Predation Risk, and Not Shelter or Food Availability, as the Main Determinant of Reproduction Investment in Island Lizards.

Reproductive investment, including the number of offspring produced, is one of the fundamental characteristics of a species. It is particularly important for island vertebrates, which face a disproportionate number of threats to their survival, because it predicts, among other things, a species' resilience to environmental disruption. Taxa producing more offspring recover more quickly from environmental perturbations and survive environmental change better. However, ecologists do not understand which primary drivers shape a species' reproductive investment well. Here, we compare the reproductive efforts of 14 island populations of the Aegean Wall Lizard (Podarcis erhardii), which lives across widely diverging environmental conditions. We test three hypotheses, namely that reproductive investment (measured as clutch size, clutch volume) is (1) positively associated with predation risk ['Predation Risk Hypothesis']; (2) positively associated with the presence of reliable vegetation cover that provides shelter ['Gravid Female Protection Hypothesis']; and (3) limited by (and hence positively correlated with) food availability ['Food Limitation Hypothesis']. Although field data are somewhat consistent with all three hypotheses, statistical analyses provide strong support for the Predation Risk Hypothesis. The results not only shed light on which fundamental forces shape reproductive investment in island vertebrates, but can also help shape conservation priorities.

Small Island Effects on the Thermal Biology of the Endemic Mediterranean Lizard Podarcis gaigeae.

Ectotherms are vastly affected by climatic conditions as they rely on external sources of heat to regulate their body temperature, and changes in their habitat thermal quality could seriously affect their overall biology. To overcome the problems of a thermally unfavorable habitat, lizards need to either adjust their thermoregulatory behavior or respond to directional selection and shift their preferred body temperatures. To assess the impact of habitat thermal quality on the thermoregulatory profile, we studied multiple islet and 'mainland' populations of the Skyros wall lizard Podarcis gaigeae, an endemic lacertid to Skyros Archipelago, Greece. We evaluated the effectiveness of thermoregulation (E) using the three main thermal parameters: body (Tb), operative (Te), and preferred (Tpref) temperatures. We first hypothesized that the spatial homogeneity, the scarcity of thermal shelters, and the exposure to higher winds on islets would result in more demanding climate conditions. Second, we anticipated that islet lizards would achieve higher E in response to the lower thermal quality therein. As hypothesized, thermal parameters differed between populations but not in the expected manner. Skyros 'mainland' habitats reached higher temperatures, had more intense fluctuations, and were of lower thermal quality. As a result, lizards showed higher accuracy, precision, and effectiveness of thermoregulation. Noteworthy, we found that lizards from different populations have shifted their thermal profile and preferred body temperatures to cope with the particular conditions prevailing in their habitats. The latter supports the labile view on the evolution of thermoregulation.

Tail regeneration alters the digestive performance of lizards.

Tissue regeneration is a fundamental evolutionary adaptation, which is well known in lizards that can regenerate their entire tail. However, numerous parameters of this process remain poorly understood. Lizard tail serves many functions. Thus, tail autotomy comes with many disadvantages and the need for quick regeneration is imperative. To provide the required energy and materials for caudal tissue building, lizards are expected to undergo a number of physiological and biochemical adjustments. Previous research showed that tail regeneration induces changes in the digestive process. Here, we investigated if and how tail regeneration affects the digestive performance in five wall lizard species deriving from mainland and island sites and questioned whether the association of tail regeneration and digestion is affected by species relationships or environmental features, including predation pressure. We expected that lizards from high predation environments would regenerate their tail faster and modify accordingly their digestive efficiency, prioritizing the digestion of proteins; the main building blocks for tissue repair. Second, we anticipated that the general food shortage on islands would inhibit the process. Our findings showed that all species shifted their digestive efficiency, as predicted. Elongation rate was higher in sites with stronger predation regime and this was also applied to the rate with which protein digestion raised. Gut passage time increases during regeneration so as to improve the nutrient absorbance, but among the islanders, the pace was more intense. The deviations between species should be attributed to the different ecological conditions prevailing on islands rather than to their phylogenetic relationships.

Tail regeneration affects the digestive performance of a Mediterranean lizard.

In caudal autotomy, lizards shed their tail to escape from an attacking predator. Since the tail serves multiple functions, caudal regeneration is of pivotal importance. However, it is a demanding procedure that requires substantial energy and nutrients. Therefore, lizards have to increase energy income to fuel the extraordinary requirements of the regenerating tail. We presumed that autotomized lizards would adjust their digestion to acquire this additional energy. To clarify the effects of tail regeneration on digestion, we compared the digestive performance before autotomy, during regeneration, and after its completion. Tail regeneration indeed increased gut passage time but did not affect digestive performance in a uniform pattern: though protein income was maximized, lipid and sugar acquisition remained stable. This divergence in proteins may be attributed to their particular role in tail reconstruction, as they are the main building blocks for tissue formation.

Sex determination by three-dimensional geometric morphometrics of craniofacial form.

SUMMARY: The purpose of the present study is to define which regions of the cranium, the upper-face, the orbits and the nasal are the most sexually dimorphic, by using three-dimensional geometric morphometric methods, and investigate the effectiveness of this method in determining sex from the shape of these regions. The study sample consisted of 176 crania of known sex (94 males, 82 females) belonging to individuals who lived in Greece during the 20(th) century. The three-dimensional co-ordinates of 31 ecto-cranial landmarks were digitized using a MicroScribe 3DX contact digitizer. Goodall's F-test was performed in order to compare statistical differences in shape between males and females. Generalized Procrustes Analysis (GPA) was used to obtain size and shape variables for statistical analysis. Shape, Size and Form analyses were carried out by logistic regression and discriminant function analysis. The results indicate that there are shape differences between the sexes in the upper-face and the orbits. The highest shape classification rate was obtained from the upper-face region. The centroid size of the caraniofacial and the orbital regions was smaller in females than males. Moreover, it was found that size is significant for sexual dimorphism in the upper-face region. As anticipated, the classification accuracy improves when both size and shape are combined. The findings presented here constitute a firm basis upon which further research can be conducted.

Sex determination by three-dimensional geometric morphometrics of the vault and midsagittal curve of the neurocranium in a modern Greek population sample.

The aim of this study is to assess sexual dimorphism of adult crania in the vault and midsagittal curve of the vault using three-dimensional geometric morphometric methods. The study sample consisted of 176 crania of known sex (94 males, 82 females) belonging to individuals who lived during the 20th century in Greece. The three-dimensional co-ordinates of 31 ecto-cranial landmarks and 30 semi-landmarks were digitized using a MicroScribe 3DX contact digitizer. Generalized Procrustes analysis (GPA) was used to obtain size and shape variables for statistical analysis. Shape, size and form analyses were carried out by logistic regression and three discriminant function analyses. Results indicate that there are shape differences between sexes. Females in the region of the parietal bones are narrower and the axis forming the frontal and occipital bones is more elongated; the frontal bone is more vertical. Sex-specific shape differences give better classification results in the vault (79%) compared with the midsagittal curve of the neurocranium (68.8%). Size alone yielded better results for cranial vault (82%), while for the midsagittal curve of the vault the result is poorer (68.1%). As anticipated, the classification accuracy improves when both size and shape are combined (89.2% for vault, and 79.4% for midsagittal curve of the vault). These latter findings imply that, in contrast to the midsagittal curve of the neurocranium, the shape of the cranial vault can be used as an indicator of sex in the modern Greek population.

Immune responses of a wall lizard to whole-body exposure to radiofrequency electromagnetic radiation.

PURPOSE: During the last three decades, the number of devices that emit non-ionizing electromagnetic radiation (EMR) at the wireless communication spectrum has rapidly increased and possible effects on living organisms have become a major concern. The purpose of this study was to investigate the effects of radiofrequency EMR emitted by a widely used wireless communication device, namely the Digital Enhanced Communication Telephony (DECT) base, on the immune responses of the Aegean wall lizard (Podarcis erhardii). MATERIALS AND METHODS: Adult male lizards were exposed 24 h/day for 8 weeks to 1880-1900 MHz DECT base radiation at average electric field intensity of 3.2 V/m. Immune reactivity was assessed using the phytohemagglutinin (PHA) skin swelling and mixed lymphocyte reaction (MLR) tests. RESULTS: Our results revealed a noticeable suppression (approximately 45%) of inflammatory responses in EMR-exposed lizards compared to sham-exposed animals. T cell-mediated responses were marginally affected. CONCLUSION: Daily radiofrequency EMR exposure seems to affect, at least partially, the immunocompetence of the Aegean wall lizard.

Effects of insularity on digestion: living on islands induces shifts in physiological and morphological traits in island reptiles.

Living on islands entails numerous challenges for animals, among which resource scarcity stands out. In order to survive, animals have to optimize energy acquisition. We examined the impact of insularity on digestion comparing a series of physiological and morphological traits of adult males between insular and mainland populations of the Balkan green lizard. Island lizards had longer gastrointestinal tracts and gut passage times and higher digestive efficiencies. The dissection of the hindgut revealed an unexpected finding, the presence of cecal valves that were more frequent in island lizards. Thanks to all above islanders retain food for longer periods and thus maximize energy income and increase the amount of the extracted nutrients. That way, they secure energy income from the limited, in time and quantity, food resources of the islands.

Evaluating the efficiency of different recording protocols for entheseal changes in regards to expressing activity patterns using archival data and cross-sectional geometric properties.

OBJECTIVES: In the past decade there have been extensive discussions on the potential utility of entheseal changes (EC) as activity markers. Nevertheless, no study to date has compared different EC recording protocols with respect to their correlation to activity patterns. MATERIALS AND METHODS: This article records changes on fibrocartilaginous entheses of the upper limbs of 90 male skeletons from the documented Athens Collection using the Hawkey and Merbs (Int J Osteoarchaeol 5 (1995) 324-338), Mariotti et al. (Collegium Antropol 28 (2004) 145-159), and Villotte et al. (Am J Phys Anthropol 142 (2010) 224-234) recording schemes in order to determine which one exhibits the highest correlation with activity. Activity is assessed by means of the recorded profession of each individual, as well as employing cross-sectional geometric properties. Generalized Linear Models are used to explore the impact of age, body mass, and activity on EC expression. RESULTS: Our results agree with previous studies that age is the primary factor determining EC, whereas body mass is the second most influential factor. In contrast, activity in the form of profession or cross-sectional geometry rarely showed a significant correlation to EC expression and no clear pattern could be discerned irrespective of the recording technique. However, bilateral differences in the impact of age and body mass in EC expression were traced and may relate to activity patterns. CONCLUSIONS: The differences found in the bilateral impact of age and body mass highlight the fact that the activity patterns of the individuals under examination must play an underlying role to EC expression, though current recording schemes for EC do not capture this, rendering further work in the direction of developing more elaborate recording standards imperative.

Application of Purkait's triangle method on a skeletal population from southern Europe.

The existence of sexual dimorphism in individual elements of the human skeleton allows sex determination and significantly enhances the identification of fragmentary skeletal material. Traditionally, the pelvis and the skull are used for an accurate sex determination. Sometimes they are not available and alternative methods have to be used. The aim of this work is to study the sexual dimorphism of the proximal portion of the posterior femur by analyzing the biometric data of the Purkait's triangle. The studied sample comprises 203 individuals (112 males and 91 females) from the Modern Human Skeletal Reference Collection (The Athens Collection) of the Department of Animal & Human Physiology (National & Kapodistrian University of Athens). The biometric data were analyzed by Discriminant Analysis and equations were generated for sex determination, which gave an overall correct classification of 78.3%. For the right and the left femur the percentages of correct classification were 77.8% and 75.9% respectively. These accuracy rates were compared to those of an Indian population and were found to be lower (86.5% for males and 86.3% for females). The study of sexual dimorphism (SD) in Greek, Indian, European-American and African-American populations gave interesting results. The AB diameter in the Greek sample shows the lowest degree of SD while the AC diameter shows the highest. In the other three population samples, the BC diameter shows the highest degree of SD. We assume that the femur anatomy of the Greeks is the major cause that the Purkait method does not give high rates of correct classification. It is therefore advisable that practitioners exercise caution when using this method on skeletal material from Greece, especially in forensic contexts.

Molecular systematics and historical biogeography of the green lizards (Lacerta) in Greece: insights from mitochondrial and nuclear DNA.

The green lizards of the genus Lacerta (Sauria, Lacertidae) comprise nine recognized species, which in Europe are mainly restricted to the southern peninsulas. Four of them (L. trilineata, L. viridis, L. bilineata and L. agilis) occur in Greece. The uncertainty of morphological diversification renders the taxonomic assignment into species and subspecies problematic. In this study sequence data derived from two mitochondrial (cytochrome b and 16S rRNA) genes and one nuclear (NKTR) gene were used to (a) evaluate the taxonomic status of the genus Lacerta in Greece with emphasis on L. trilineata group and (b) investigate the evolutionary history of the genus through the application of phylogenetic and phylogeographic analyses, using Gallotia and Timon as outgroups. The phylogenetic analyses revealed the existence of four major clades. The first clade corresponds to L. trilineata group, the second to L. media, the third to L. agilis and the fourth to a complex of L. viridis and L. bilineata. However, the produced phylogenetic relationships are not congruent with the current taxonomy, especially in the first clade in which L. trilineata appeared to be paraphyletic in regard to L. pamphylica. Six distinct lineages were inferred within L. trilineata, despite the current recognition of nine morphological subspecies, the genetic differentiation of which exceeds that of other Lacerta species, imposing a thorough taxonomic revision of the species. Our results suggested a rapid diversification of L. trilineata group during the late Miocene. We believe that the present distribution of the genus in Greece is the result of several dispersal and vicariant events that took place during the late Miocene and early Pliocene.

Sex determination by three-dimensional geometric morphometrics of the palate and cranial base.

The purpose of this study is to assess sexual dimorphism in the palate and base of adult crania using three-dimensional geometric morphometric methods. The study sample consisted of 176 crania of known sex (94 males, 82 females) belonging to individuals who lived during the 20th century in Greece. The three-dimensional co-ordinates of 30 ectocranial landmarks were digitized using a MicroScribe 3DX contact digitizer. Generalized Procrustes Analysis (GPA) was used to obtain size and shape variables for statistical analysis. Three discriminant function analyses were carried out: (1) using PC scores from Procrustes shape space, (2) centroid size alone, and (3) PC scores of GPA residuals which includes InCS for analysis in Procrustes form space. Results indicate that there are shape differences between sexes. In males, the palate is deepest and more elongated; the cranial base is shortened. Sex-specific shape differences for the cross-validated data give better classification results in the cranial base (77.2%) compared with the palate (68.9%). Size alone yielded better results for cranial base (82%) in opposition to palate (63.1%). As anticipated, the classification accuracy improves when both size and shape are combined (90.4% for cranial base, and 74.8% for palate).

Mitochondrial phylogeny and biogeographic history of the Greek endemic land-snail genus Codringtonia Kobelt 1898 (Gastropoda, Pulmonata, Helicidae).

The aim of this work was to infer the phylogeny of the Greek endemic land-snail genus Codringtonia Kobelt 1898, estimate the time frame of the radiation of the genus, and propose a biogeographic scenario that could explain the contemporary distribution of Codringtonia lineages. The study took place in the districts of Peloponnese, Central Greece and Epirus of mainland Greece. Sequence data originating from three mtDNA genes (COI, COII, and 16S rDNA) were used to infer the phylogeny of the eight nominal Codringtonia species. Furthermore, the radiation time-frame of extant Codringtonia species was estimated using a relaxed molecular clock analysis and mtDNA substitution rates of land snails. The phylogenetic analysis supported the existence of six Codringtonia lineages in Greece and indicated that one nominal species (Codringtonia neocrassa) might belong to a separate genus distantly related to Codringtonia. The time frame of differentiation of Codringtonia species was placed in the Late Miocene-Pleistocene epoch. The dispersal-vicariance analysis performed indicated that most probably Codringtonia exhibited a north-to-south spread with the ancestral area being that of central Greek mainland, accompanied with duplication (speciation) and vicariance events.

Island biology and morphological divergence of the Skyros wall lizard Podarcis gaigeae: a combined role for local selection and genetic drift on color morph frequency divergence?

BACKGROUND: Patterns of spatial variation in discrete phenotypic traits can be used to draw inferences about the adaptive significance of traits and evolutionary processes, especially when compared to patterns of neutral genetic variation. Population divergence in adaptive traits such as color morphs can be influenced by both local ecology and stochastic factors such as genetic drift or founder events. Here, we use quantitative color measurements of males and females of Skyros wall lizard, Podarcis gaigeae, to demonstrate that this species is polymorphic with respect to throat color, and the morphs form discrete phenotypic clusters with limited overlap between categories. We use divergence in throat color morph frequencies and compare that to neutral genetic variation to infer the evolutionary processes acting on islet- and mainland populations. RESULTS: Geographically close islet- and mainland populations of the Skyros wall lizard exhibit strong divergence in throat color morph frequencies. Population variation in throat color morph frequencies between islets was higher than that between mainland populations, and the effective population sizes on the islets were small (Ne:s < 100). Population divergence (FST) for throat color morph frequencies fell within the neutral FST-distribution estimated from microsatellite markers, and genetic drift could thus not be rejected as an explanation for the pattern. Moreover, for both comparisons among mainland-mainland population pairs and between mainland-islet population pairs, morph frequency divergence was significantly correlated with neutral divergence, further pointing to some role for genetic drift in divergence also at the phenotypic level of throat color morphs. CONCLUSIONS: Genetic drift could not be rejected as an explanation for the pattern of population divergence in morph frequencies. In spite of an expected stabilising selection, throat color frequencies diverged in the islet populations. These results suggest that there is an interaction between selection and genetic drift causing divergence even at a phenotypic level in these small, subdivided populations.

Anatomical and physiological changes associated with a recent dietary shift in the lizard Podarcis sicula.

Dietary shifts have played a major role in the evolution of many vertebrates. The idea that the evolution of herbivory is physiologically constrained in squamates is challenged by a number of observations that suggest that at least some lizards can overcome the putative physiological difficulties of herbivory on evolutionary and even ecological timescales. We compared a number of morphological and physiological traits purportedly associated with plant consumption between two island populations of the lacertid lizard Podarcis sicula. Previous studies revealed considerable differences in the amount of plant material consumed between those populations. We continued the investigation of this study system and explored the degree of divergence in morphology (dentition, gut morphology), digestive performance (gut passage time, digestive efficiency), and ecology (endosymbiont density). In addition, we also performed a preliminary analysis of the plasticity of some of these modifications. Our results confirm and expand earlier findings concerning divergence in the morphology of feeding structures between two island populations of P. sicula lizards. In addition to the differences in skull dimensions and the prevalence of cecal valves previously reported, these two recently diverged populations also differ in aspects of their dentition (teeth width) and the lengths of the stomach and small intestine. The plasticity experiment suggests that at least some of the changes associated with a dietary shift toward a higher proportion of plant material may be plastic. Our results also show that these morphological changes effectively translate into differences in digestive performance: the population with the longer digestive tract exhibits longer gut passage time and improved digestive efficiency.

Tail shedding in island lizards [Lacertidae, Reptilia]: decline of antipredator defenses in relaxed predation environments.

The ability of an animal to shed its tail is a widespread antipredator strategy among lizards. The degree of expression of this defense is expected to be shaped by prevailing environmental conditions including local predation pressure. We test these hypotheses by comparing several aspects of caudal autotomy in 15 Mediterranean lizard taxa existing across a swath of mainland and island localities that differ in the number and identity of predator species present. Autotomic ease varied substantially among the study populations, in a pattern that is best explained by the presence of vipers. Neither insularity nor the presence of other types of predators explain the observed autotomy rates. Final concentration of accumulated tail muscle lactate and duration of movement of a shed tail, two traits that were previously thought to relate to predation pressure, are in general not shaped by either predator diversity or insularity. Under conditions of relaxed predation selection, an uncoupling of different aspects of caudal autotomy exists, with some elements (ease of autotomy) declining faster than others (duration of movement, lactate concentration). We compared rates of shed tails in the field against rates of laboratory autotomies conducted under standardized conditions and found very high correlation values (r > 0.96). This suggests that field autotomy rates, rather than being a metric of predatory attacks, merely reflect the innate predisposition of a taxon to shed its tail.

Cross-species testing of 27 pre-existing microsatellites in Podarcis gaigeae and Podarcis hispanica (Squamata: Lacertidae).

We tested 27 microsatellite loci for cross-species amplification in the lacertids Podarcis gaigeae and Podarcis hispanica. We detected 11 and 15 polymorphic loci in the former and the latter species, respectively. In a larger sample of individuals from a single population of each species, the number of alleles ranged from five to 23 in 10 of the polymorphic loci in P. gaigeae, and between four and 13 in nine of polymorphic loci in P. hispanica. Two locus deviated from Hardy-Weinberg equilibrium in P. hispanica. Between 11 and 16 of the 27 loci also amplified successfully in three other Podarcis species.

Combining immunological and molecular data to assess phylogenetic relations of some Greek Podarcis species.

Most recent molecular studies revealed the phylogeny of Greek Podarcis species, which for years remained elusive, due to discordant data produced from various chromosomal, complement fixation and protein studies. In this report, we analyzed cellular immune responses of spleen-derived lymphocytes from six allopatric Podarcis species encountered in Greece, by assessing two-way mixed lymphocyte reaction (MLR)-induced proliferation. On the basis of stimulation indices (S.I.) as determined from cultures set up from xenogeneic splenocytes coincubated in pairs, we generated a phylogenetic tree, fully consistent with the phylogenetic relationships of Podarcis as determined by parallel analyses based on partial mitochondrial (mt) DNA sequences. Although the exact mechanisms triggering lymphocyte responses in lizard two-way xenogeneic MLR are not fully understood, our results show the potential use of cell-mediated immune responses as an additional approach to mtDNA analysis, for species delimitation within specific lizard taxa.

Comparative postautotomy tail activity in six Mediterranean lacertid lizard species.

Tail autotomy, the self-induced tail separation from the body, is a common and effective antipredator mechanism in lizards. In this study, we examine the muscle energetics of tail shedding in six lacertid lizard species (Podarcis erhardii, Podarcis peloponnesiaca, Podarcis muralis, Podarcis gaigeae, Podarcis milensis, and Lacerta graeca) from the northeast Mediterranean region. Very long periods of postautotomy tail movement were demonstrated for all species (range=6-8 min), and differences among species were not statistically significant. Postautotomy tail movement, powered by anaerobic muscle activity, resulted in a strong increase in lactate concentrations and a concomitant depletion of muscle glycogen of exhausted tails relative to resting tails. No significant differences were found in either lactate or glycogen concentrations among the species examined. Duration of movement was negatively correlated with final lactate concentrations. The lack of differentiation in postautotomy energetic physiology in this group of species that have evolved under very different predation environments indicates that postautotomy muscle metabolism involves an overall conservative suite of characters.