Anole Annals

In his Masters thesis conducted in Simon Lailvaux’s lab at the University of New Orleans and presented this week at Evolution 2017, David Weber used a multiyear data set of Anolis carolinensis lizards’ locations and morphology as well as a DNA-based pedigree to investigate the effects of body size and relatedness on the spatial distribution of these lizards. Specifically, he set out to test three hypotheses: first, are males’ home ranges larger than females’ home ranges? Second, are bigger males more likely to be surrounded by smaller males that are related to them? And third, is there any evidence for the inheritance of home ranges from parent to offspring?

Anolis carolinensis dewlapping. Photo by Cowenby available on Wikipedia.

Lizard locations were sampled in an urban New Orleans park twice a year, in the fall and in the spring, from 2010 to 2015. The dataset included over 800 individuals, and what struck me most about these data was that, of these 800+ individuals, fewer than 100 were observed often enough to estimate home range volumes–death and dispersal can rule these lizards’ lives! Male and female home range volumes did not differ significantly (and the trend was in the direction of females moving over larger areas, which concurs with data from Robert Gordon’s 1956 thesis on green anoles, but with little else, I think). Curiously, smaller neighbours of the biggest males were less related to them than were males found farther away, suggesting that male anoles don’t preferentially tolerate their kin over non-kin. And though philopatry  (aka site fidelity aka staying the same place) was rare overall, females were a bit more likely to co-occur with their male offspring than males were. In a result that conforms to traditional wisdom, Weber found that the biggest males in the site seemed to avoid each other, potentially spacing themselves as far apart as possible.

Following a kind shout-out to my and Jonathan Losos’ recent paper on Anolis territoriality or the lack thereof, Weber chose to interpret his results as making sense only outside of a territorial framework. Unsurprisingly, I concur with this decision entirely, and am excited to see where Weber goes with this idea in the publications resulting from this mammoth dataset!

The arrival of an outsider that overlaps in resource use and habitat with local species can lead to intense competition between the two. A result of this competition can be character displacement, where traits of the species (one or both) change in sympatric populations (where the co-occur), but not in allopatric populations. Claire Dufour (Post-Doctoral researcher at Harvard University) presented her work on character displacement for two anole species on the island of  Dominica: the native Anolis oculatus and the introduced Anolis cristatellus. Her objective was to integrate ecological, antagonistic and reproductive character displacement. Specifically, she tested whether competition  between these new island-mates leads to changes in habitat use, morphology, and display behavior.

Location of populations of the introduced A. cristatellus with the sampled area, Calibishie inset

Claire compared allopatric populations of the two species with sympatric populations in the northern area of the island in Calibishie, where Anolis cristatellus has been present for two years. She found that in sympatry, both morphological and behavioral shifts have occurred. In sympatry, Anolis oculatus perched higher and had shorter limbs. She also found differences in display behavior, which she tested with an anole robot programmed to dewlap and do push-ups. This experiment showed that in sympatry, Anolis cristatellus dewlapped less, but Anolis oculatus does not alter its display behavior. Future work will test for reproductive character displacement and contrast populations where Anolis cristatellus has been present for a longer time span.

The Sensory Drive hypothesis predicts that species will evolve communication signals that are effective in the particular light environment in which they occur. Anolis lizards are an excellent example: in dark habitats, they tend to have light-colored, highly reflective (and transmissive) dewlaps that are usually yellow or white in color, whereas in bright, open environs, dewlaps tend toward blue, black, orange or red. However, demonstrating that these dewlaps are actually effective at being visible in their particular habitats has proven surprisingly challenging.

Leo Fleishman has been a leader in this area and in a talk at the sensory ecology symposium at the evolution meetings, he presented new and exciting developments. First, in line with previous work, he showed that the spectral reflectance/transmittance of dewlaps is not particularly well-matched to that of the background. Rather, the same colored dewlaps appear to be maximally contrasting with the radiance of the background across all habitats:  basically all habitats have mostly green backgrounds, and red or orange stands out the best against the green background, no matter what the habitat.  So much for sensory drive, it would seem!

But more recent work saves the day: it turns out that habitats differ in the total intensity of light (number of photons coming down) they receive and that, furthermore, across species, dewlap intensity (total photons reflected and/or transmitted) is negatively related to habitat intensity (with one notable outlier, the enigmatic A. gundlachi). Under the relatively low light conditions of forest shade or partial shade, color discrimination becomes more difficult, and colors such as red and orange and other dark colors do not stand out well against the background, because they simply do not emit enough photons to efficiently drive color vision.  Yellow or white works better. Conversely, in intense light environments, there is enough light to easily see the darker colors, and these stand out well against the green background. Moreover, behavioral experiments confirm that in bright light conditions red stimuli are most visible against a green background, whereas in low light yellow stimuli are more visible.  Thus, even though most Anolis habitats have similar spectral properties, differences in total light intensity strongly influence what colors are most effective, and thus appear to have played a major role in the shaping the evolution of dewlap colors.

Leo Fleishman discusses color space in four dimensions, corresponding to the four cones in the anole eye. For each species, red dots are color of the dewlap and green dots are the color of the background, indicating that dewlaps stand out against their background.

Bright and early on the last day of the annual Evolution meeting, James Stroud (Florida International University) presented his work on character displacement in novel communities of introduced anoles in Miami. In this elegant use of a natural experiment, James looked at the novel co-existence of two anoles in their introduced range and wondered if character displacement was occurring as predicted when two ecologically similar species are found in sympatry. Specifically, James wanted to know if Anolis cristatellus and Anolis sagrei would shift their habitat use when in sympatry, resulting in correlated shifts in morphology. These species are both trunk-ground anoles of roughly the same body size. They are native to Cuba/Bahamas and Puerto Rico (respectively) and are diverged by ~50 million years.

James hypothesized that in their introduced range in Florida, these two species would diverge ecologically in sympatry but be more similar in allopatry. He found that in allopatry, both species attained similar relative abundances and perched at similar heights. However, in sympatry, both decline in relative abundance suggesting that these species are interacting strongly with one another. Even more interesting, in sympatry A. sagrei perches lower and spends more time on the ground than it does in allopatry, while A. cristatellus perches higher!

Next James hypothesized that these ecological shifts could lead to shifts in morphology. If A. sagrei is spending more time on the ground, perhaps longer limbs would be favored. Similarly, if A. cristatellus is spending more time higher up in the trees, perhaps there would be selection for stickier toepads. As predicted, A. sagrei had longer forelimbs and hindlimbs in sympatry. However, he did not find any difference in toepad morphology between sympatric and allopatric populations of A. cristatellus. Instead, he observed that A. cristatellus in sympatry with A. sagrei had significantly smaller heads.

James ended by wondering if alternative behavioral and social mechanisms may drive these observed shifts in head morphology. Either way, this case study provides an interesting insight into how a complex range of adaptive responses can result from a seemingly simple ecological interaction.

Cities and urban areas are expanding rapidly around the world, altering the environment and creating very different ecological and selective pressures for organisms that live in urban habitats. A few of the most striking differences between urban and natural habitats are higher temperatures and a huge increase in artificial substrates like the walls of buildings. These artificial substrates (e.g., metal, concrete) are not only significantly smoother than natural (i.e., trees) substrates, but also absorb, retain, and radiate heat differently. Consequently, organisms may alter their behavior to better deal with these and other challenges of city life. Since anoles cannot internally regulate their temperature, behavioral shifts may be driven by perch substrate properties, temperature, or some interaction of the two.

Kevin Aviles-Rodriguez (U. Mass. Boston) addressed this question in urban Anolis cristatellus in San Juan, Puerto Rico. He created experimental enclosures in which each wall was a different substrate: wood, plastic, painted cement, and metal. He placed individual lizards into the enclosures and observed which wall they were perched on throughout the day. He also recorded the temperature of each wall, to determine how perch temperature of each substrate type influenced perch choice. Aviles-Rodriguez conducted this experiment in both urban and forest populations, and predicted that urban lizards would use artificial substrates more readily than forest lizards.

Interestingly, he did not find that to be the case – lizards from both urban and forest habitats used bark much more than any other surface. However, when lizards did use artificial substrates, they tended to use metal and cement when these perches were cooler, suggesting that perch temperature is a factor in perch choice. Aviles-Rodriguez plans to test these hypotheses more thoroughly by conducting additional experiments across more urban replicates to see if the same pattern emerges. He also plans to experimentally control the temperatures of different perch substrates in his enclosures to see whether lizard choices are primarily driven by perch substrate or temperature.

I find Transposable Elements (TEs) to be some of the most fascinating features of genomes. Also known as selfish genetic elements, these sequences contain the genetic machinery to create copies of themselves and insert these new copies in locations throughout the genome. The genomes of different organisms vary widely in their degree TE abundance. For example, 20% of the human genome is composed of just one kind of TE!

This morning Robert Ruggiero, a Postdoctoral Fellow in the lab of Stephane Boissinot at NYU Abu Dhabi, presented his work on the population genomics of TEs in the genomes of Anolis carolinensis populations. Robert employed a clever approach that uses a feature of next-generation sequence data to identify TE insertions. In this way, he can characterize all of the TE insertions in an individual’s genome and determine what portion of a population contains any particular insertion.

It’s easy to see how Transposable Elements could be bad for an organism. If a TE inserts itself into the middle of an important gene, the function of that gene could be interrupted, and render the bearer of that insertion less evolutionarily fit. The ability of natural selection to purge this type of deleterious insertion is governed in part by the effective population size of the group where that insertion arises. In essence, natural selection is more effective in larger populations.

Using the information he collected on TE insertions in anole populations, Ruggiero created a population genetic summary called an Allele Frequency Spectrum, the count of insertions that exist at a particular frequency in a population. This distribution can then be used to infer how well populations control the frequency of TE insertions, and in addition, estimate the effective size of those populations. Robert found TE insertions in Floridian populations of Anolis carolinensis were maintained at lower frequencies than other populations suggesting that selection is better able to purge deleterious insertions in the Florida population. He also found that different families of TEs appear to employ strategies that mirror ecological r/K theory. Some TEs create insertions frequently but few of these insertions get to high frequency, whereas other TEs insert infrequently, but those insertions that do occur are more likely to reach high frequency. Moving forward, using this line of inquiry in anoles will be an excellent opportunity to understand the control and evolutionary consequences of TEs, particularly as more Anolis genomes come online allowing comparative analyses.

Anolis distichus is well-known in the anole world for the high degree of ecomorphological variation within the species, especially in dewlap color. In fact, there are 18 described subspecies! While there is some gene flow between various subspecies and populations, the phenotypic differences are maintained, which suggests strong selection. But the fine-scale genetic structure underlying these traits is not well understood. Anthony Geneva and colleagues decided to explore the genomic basis of adaptive divergence in a well-described hybrid zone between two A. distichus subspecies. The first, A. d. ignigularus, has a white dewlap, and occupies a dry forest habitat while the second, A. d. ravitergum, has a red dewlap and inhabits a wetter habitat. The two subspecies occur along a transect from dry to wet, and they hybridize in a narrow contact zone in the middle. These two subspecies provide a great system to explore the link between adaptive and genetic divergence.

Anolis distichus. Photo by Rich Glor

Geneva sequenced individuals using RNASeq across an environmental transect from wet to dry, including allopatric and sympatric populations of both species. He examined levels of divergence and introgression to explore which genomic loci might be the basis for the ecological adaptive divergence between these two species. He found a suite of candidate genes that differ between the two subspecies, as well as several that show signs of introgression between the two. Interestingly, several of the divergent genes are involved in two traits that likely are impacted the environment – insulin signaling, which may relate to metabolic differences between hot and cool climates, and vision, which may relate to differences in light availability and signal efficiency. Most of the introgressed genes, on the other hand, relate to conserved pathways, suggesting that these genes play similar roles in both subspecies.

Adpative divergence in anoles has been a topic of interest for a long time, and Geneva’s study provides and a valuable insight to the genetic basis of this interesting phenomenon.

On each of the Greater Antillean islands, habitat-specialist Anolis ecomorphs have independently evolved complex suites of shared phenotypes and behaviors. This remarkable convergence has motivated the work of generations of anolologists. With anoles entering the once-exclusive club of genome-enabled organisms, a new line of investigation has become possible: Is the convergence observed in anole ecomorphs caused by molecular convergence? Such convergence can take many forms, including shared changed at individuals sites, or shared changes in the rates of protein evolution of individual genes.

Russ Corbett-Detig of UCSC sought to answer this question using whole-genome sequence data from 12 species – four from each of the Trunk-Ground, Trunk-Crown, and Grass-Bush ecomorphs drawn from different islands and different evolutionary lineages. Accurately detecting molecular convergence is fraught and much recent research has focused on avoiding pitfalls that could lead to a positively misleading inference of convergence where none actually exists. Previous studies have trumpeted amazing cases of molecular convergence in a variety of animals, only to be later shown to be artifacts of data analysis.

Corbett-Detig did everything right. He used null models that account for the expected background levels of convergence caused by processes other than natural selection. He found no evidence of extra shared non-synonymous mutations in any of the three ecomorph groups. Similarly, he found no signal of shared changed in protein evolution in Trunk-Ground or Trunk-Crown but Grass-Bush anoles seemed to share elevated rates of changes in many genes. This result was exciting, but Corbett-Detig dug deeper and discovered a new way this type of analysis could be mislead – two of the four Grass-Bush anoles exhibited accelerated evolution across their entire genomes and, as a result, seemed to share faster rates at more genes than expected by chance. When Corbett-Detig corrected for this bias, the signal of convergence disappeared.

While this result was in one sense disappointing, it is also fascinating and suggests the evolutionary pathways to shared ecomorphological traits are numerous and strongly influenced by contingency. Furthermore, anole ecomorphs have evolved such a stunning set of similarities that other forms of convergence like genome structure, gene family expansion, or convergence in gene regulation may still hold the key to understanding the genetic basis the remarkable convergence of Anolis ecomorph classes.

Sexually antagonistic selection occurs when traits are beneficial for one sex, but detrimental to the other. This commonly occurs in species with sexual dimorphism, such that one trait is positively correlated with fitness in one sex, and negatively correlated with fitness in another. But in many organisms, the sexes do not become dimorphic until maturity – that is to say, juveniles all look pretty much alike, even when adults show clear differences between males and females. Which leads to the question: how does sexually antagonistic selection change over an organisms’ lifespan? Research from studies of Drosophila flies suggests that this is the case, but the question hasn’t been well-studied in vertebrates.

Everyone’s favorite anole, Anolis sagrei

Until now. In his Evolution talk, Aaron Reedy (University of Virginia) described his work testing whether sexually antagonistic selection changes over ontogeny using our favorite workhorse of evolutionary ecology, the brown anole (A. sagrei). Anolis sagrei are sexually dimorphic, with adult male body sizes up to 30% larger than females, but juveniles are monomorphic. Reedy and colleagues  sampled A. sagrei on several small islands in a Florida watershed four times a year, capturing thousands of adults and juveniles. They measured the body size of all lizards captured, and combined this morphological data with survivorship data to determine how selection was acting on body size in adults and juveniles.

They predicted that juvenile males and females would experience concordant selection, while adult males and females would experience antagonistic selection. And this is exactly what they found: for juveniles, body size was correlated with survival in the same way between sexes. But in adults, this was not the case. In the first year of sampling, there was no selection on body size for adult females, but positive selection for males, such that bigger males survived better. Interestingly, during the second year of sampling, the relationship flipped – females experienced positive selection on body size, and males experienced negative selection. The reasons for this shift are uncertain, but the main point is clear – sexually antagonistic selection does indeed change over ontogeny. Reedy et al. are planning to follow up this great new research by expanding their study to look at more islands and more traits to get at the finer points of these selective differences, so stay tuned!

Oriol Lapiedra opened up the penultimate day of Evolution by discussing his results of a recent field experiment in the Bahamas. In this project, Lapiedra and colleagues evaluated how inter-individual variation in behavior – specifically risk-taking – influenced survival. To do this, the research team took advantage of a well-understood model system in evolutionary ecology: brown anoles (Anolis sagrei) on islands with and without anole-predators (curly-tailed lizards; Leiocephalus carinatus) in the Bahamas. Male and female brown anoles were collected and subjected to a behavioural trial which measured the amount of time it took for a lizard to leave a refuge after being exposed to a predator. These observations were used to quantify each individual’s propensity to take risks. For example, those individuals that left their refuge shortly after seeing a predator were interpreted as being more ‘risky’ than more conservative individuals. Following these trials, each lizard was x-rayed to assess morphology and individually tagged, before being released onto one of 4 predator-free islands or 4 predator-present islands, all of which were currently void of anoles.

Lapiedra et al. started with a priori hypotheses that overall survival would be lower on those islands with predators, and those that did survive would be individuals considered less risky. After waiting 4 months, the research team returned to the Bahamas to collect all lizards from each island and see which individuals had survived. The authors report that, as expected, overall survival was lower on islands with predators, and that there was a significant relationship between behaviour and survival such that high risk-taking individuals had much lower survival when predators were present. This suggests that under those biotic conditions, natural selection operates against those riskier phenotypes. On closer inspection, this relationship was largely driven by a strong relationship in females, with no significant relationship existing between risk-taking behavior and survival of males.

Lapiedra et al. then contrasted these results by independently assessing how morphology was related to survival. The authors found that both risk-taking behavior and morphology influenced survival, however – and, important to this study – the relative effect of an individual’s risk-taking behaviour was much more influential on survival.