Effects of temperature on plasma corticosterone in a native lizard

The glucocorticoid stress response is frequently used to indicate vertebrate response to the environment. Body temperature may affect glucocorticoid concentrations, particularly in ectotherms. We conducted lab manipulations and field measurements to test the effects of body temperature on plasma corticosterone (predominant glucocorticoid in reptiles) in eastern fence lizards (Sceloporus undulatus). First, we acclimated lizards to one of 4 treatments: 22 °C, 29 °C, 33 °C, or 36 °C, and measured cloacal temperatures and plasma corticosterone concentrations at baseline and after exposure to a standardized stressor (cloth bag). Both baseline and stress-induced corticosterone concentrations were lower in lizards with lower body temperatures. Second, we acclimated lizards to 22 °C or 29 °C and exposed them to a standardized (cloth bag) stressor for 3 to 41 min. Lizards acclimated to 29 °C showed a robust increase in plasma corticosterone concentrations with restraint stress, but those at 22 °C showed no such increases in corticosterone concentrations. Third, we measured lizards upon capture from the field. There was no correlation between body temperature and baseline plasma corticosterone in field-caught lizards. These results suggest body temperature can significantly affect plasma corticosterone concentrations in reptiles, which may be of particular concern for experiments conducted under laboratory conditions but may not translate to the field.

A chromosome-level genome assembly for the Eastern Fence Lizard (Sceloporus undulatus), a reptile model for physiological and evolutionary ecology

High-quality genomic resources facilitate population-level and species-level comparisons to answer questions about behavioral ecology, morphological and physiological adaptations, as well as the evolution of genomic architecture. Squamate reptiles (lizards and snakes) are particularly diverse in characteristics that have intrigued evolutionary biologists, but high-quality genomic resources for squamates are relatively sparse. Lizards in the genus Sceloporus have a long history as important ecological, evolutionary, and physiological models, making them a valuable target for the development of genomic resources. We present a high-quality chromosome-level reference genome assembly, SceUnd1.0, (utilizing 10X Genomics Chromium, HiC, and PacBio data) and tissue/developmental stage transcriptomes for the Eastern Fence Lizard, Sceloporus undulatus. We performed synteny analysis with other available squamate chromosome-level assemblies to identify broad patterns of chromosome evolution including the fusion of micro- and macrochromosomes in S. undulatus. Using this new S. undulatus genome assembly we conducted reference-based assemblies for 34 other Sceloporus species to improve draft nuclear genomes assemblies from 1% coverage to 43% coverage on average. Across these species, typically >90% of reads mapped for species within 20 million years divergence from S. undulatus, this dropped to 75% reads mapped for species at 35 million years divergence. Finally we use RNAseq and whole genome resequencing data to compare the three assemblies as references, each representing an increased level of sequencing, cost and assembly efforts: Supernova Assembly with data from10X Genomics Chromium library; HiRise Assembly that added data from HiC library; and PBJelly Assembly that added data from PacBio sequencing. We found that the Supernova Assembly contained the full genome and was a suitable reference for RNAseq, but the chromosome-level scaffolds provided by the addition of the HiC data allowed the reference to be used for other whole genome analysis, including synteny and whole genome association mapping analyses. The addition of PacBio data provided negligible gains. Overall, these new genomic resources provide valuable tools for advanced molecular analysis of an organism that has become a model in physiology and evolutionary ecology.

Testosterone Reduces Growth and Hepatic IGF-1 mRNA in a Female-Larger Lizard, Sceloporus undulatus: Evidence of an Evolutionary Reversal in Growth Regulation

Synopsis Previous research has demonstrated that testosterone (T) can inhibit growth in female-larger species and stimulate growth in male-larger species, but the underlying mechanisms of this regulatory bipotentiality have not been investigated. In this study, we investigated the effects of T on the expression of hepatic insulin-like growth factor-1 (IGF-1) mRNA and circulating IGF-1 hormone in Sceloporus undulatus, a species of lizard in which females grow faster to become larger than males and in which T inhibits growth. Experiments were performed in captivity on mature female and male adults in the asymptotic phase of their growth curve and on actively growing, pre-reproductive juveniles. In adult males, the expression of hepatic IGF-1 mRNA increased following surgical castration and returned to control levels with T replacement; in intact adult females, exogenous T had no effect on IGF-1 mRNA expression. In juveniles, T significantly reduced both growth and the expression of hepatic IGF-1 mRNA to similar extents in intact females and in castrated males. The relative inhibitory effects of T on mRNA expression were greater in juveniles than in adults. Plasma IGF-1 hormone was about four times higher in juveniles than in adults, but T had no significant effect on IGF-1 hormone in either sex or in either age group. Our finding of inhibition of the expression of hepatic IGF-1 mRNA stands in contrast to the stimulatory effects of T in the published body of literature. We attribute our novel finding to our use of a species in which T inhibits rather than stimulates growth. Our findings begin to explain how T has the regulatory bipotentiality to be stimulatory in some species and inhibitory in others, requiring only an evolutionary reversal in the molecular regulation of growth-regulatory genes including IGF-1. Further comparative transcriptomic studies will be required to fully resolve the molecular mechanism of growth inhibition.

History of predator exposure affects cell-mediated immunity in female eastern fence lizards, Sceloporus undulatus (Squamata: Phrynosomatidae)

On exposure to stressors, energy is diverted from non-urgent functions towards those important for immediate survival. The degree and nature of reallocation may be affected by the evolutionary history of the animal. The eastern fence lizard (Sceloporus undulatus) coexists in parts of its range with invasive fire ants (Solenopsis invicta), which attack and wound lizards and elevate stress-relevant hormones (corticosterone), whereas other populations have never been exposed to fire ants. We examined how a history of fire ant invasion affected the immune response in female lizards after exposure to exogenous corticosterone (mimicking exposure to a stressor) during gestation (dosing regimens differed among corticosterone-exposed lizards owing to the constraints of the original studies, but we found no evidence that this affected the outcome of the present study). A history of exposure to predatory stressors (fire ants) and corticosterone treatment affected cell-mediated immunity. Lizards from fire ant-invaded sites had a reduced immune response compared with those from uninvaded sites. Corticosterone treatment had no effect on the immune response of lizards from invaded sites but reduced the immune response of lizards from uninvaded sites. This suggests that an evolutionary history of exposure to wounding alters the immune response to corticosterone. Future work on how the immune system responds to environmental threats will be informative for the prediction and management of these threats.

Contrasting Responses of Lizards to Divergent Ecological Stressors Across Biological Levels of Organization

It is frequently hypothesized that animals employ a generalized “stress response,” largely mediated by glucocorticoid (GC) hormones, such as corticosterone, to combat challenging environmental conditions. Under this hypothesis, diverse stressors are predicted to have concordant effects across biological levels of an organism. We tested the generalized stress response hypothesis in two complementary experiments with juvenile and adult male Eastern fence lizards (Sceloporus undulatus). In both experiments, animals were exposed to diverse, ecologically-relevant, acute stressors (high temperature or red imported fire ants, Solenopsis invicta) and we examined their responses at three biological levels: behavioral; physiological (endocrine [plasma corticosterone and blood glucose concentrations] and innate immunity [complement and natural antibodies]); and cellular responses (gene expression of a panel of five heat-shock proteins in blood and liver) at 30 or 90 min post stress initiation. In both experiments, we observed large differences in the cellular response to the two stressors, which contrasts the similar behavioral and endocrine responses. In the adult experiment for which we had innate immune data, the stressors affected immune function independently, and they were correlated with CORT in opposing directions. Taken together, these results challenge the concept of a generalized stress response. Rather, the stress response was context specific, especially at the cellular level. Such context-specificity might explain why attempts to link GC hormones with life history and fitness have proved difficult. Our results emphasize the need for indicators at multiple biological levels and whole-organism examinations of stress.

Eastern Fence Lizards (Sceloporus undulatus) display an ontogenetic shift in relative consumption of native and invasive prey

Interactions between invasive prey and native predators can provide an opportunity to better understand predator–prey dynamics and how these may change through ontogeny. Eastern Fence Lizards (Sceloporus undulatus (Bosc and Daudin in Sonnini and Latreille, 1801)) are ant specialist, particularly as juveniles. Invasive red imported fire ants (Solenopsis invicta Buren, 1972) pose a lethal risk to S. undulatus that eat them, especially smaller-bodied juveniles. We examine ontogenetic shifts in S. undulatus consumption of toxic invasive fire ants versus palatable native pyramid ants (Dorymyrmex bureni (Trager, 1988)). We predicted that hatchlings should avoid eating fire ants in favor of native ants, whereas less-vulnerable adults should take advantage of both prey sources. However, when given the choice between fire ants and native ants, hatchlings consumed similar numbers of these species, whereas adults consumed nearly three times as many native ants as invasive fire ants. Increased consumption of fire ants in adulthood could be the result of lifetime experience, strategies to safely consume fire ants, ontogenetic shifts in the ability to distinguish between ants, or reduced costs to adults of eating venomous ants. Future research should aim to distinguish these alternative mechanisms and examine the long-term consequences of native species incorporating toxic invasive prey into their diets.