Noisy invasive treefrogs

I’m Jenny Tennessen, a doctoral candidate in the Langkilde lab.  This week we’re going to leave the world of reptiles behind, to focus on a different order; one that includes animals often characterized by their large eyes, moist skin, and brilliant voices.  That’s right, anurans – frogs and toads.  Much of my research in the Langkilde lab focuses on how these inquisitive-looking animals respond physiologically and behaviorally to their surrounding acoustic environments.  In this week’s blog I’d like to tell you about an exciting project in which I got to chase around treefrogs in the southeast to study their acoustic behavior.  You can call me the Treefrog Whisperer.

Despite their name, not all treefrogs hang out in trees.  They do, however, spend a large portion of their lives elevated on various climbing perches – a behavior facilitated by their relatively small size and broadened toe pads equipped with discs that aid in climbing and grasping.  In fact, this body plan is so effective for a life of climbing, that different evolutionary trajectories of treefrogs have resulted in similar body plans through convergent evolution!

During the breeding season, treefrogs come down from their aerial perches to breed in water bodies, which can be anything from large lakes with fish to flooded roadside ditches following rainstorms.  Have you ever gone for a walk on a summer evening in Florida just after a deluge of rain, and heard sounds so loud you thought you might lose your hearing?  If so, the likely culprits included treefrogs, whose post-rain acoustic nuptials can be nearly deafening at the peak of breeding season.

A male and female green treefrog (Hyla cinerea) in amplexus.

Something interesting is going on in Florida right now – a treefrog war, of sorts, is being waged, though only one side is fighting the war because the other side is mostly naïve to any problems.  Cuban treefrogs (Osteopilus septentrionalis), an invasive species native to Cuba and parts of the Caribbean, have been spreading northward throughout Florida since the 1930s or so, having reached the mainland likely as stowaways on ships for the nursery trade.  These large treefrogs have gigantic toepads that make them supremely equipped to cling to things – including cars and RVs – and they have hitchhiked all over the state.  These unfriendly giants have voracious appetites, and devour unsuspecting native frogs that cross their paths or climb into the same shelters.  Because this behavior is unique to the native treefrog community in Florida, it has been very effective.  Additionally, their toxic skin secretions help keep them safe from would-be predators.  For all of these reasons, Cuban treefrog populations have exploded throughout Florida – in fact, it seems that one of the only natural factors able to limit their populations is cold temperatures during the winter season.

Why did the Cuban treefrog cross the road?  To advance the invasion tide! ….. (c’mon, I had to try!)

After learning about the Cuban treefrog invasion in Florida, I had an idea: since treefrogs use sound to attract mates and defend territories, maybe Cuban treefrog calls are interfering with native treefrogs’ acoustic behavior.  There have recently been many discoveries about ways that animals change their acoustic behavior in noisy environments – for example, city birds sing at higher frequencies, and humpback whales lengthen their songs in response to sonar – but few studies have considered the sounds of an invasive species to be a source of noise disturbance.

To test our hypothesis that Cuban treefrog choruses interfere (cause acoustic competition) with native treefrog communication, we selected two treefrog species native to Florida: the green treefrog (Hyla cinerea), whose breeding call is very similar to the Cuban treefrog’s, and the pine woods treefrog (Hyla femoralis), which calls at a faster rate and a higher frequency. Click here to see a video of a calling green treefrog and here for a video of a calling pine woods treefrog.

A “spectrogram” comparing the breeding calls of Cuban, green, and pine woods treefrogs. Greater acoustic intensity is illustrated by lighter coloration. The orange box that continues across the graph illustrates the frequency range of greatest energy for Cuban treefrogs and the portion of green and pine woods treefrog breeding calls with which it overlaps. The individual red boxes illustrate the frequency ranges of greatest energy for green and pine woods treefrogs. It is clear that Cuban and green treefrog breeding calls are similar in rate an frequency, whereas pine woods treefrog breeding calls occur at a much greater rate with most energy at a higher frequency.

We approached these treefrogs as they were calling from perches on grasses above the water, and played recordings of Cuban treefrog choruses (or control sounds within or above the frequency range of the Cuban treefrog chorus).  We made acoustic recordings of their pre-, during- and post-playback acoustic behavior.  If our hypothesis was correct, we predicted we would see that species with more similar acoustic behavior (green treefrogs) would change their calls, whereas those with less similar acoustic behavior (pine woods treefrogs) would not.

The setup for our acoustic playback experiment. Lots of expensive acoustic equipment suspended over the water! Fortunately nothing went for a swim.

 

We are still in the process of analyzing the data but I’ll share one of our most interesting discoveries so far.  In response to Cuban treefrog playback, green treefrogs increased their call rate by up to double the original rate, while pine woods treefrog call rate did not change!  These results are consistent with our predictions, and lend support to our hypothesis that sounds from invasive species can interfere with native species’ communication.  We are still exploring other acoustic parameters that green and/or pine woods treefrogs may modifiy to avoid interference, including call frequency, duration, and intensity.  To learn more about this work, check out my radio interview on Quirks & Quarks, or feel free to email me (jbt148@psu.edu).

 

Redemption in the Deep South

This undergraduate guest blog post has been dispatched by Mark Herr, a junior from Los Angeles who is majoring in Wildlife and Fisheries Science at Penn State. Mark is a member of the Penn State Presidential Leadership Academy, and is interested in continuing on to graduate school to study behavior, ecology, and, of course, reptiles & amphibians:

This past spring I applied for, and with the help of Dr. Langkilde received, a Penn State Discovery Summer Grant to conduct independent research. Initially, I planned to conduct two projects on similar systems. Unfortunately, only the second project (hence the title of this post) resulted in a successful field season. Which isn’t to say that the first project wasn’t valuable – it was perhaps the most valuable research experience I’ve had yet – but its value really lay in lessons learned rather than any publishable results!

Before I explain the project, I want to first give a shout-out to Drs. Langkilde and Sean Graham. Dr. Langkilde managed to let me know that my initially proposed projects were all rather grandiose and unfeasible, while convincing me to keep on firing away with ideas. Eventually, I sent her an idea that we both agreed was novel, and more importantly, possible. Sean went out in the field with me during every day of sampling and made the project possible. This is all the more impressive because he is a post-doc researcher and I am just an undergraduate – which says much both about him and the atmosphere that Dr. Langkilde fosters in her lab.

Dr. Sean Graham does his best to become an internet sensation by holding a cottonmouth safely out of striking range while using the standard forced perspective method so that it appears to be of monstrous size. However, this snake is actually only a little over 3 ft. long. The camera really does add a few pounds.

I don’t want to delve into the specifics of my project idea in California, both because it will bring back the stinging memory of defeat (just joking!) and poison oak rashes so bad that they probably warranted hospital visits – both for Sean and I! Luckily, Sean and Dr. Langkilde had foreseen the fact that all might not go as planned with one of the projects – and so they had suggested that we conduct a second project as insurance. And so it was this “insurance policy” that became my last true hope for some publishable research during the summer break.

My project ran side by side with other work being conducted in Alabama by the Langkilde Lab, which allowed me to help out with other projects when I wasn’t out in the field with Sean collecting snake blood. Yes. Snake blood, and no, this didn’t involve door-to-door salesmen selling cure-alls or primitive rituals by witch doctors.

Sean and I investigated how the stress hormone (corticosterone) concentrations in Cottonmouths (Agkistrodon piscivorus) were related to their anti-predator behavior. Cottonmouths are large bodied, aquatic pit vipers native to the southeastern US. Most importantly, they are abundant. The snakes were honestly even more abundant than I had expected – even though we did have a period where we had trouble finding any, but more on that later.

They are also famous for being aggressive, or, at least that is what the man on the street will say – the consensus among scientists and in peer-reviewed research is that they aren’t anything close to the bloodthirsty mankillers they are made out to be. Cottonmouths do, however, have an extensive suite of anti-predator behaviors. They vibrate their tails (even though they have no rattle), they hiss, they can strike, and they can open their mouth wide in what is called a ‘gape’ in order to convince a possible predator that they aren’t worth the trouble. Actually, this gaping behavior is what gives them their common name – the inside of their mouths has a white lining that is highly visible (especially in their often dark habitats).

A Cottonmouth from the Everglades showing the display for which it is named.

The fact that this species has so many different anti-predator behaviors means that I was able to formulate a point system that would rank each individual snake based on how “defensive” it was acting. The procedure was to approach the snake and stand in close (but safe) proximity to it for 15 seconds and then grasp it mid-body with a set of snake tongs for another 15 seconds – all the while taking note of every behavior that the snake exhibited. We would then use a snake tube to restrain the animal and take a blood sample.

As I write this I am laughing out loud at how simple that last sentence makes the blood drawing seem.

The problem, as you might imagine, is that often times (read: every time) the already defensive snake wants nothing more than to avoid slithering up a tight clear plastic tube so that we might get our blood sample safely. For obvious safety reasons, the entire tubing maneuver has to be completed using nothing but a set of snake tongs and a tremendous amount of patience – tubing is the safest way to handle venomous snakes – by far (both for the snakes and the researcher). It’s the industry standard technique.

The process goes something like this:

1. Using a pair of snake tongs, the snake is grasped firmly enough at the midbody to prevent the snake from escaping but not so tightly as to injure the animal.
2. The tube is maneuvered such that it fits over the snake’s head, and ideally the snake will crawl up the tube such that its body is half inside and half outside the tube, with the snake’s posterior body portion and tail hanging out of the end.
3. Swiftly and steadily, the snake is grasped at midbody at the exact point where it hangs out of the tube – with the hand holding the snake firmly grasping both the animal itself and the tube to prevent the snake from either moving further forward or backing out.

Step #2 is sometimes easy, as the snake will cooperate and crawl up the tube as soon as it is able. Usually, though, it is not so easy. More often it would rather keep dodging and moving its head away. Even more often it would rather just strike the tube repeatedly. This process was further complicated by the fact that we were on a tight time schedule –we needed to obtain blood from the snake before its hormone levels had risen significantly (which takes only a few minutes). Did I mention the fact that we were unable to physically touch the snakes until they were in the tube?

A safely and successfully tubed Cottonmouth on display after obtaining a blood sample.

We went out during the day and at night searching for snakes, usually spending at least 4-5 hours at a stretch slogging through swamps in waist deep swamp water, and we managed to get to within about 5 snakes of our sample size when we hit a wall. No. More. Snakes.  I can’t even remember how many times Sean and I ventured forth without finding even a single snake. It was as though cottonmouths had gone from being the most common snake in the Conecuh National Forest to almost nonexistent. A change in the weather may have been the culprit, but I wasn’t nearly as concerned with the source of the problem as with the prospect that I might not find enough snakes to complete the research.

Luckily, we had a solution. Sean had a location that he knew would guarantee snakes en masse – unfortunately, it was too far away for just a day trip. Our solution was to hit this magical spot on the way back to Pennsylvania on our very last day.

I can’t tell you how nervous I was when we went out that final night.

To set the stage:

1. We didn’t have enough snakes. More importantly, we really needed to find a specific subset of snakes, adult males, which are significantly more difficult to locate than adult females or juveniles.
2. We absolutely had to leave for Penn State the next morning, making this the absolute last possible time to get enough snakes to complete the project.
3. The previous four days hadn’t yielded even a single snake.
4. Just for dramatic effect, there was an absolutely MASSIVE electrical storm just prior to our sojourn, shaking the ground and lighting up the sky like a child repeatedly turning the lights on and off inside an otherwise pitch black room.

Well, that night could have shared a title with this post: Redemption in the Deep South. After working from 8:00 pm to 2:00 am, slogging through waist deep (read: sometimes neck deep) blackwater while exhausting our drinkable water, getting lost, and receiving (conservatively, of course) 1,000,000 mosquito bites, we found our 5 snakes. Three of the five were adult males. Our goals were complete. Victory.

I am still in the process of inputting data and running samples, so I can’t yet tell you how stress hormone levels in cottonmouths relate to their behavior. What I can tell you is that working through the trials and tribulations of this summer’s research has made me a better scientist by far. I have many to thank for making this experience possible, but my highest gratitude extends to Tracy Langkilde, Sean Graham, and the cottonmouths of south Alabama. Thanks guys.

Adapted from Mark Herr’s Presidential Leadership Academy Blog

Do Bearded Ladies Have Sexier Sons?

Our next undergraduate guest post is from Courtney Norjen, a senior majoring in Veterinary and Biomedical Sciences. Courtney is in the Schreyer Honors College and is also the president of the Pre Vet Club. Courtney spent the summer running multiple projects in the Langkilde Lab, and she discusses her independent research below:

This summer, I had the opportunity to begin working on my honors project in the Langkilde Lab.  In eastern fence lizards (Sceloporus undulatus), males have bright blue badges on their throats and bellies, while some females have paler forms of these badges and others are generally white underneath. The hormone testosterone regulates the development of these blue badges; lizards with more testosterone have larger, brighter badges.  These color patterns are especially important for social interactions, particularly during mating season. Males will behave aggressively towards lizards with male-typical coloration (they’ll display their own bright colors by doing push-ups to show that they’re better than the other males), while they court lizards with female-typical coloration. Recently, however, we’ve started noticing “bearded ladies,” females that display light blue badges, a male-typical trait.  Lindsey wanted to determine if males liked these bearded ladies as much as normal females.  She set up arenas where males had a choice between a bearded lady and a normal one, and her results showed that males prefer to mate with normal, white females rather than bearded ladies.  These blue females also tend to lay their eggs later in the season, which doesn’t give their offspring as much time to forage for prey and build up energy reserves before the coming winter.

A male (left) and female (right) fence lizard. The female has pale blue badges.

If having blue badges is bad for the females and potentially for their offspring, then why are there still bearded ladies in the population?  This is the question I hope to answer through my project!  In order for that male-typical trait to be retained in the females, there theoretically must be some positive outcome.  We hypothesized that females with higher testosterone levels (the ones that are blue) would produce bluer offspring, regardless of sex, and would produce more sons than daughters.  This is also known as the “sexy son hypothesis:” perhaps masculinized females will produce more magnificent sons!

To test this, we used two different experimental designs.  First, I took normal female lizards that were in the early stages of pregnancy and gave half of them testosterone five days per week while they were developing their eggs.  The testosterone comes in a powder form; I took the powder and dissolved it in sesame oil, and then applied the oil to their backs.  The oil, along with the testosterone, gets absorbed through their skin!  The other half of the lizards was treated with plain sesame oil as a control.  The second design involved using eggs that had already been laid by normal lizards.  Half of the normal eggs were dosed once with testosterone prior to incubation, and the other half were given plain oil instead. In about 45 days, the eggs started hatching!

A brand new hatchling fence lizard.

I am now in the midst of raising 70 hatchlings!  I will be comparing the male to female sex ratios, the growth rates, and the coloration of these offspring between the two hormone treatment groups and between the two experimental designs.  If my hypothesis is correct, the lizards that were dosed with testosterone will have more sons than daughters, and their offspring will generally be bluer than the lizards that were not given testosterone.  I don’t have any results to present to you yet, but I’m really excited to see what happens!

Courtney holding one of her study subjects while in the lizard room.

Hanging-thieves!

No, it’s not the name of a cool new band you haven’t discovered yet; the Hanging-thieves are a genus (Diogmites) of robber flies that is known for hanging by one or two legs from a perch site while consuming their prey. Robber flies (family Asilidae) are also known as assassin flies, because they catch other bugs while on the wing and eat them with their piercing mouthparts. In general, robber flies are considered to be beneficial, as they eat many pest insects.

A hanging-thief chows down on a freshly caught wasp .

I was lucky enough to stumble upon a Hanging-thief (it looks like Diogmites salutans) as it was engrossed in eating a wasp. The robber fly was perched on one of my AED’s (Armadillo Excluder Devices), small wire cages that I set up to protect my fake lizard nests from predation by wily armadillos (that story another time…). The fly had already paralyzed its prey with venom, which also contains enzymes which liquefy the prey’s innards (as in spiders). As I watched, the robber fly carefully manipulated the wasp, turning its body over and around, and inserting its proboscis into likely sites, as if its prey were some sort of delicious juice box. In the video below, you can see the mouthparts of the fly working to slurp up its presumably delicious meal.

My favorite part of seeing this robberfly was observing its moustache, or mystax, which is so tough that it may help protect the fly from injury when it is subduing struggling prey.

The mystax is among the most fearsome of moustaches in the animal kingdom.