The Lizard Log

The Langkilde Lab in Action


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Tree selection is linked to locomotor performance and associated noise production in fence lizards

New paper in Journal of Zoology featuring Kirsty, Tracy, and Nicole! The substrates on which animals spend their time can affect how they look, move, and sound. We found fence lizards more frequently on deciduous trees, on which they sprint faster and produce less noise relative to coniferous trees, which may affect their ability to catch prey or evade detection by predators. Noisiness and performance have received less attention in the context of substrate preferences than visual camouflage, but our results suggest they may also be important in determining the surfaces on which lizards prefer to be. (Check out the paper: Tree selection is linked to locomotor performance and associated noise production in a lizard.)

In the summer of 2016 I began a large-scale experiment investigating the effects of maternal stress on offspring characteristics in the eastern fence lizard, Sceloporus undulatus. My primary mission for the first part of that summer was simple: catch as many lizards as possible in the longleaf pine forests of southern Alabama. When you spend most of your day pursuing a small prey species, you quickly start to think like a predator. In what areas are you most likely to find them? At what times? On what surfaces? The cumulative years of experience of my fieldwork team (colleagues from the Langkilde lab) suggested that fence lizards were mostly found where there was a mix of hardwood deciduous trees and pines, and that they preferred the deciduous trees, like oaks and hickory, to the famous pines of the region. The longer I spent looking for lizards, the more I noticed that this observation held true. I didn’t put much thought into why until one day when I followed a lizard into a small stand of pine trees. I momentarily lost sight of the lizard, until I heard a loud scrabbling from a few metres away – there was the lizard, scuttling up a pine tree on the smooth, dry flakes of its bark. If the noise of the lizard’s claws moving on the pine bark alerted me so easily to its presence, I thought, perhaps the same was true for its real predators! Also – perhaps that noise was indication that this type of bark, with fewer crenulations and ridges on which to grip, was more difficult for the lizard to run on. Together, could these provide a reason that fence lizards seem to avoid pine trees despite their prevalence?

We decided to test this in the field. First, we quantified whether our anecdotal hunch that lizards prefer deciduous trees to conifers (pines) was really true by conducting thorough searches for fence lizards throughout our field sites, and noting the tree type we found them on, as well as the availability of trees in that area. This allowed us to test whether lizards were “choosing” deciduous trees in areas where they could also choose pines, as opposed to just being found in areas with only deciduous trees. As we expected, we found that even when availability of coniferous:deciduous trees was more or less 1:1, lizards were overwhelmingly found on deciduous trees, not pines.

Next, we tested our hypotheses that tree type changes how noisy lizards are when they move, and how quickly they are able to move. We did this by releasing wild lizards on either coniferous or deciduous trees, and then recording them as we stimulated them to run upwards on the tree by gently tickling their back legs. We then analysed these recordings and found, as we predicted, that the noise of lizards running (the sound level they produced when running compared to the background noise when they were still) was significantly higher when they were running on the smooth, flakier bark of coniferous trees. We also found that the sprint speed they attained on coniferous trees was lower than on deciduous trees. In other words: they are noisy and slow on pine bark compared to the bark of trees like oaks and hickorys.

Studies investigating where animals spend their time (either in terms of broader habitat preference, or more localised use of substrates) has often focused on coloration, and the camouflage it may or may not afford. Our study shows that other aspects of camouflage, such as acoustic camouflage, may also be important. It’s also important to consider how substrate affects performance, like sprinting speed: once you’re spotted by a predator, the speed at which you’re able to escape may be just as important as trying to remain hidden in the first place.

This was one of my favourite studies to be involved in, for a number of reasons! First, I love that we were able to find ways to test hypotheses based on a very simple natural history observation. Understanding the natural history of an organism is crucial for developing new ideas – and the “why does this happen?” questions are the bedrock of behavioural ecology. Second, this study was an opportunity to bring together friends and start new collaborations! Langkilde lab alum Nicole Freidenfelds brought her great knowledge and understanding of herpetofauna and natural history; local friends in Alabama helped me to identify tree species; I knew of Gavin’s prowess in acoustic analysis through Twitter, and asked him to help with this aspect of the project; and Tracy and I had a blast exploring these ideas with them!

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Maternal glucocorticoid effects across life stages in fence lizards – new paper!

“Anthropogenic disturbance is a growing threat, and the physiological consequences of exposure to such stressors is gaining increasing attention. A recent paper published in the Journal of Animal Ecology explores the consequences of stress-relevant hormones for mothers and their offspring…”

Read more in David’s new paper, and featured blogpost on the Journal of Animal Ecology blog!

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Sex-dependent effects of maternal stress in lizards

Check out the second chapter of Dustin’s thesis on the “Sex‐dependent effects of maternal stress: Stressed moms invest less in sons than daughters”!

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Multigenerational effects can have important and sex‐dependent effects on offspring. Sex allocation theory predicts that females should differentially invest in sons and daughters depending on sex‐specific fitness returns and costs of investment. Maternal stress‐relevant (glucocorticoid) hormones may be one mechanism driving this effect. We investigated how maternal stress hormones differentially affected sons and daughters by manipulating levels of the glucocorticoid, corticosterone (CORT), in gravid female eastern fence lizards (Sceloporus undulatus) and quantifying reproductive investment and sex ratio of resulting clutches, and the mass, snout‐vent length, and body condition of sons versus daughters at hatching. We found no effect of maternal CORT‐treatment on the number or size of eggs laid or on the sex ratio of resulting offspring, but sons of CORT‐treated mothers were shorter, lighter, and of poorer body condition at hatching than were sons of control mothers. We found no difference in size or condition of daughters with maternal treatment. Our results suggest that maternal stress, mediated by elevations in maternal CORT concentrations, can have sex‐specific effects on offspring manifesting as lower investment in sons.


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Undergraduate research in the spotlight!

The Langkilde lab was well-represented at this year’s Undergraduate Research Symposium by three of our lab researchers! It’s been an excellent year for undergraduate research. Congratulations to Richard Novak, Kristen Sprayberry, and Andrea Racic on their poster presentations! Not pictured is Jennifer Heppner, who also completed a brilliant thesis in the Langkilde lab this year. It’s been a pleasure having you all in the lab, and we will be sorry to see you go – but look forward to hearing about your future endeavours!

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A New Blood Sampling Method for Smaller Anurans that Preserves Critical Features of Specimens

Another new paper from Dustin In Herpetological Review! Summary below.

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Obtaining adequate blood samples is vital for most studies involving immunology or physiology. Anurans (frogs and toads) present a particular challenge for obtaining adequate samples, largely because of their relatively small size compared to other vertebrates.

Here, we propose a new method for obtaining large amounts of blood from the ventral abdominal vein of euthanized frogs, which we call the lethal abdominal vein of anurans (LAVA) technique.

We tested this method on the locally common Wood Frog (Lithobates sylvaticus –  pictured above). Using the LAVA technique, we were able to collect blood from 100% of frogs. Each frog yielded an average of 0.09 mL (range: 0.03 to 0.17 mL) of blood, which contained an average of 40 µL (range: 15 to 100 µL) of plasma.

We also found that neither size, ambient temperature, nor site affected our blood yields. We show that the LAVA technique is an easy-to-use method that yields high amounts of blood from anurans, and could be potentially viable in other small vertebrates.


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Offspring influenced by their evolutionary history more than their own experience in fence lizards

Check out the first chapter of Dustin’s thesis on the “Trans-generational but not early life exposure to stressors influences offspring morphology and survival”, recently published in Oecologia!
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Environmental changes, such as the introduction of non-native species, can impose novel selective pressures. This can result in changes in fitness-relevant traits within an individual’s lifetime or across multiple generations. We investigated the effects of early life versus trans-generational exposure to a predatory invasive insect stressor, the red imported fire ant (Solenopsis invicta), on the morphology and survival of the eastern fence lizard (Sceloporus undulatus). We captured gravid lizards from high-stress populations with long histories of invasion by fire ants and from uninvaded sites. Resulting hatchlings were exposed weekly to one of the three treatments until they reached maturity (42 weeks): (1) sub-lethal attack by fire ants; (2) topical application of the stress-relevant hormone, corticosterone (CORT), to mimic the stress of fire ant attack; or (3) control handling. Exposure to post-natal early life stress (fire ants or CORT) did not interact with a population’s evolutionary history of stress to affect morphology or survival and early life stress did not affect these fitness-relevant traits. However, morphology and survival were associated with the lizards’ evolutionary history of exposure to fire ants. Offspring of lizards from fire ant invaded sites had longer and faster growing hind-limbs, gained body length and lost condition more slowly in the first 16 weeks, and had lower in-lab survival to 42 weeks, compared to lizards from uninvaded sites. These results suggest that a population’s history of stress/invasion caused by fire ants during ca. 38 generations may be more important in driving survival-relevant traits than are the early life experiences of an organism.

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You can read more about Dustin’s research here and here! 


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Immunapalooza

Not all of the lab went south this summer; Kristen (one of our awesome lab undergrads) and I stayed at Penn State most of the summer, working on immune assays.  Kristen was the recipient of the Erickson Discovery Grant, and spent much of her summer on an independent research project, which involved measuring the effects of corticosterone (CORT) on the cell-mediated immunity (i.e. one way the body responds to a toxic or foreign substance) of eastern fence lizard females. She was also trying to determine if the lizards’ life history (whether they were from sites with or without fire ants) affected their immune function or interacted with the CORT treatment. Kristen just recently gave an excellent talk on her research at the Three Rivers Evolution Event (TREE) on Sept. 9th, where she was one of the only undergraduates to present a talk.

We also spent a lot of time this summer developing, improving, and validating several different immune assays for use in fence lizards, including ELISA assays for measuring anti-fire ant antibodies (IgY and IgM), complement function, natural antibodies, and the activity levels of heterophils (a type of immune cell that kills bacteria). Work on the assays for IgY, complement function, and natural antibodies is ongoing, but the IgM and heterophil activity assays are ready to be used.

The IgM ELISA assay was developed to work with as little as 10μl of plasma, and accurately detected anti-fire ant antibodies in a pool of plasma of lizards from Alabama, where the lizards are regularly exposed to fire ants. It did not detect any antibodies in a pool of plasma of lizards from Tennessee, at sites which have not yet been invaded by fire ants. The next step is to test the plasma of individual lizards from different sites, to see what proportion of lizards in various invaded sites have actually developed IgM antibodies to fire ants. Once the IgY assay is working, we should be able to better characterize the antibody response of the lizards to fire ants, and see if this helps them recover faster from fire ant stings.

IgM in the plasma of Alabama lizards

The higher the proportion of plasma from invaded (Alabama) lizards, the higher the signal from the IgM antibody.

Our heterophil activity assay is based off the assay described in Merchant, Williams, and Hardy (2009) for use in American alligators. To account for the much smaller blood volume of fence lizards, I altered the assay to work with 10μl of whole blood, and validated it in this species. This assay specifically tests for the presence of superoxide radicals, which are produced by heterophils as part of the oxidative burst used to kill bacteria and other organisms. When heterophils are more active (either because there are more heterophils or because the existing heterophils have been stimulated by something), the amount of superoxide in the blood increases. As part of the validation, we ran the assay with pools of blood treated with superoxide dismutase, which destroys superoxide, to test that the signal is actually caused by superoxide. We also ran blood with and without a stimulant of heterophil function, to determine if the signal reliably increases when heterophils are more active. The signal reliably decreases when inhibited by superoxide dismutase, and reliably increases when stimulant is added, indicating that this is a reliable test of heterophil function.

We also did a little bit of work optimizing the natural antibody test, increasing the sensitivity of the test so that it will work with less lizard plasma. And we also found a promising lead for testing alternative pathway complement function in fence lizards.

Aside from all the immunology work, we also got out into the field up here in Pennsylvania a little bit, although we didn’t find many lizards. All in all, it was a fun, productive summer.