and silver morphs in the Eastern mosquitofish, gambusia holbrooki, are ideal for understanding how variation within populations influence responses to predation. The overarching goals of this dissertation were to evaluate variation in behavior and physiology from predation risk as a result of differences between populations of the least killifish and within a population of the Eastern mosquitofish as well as to review pleiotropic effects that stem from the central melanocortin system and their potential influence on behavior and stress within and between populations. To answer our first questions in the least killifish we created control and predator treatments and measured both populations’ differences in male mating behavior when exposed to a predator. We also analyzed female cortisol levels between the two populations to see if their stress responses varied. We exposed male and female least killifish from both populations to predator and control treatments over 30 days to measure the effect of treatment and population on reproductive output. Results showed that males from the high-predation population were more active overall than males from the low predation population in the control treatment but that males from both populations responded to the presence of predators. Females from the low predator population had higher cortisol levels in predator treatments however cortisol levels from females in high predator population were similar in control and predator treatments. Our results reflect conclusions of another study in which predator presence did not influence reproductive output in another population of H.formosa. In the Eastern mosquitofish we exposed silver and melanic males to predator and control treatments and followed behavior assays with cortisol analyses. We found that melanic males were more active than silver males in their mating behavior and this result is consistent with other studies. All males decreased their mating behaviors in the presence of predators and increased antipredator freezing behavior in predator treatments. Melanic males had higher cortisol levels on average than silver males in control treatments; melanic male cortisol levels were only half as high in the predator treatments as in the control treatments. Differences in morph behaviors and physiology may be mediated by the pleiotropic effects correlated with black coloration and also may help rare eastern mosquitofish morphs persist. Finally, we reviewed the melanocortin system and its pleiotropic effects determined the frequencies of melanic morphs within natural populations and reflected on other pathways that influence black coloration. We found that melanic morphs are rare in polymorphic populations. We also found that populations with discrete polymorphisms mostly consisted of melanic morphs that occurred 33% or less of the time in the population with melanic morphs frequencies higher than 75%. In conclusion, this work is the first to review the frequencies of melanic morphs in populations and assess behavior and stress response in the least killifish and Eastern mosquitofish. We highlight the importance of assessing short-term behavioral responses and also measuring long-term responses through predation exposure in H.formosa. We also provide evidence for the use of short and long-term measures and not solely relying on short-term responses as a means to predict fitness. We also demonstrate how variation within populations, specifically melanic coloration may influence differences in morph behaviors, the propensity to be approached or attacked by predators, and additional behavioral and physiological responses that may arise because of pleiotropy in the melanocortin system. / A Dissertation submitted to the Department of Biological Science in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / 2019 / August 28, 2019. / Behavior, Life History, Melanin, Poeciliids, Predation / Includes bibliographical references. / Joseph Travis, Professor Directing Dissertation; Lisa Eckel, University Representative; Kimberly Hughes, Committee Member; Emily DuVal, Committee Member; Scott Burgess, Committee Member.

Biological hard materials are a remarkable class of materials combining large volumes of mineral with minute organic components into often complex, hierarchical microstructural arrangements. These intricate microstructures offer ideal systems from which form-function relationships can be dissected due to their limited functional demands. They are also of increasing interest to the materials science community due to their high combinations of stiffness and toughness unexpected of ceramic-like materials. Individually, each approach for understanding these materials has suffered from a lack of insight from the other field: the biological perspective has suffered from a lack of analytical rigor while the engineering perspective has been ignorant to the intricacies of evolution as needed to accurately infer the original and current function of these structures. Here I present and execute a unified framework for examining biological hard materials. In order to identify the mechanical import of microstructural changes, this framework tests changes in biologically relevant material properties by measuring mechanical response across the transformation series of microstructures observed in conjunction with ecological shifts. In order to apply this framework, I use mammalian dental enamel as a model system. Dental enamel is the most mineralized tissue in the vertebrate body and is non-repairable and irreplaceable if damaged. Arguably, it has only two functions: transfer masticatory loads to ingesta and resist its own degradation. In mammals, the evolution of a critical tissue constituent--the enamel prism--has resulted in a multitude of enamel microstructural arrangements, some of which have independently evolved consistently in ecologically similar contexts. I sought to characterize changes in the mechanical response of enamel microstructures by providing a survey of elastic modulus and fracture toughness for a diversity of mammals showing a broad array of microstructural forms. Considering the mechanics of damage to mammalian enamel as they pertain to documented microstructural changes within lineages, I then identified three critical functional transitions in enamel microstructures. These functional transitions include: (1) the evolution of the enamel prism, (2) the adaptation to a high wear diet, and (3) the adaptation to a high fracture diet. I investigated potential changes in material response across these transitions. Methodologically, I measured elastic modulus using instrumented nanoindentation across a series of reptilian and mammalian enamels to examine differences in resistance to elastic deformation. I then verified and executed a new method for determining the intrinsic fracture toughness of enamel, crack tip opening displacement, and identified changes in small scale resistance to fracture. I used Vickers microindentation to evaluate differences in resistance to plastic deformation. Lastly, I developed a novel method for quantifying fracture orientation, called Crack Analysis of Propagation Orientation (CAPO). CAPO identifies directions of preferred cracking and provides a proxy of resistance to large-scale fracture effects. These data provide consistent evidence that mammalian enamel microstructures are remarkably consistent in elastic modulus, intrinsic fracture toughness, and hardness. This consistency and their correspondence to values reported in the literature suggests that selection has acted to make enamel microstructures as stiff, hard, and intrinsically tough as possible given the inherent developmental constraints of amelogenesis and material constraints of hydroxyapatite. However, they display marked quantitative and qualitative differences in their resistance to large-scale fracture. Contact with hard particulates in the environment such as plant phytoliths or exogenous grit are expected to result in local indentation damage and the removal of enamel through microcrack growth. Grazing taxa have enamels which include modified radial enamel, a microstructure that channels indentation crack growth into a single direction and suppresses subsurface lateral crack growth. Together, these mechanisms would reduce the removal of enamel pieces by inhibiting microcrack coalescence and offer increased resistance to severe wear. Conversely, contact with large objects such as bone are expected to result in fractures which propagate across the tooth surface. Carnivoran Hunter-Schreger bands qualitatively suppress fracture across bands; this behavior could provide resistance to fatigue crack growth. These results provide evidence that mammalian enamel microstructures are consistent in many of the commonly reported material properties but differ primarily in their large-scale fracture behavior. They further offer avenues for biomimetic ceramic composites with consistent hardness and moduli but with potential damage and fatigue tolerance specific to the loading scenario. / A Dissertation submitted to the Department of Biological Science in partial fulfillment of the requirements for the degree of Doctor of Philosophy. / Summer Semester 2018. / July 6, 2018. / Biomimetic, Enamel, Mammal, Material Properties, Microstructure, Teeth / Includes bibliographical references. / Gregory Erickson, Professor Directing Dissertation; William Oates, University Representative; Brian Inouye, Committee Member; William Parker, Committee Member; Scott Steppan, Committee Member.