The Lizard Log

The Langkilde Lab in Action


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Hormone assays in the lab this Fall

Now that our field seasons are (mostly) over, the members of the Langkilde lab have been busy processing the blood samples we collected over the summer.

As I wrote last time, most of us are looking in some way at the impacts of environmental stressors on an animal’s behaviour, and the characteristics of the offspring they produce. In order to test these questions, we need to be able to quantifiably measure the stress levels of the animals we study.

To do this, we took blood samples from our study species in the field to measure levels of “stress hormones” (glucocorticoids, factors produced by the adrenal glands in response to stress). These hormones circulate in the blood, and correlate with the baseline stress levels of an individual – the more glucocorticoids we find in the blood, the more stressed an animal is. We determine the concentration of glucocorticoids in our blood samples by first centrifuging the sample to separate red blood cells from the plasma (the clear fluid in the eppendorf tube below), and then running the sample through an enzyme immunoassay.

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Enzyme immunoassays work by using antibodies that bind to the factor of interest in a sample – in our case, the steroid hormone corticosterone. We add a known amount of plasma to each well of the plate (above right), and in each well, the corticosterone in the sample binds to the antibodies. The antibodies that aren’t bound by corticosterone are bound by a conjugate tracer, which gives off a colour. So, in the plate above, the more “yellow” the well appears, the less hormone it contains (meaning that more free antibody sites have been left to bind with the yellow tracer-bearing conjugate). We can compare the “yellowness” of each well with wells containing a known amount of hormone, and this allows us to calculate the concentration of hormone in each sample:

I’ve had a lot of fun learning these techniques this Fall, processing my lizard samples, and helping Chris process some of his rattlesnake samples! I’m looking forward to reporting back on the exciting results these data contribute to over the next few months.

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Preparing samples with Chris and Danielle!

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Basking Site Use by Timber Rattlesnake Morphotypes – By Shawn Snyder

My name is Shawn Snyder and I am currently a senior majoring in Wildlife and Fisheries Science.  This is my first and only year working in the Langkilde Lab.  During the summer of 2016, I worked under Dr. Chris Howey as a Research Technician studying the effects of prescribed fire on timber rattlesnake populations.  This position provided me the opportunity to radio-track timber rattlesnakes, record habitat data on tracked snakes, catch new snakes (extremely fun), learn how to safely tube a venomous snake (even more fun), and conduct vegetation surveys.  Also, this position provided me the opportunity to formulate my own scientific question to test! Together, Chris and I thought up a small side-project that I could conduct throughout the summer, which provided me the fantastic experience of going through the scientific process, collecting my own data, analyzing those data, and now writing a manuscript so that I can share those results with the scientific world.

When we first started collecting data for my side-project I was a little apprehensive.  Once the data was collected and analyzed I realized that this project was going to take time and a large amount of effort to complete.  As the process of analyzing the data and then coming up with a plan for the manuscript began to take shape, I started to feel challenged and nervous by this new task. But weekly meetings with Chris to discuss the process of writing a manuscript have helped immensely.  This is my first manuscript and yes it is challenging, but it will all be worth it once we have a finished product. I have ambitions to continue on to a Graduate program after I graduate and this manuscript will help me build my C.V. to apply to Grad schools.

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Two yellow morphs bask alongside three black morph timber rattlesnakes at a gestation site. Although we did not use gestating (i.e., pregnant) females as part of this project, this shows you the posture of a basking snake and the difference in color morphs.

My research is investigating if the two distinct morphotypes of timber rattlesnakes (a dark, black morph and a lighter, yellow morph; see above picture) use basking habitat with differing amounts of canopy openness and solar radiation. Previous research suggests that the dark morph evolved in response to thermal limitations in the northern parts of its range.  Darker snakes have more melanin in their skin, which allows them to absorb more solar radiation and maintain a higher body temperature than yellow morphs.  Yellow morphs having this thermal disadvantage, in theory would have to choose basking sites that receive more solar radiation to compensate for this limitation if they wanted to maintain a similar body temperature to the black morphs.  Specifically, I am testing the hypothesis that yellow morphs use basking habitat that has more canopy openness and receives more direct solar radiation (i.e., sun) than basking habitat used by black morphs.

 

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A black morph male timber rattlesnake is seen courting a basking yellow morph female.  Once again, the difference in color morphs is striking and has led many to ask what selective pressures are maintaining this polymorphism.

To test this hypothesis, I measured canopy openness over basking yellow and black morphs. I used the timber rattlesnakes that are being radio-tracked for Dr. Howey’s main study as my sample population and placed a flag where a snake was found exhibiting basking behaviors (see picture below  for example).  We took a picture facing skyward directly over the snake using a camera with a fisheye lens.  This lens takes a picture of 180 degrees and captures an image of all of the canopy over the snake (see picture).  We can then analyze these hemispherical photographs using a computer program called Gap Light Analyzer to measure the percent canopy openness and the amount of direct solar radiation transmittance (i.e., rays of sunlight) for each basking site.  Direct solar radiation is when the sunlight reaches the forest floor with no obstructions from the canopy; as opposed to indirect solar radiation which may be radiation that is being reflected off of clouds, trees, or the ground itself.  Our study site is characterized as having a mature Oak/Maple forest with an abundance of closed canopy throughout the area.  Both morphotypes use this “closed canopy” forest throughout the summer as foraging grounds, and when they need to bask they must seek out areas where some sunlight is making its way through the canopy.  This is where my question becomes very important comparing the habitat used by each morph.

 

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A flag is placed next to a basking yellow morph.  An exact description of the habitat is recorded so that I can come back at a later time (when the snake is not there) and take a photo of the canopy directly over where the snake had been.

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Two examples of hemispherical photographs taken over two different basking timber rattlesnakes.  Both canopies actually have similar canopy openness, but the canopy on the left receives far more direct solar radiation based on the placement of those canopy openings.

So far, my results show that the two morphs use habitat that have similar percent canopy openness, however, there was a difference in the amount of UV transmittance between the basking sites used by the two morphs.  Canopy openness doesn’t necessarily designate a “warmer” site because the sun path may not go directly over the gaps in the canopy of that site, thus, the site wouldn’t receive large amounts of direct solar radiation.  Black morphs use basking sites that received lower amounts of direct sunlight.  They may be able to do this because the greater amount of melanin in their skin provides a greater ability to absorb whatever direct or indirect solar radiation is available more effectively. Yellow morphs use basking sites that received more direct solar radiation.  They could be forced to use these sites to compensate for their disadvantage in their thermal ability.  I am currently working on writing a manuscript for these data and hope to have it completed by the end of 2016.  Stay tuned for more on this manuscripts progress!

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Here is a picture of Shawn (holding a Hellbender!!) while on a break from collecting some amazing data.


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Efficacy of Daylighting… Part I

Pregnant female rattlesnakes prefer to maintain an elevated body temperature (~32 °C), which allows for a more optimal development of embryos.  In Pennsylvania forests, however, these warm temperatures are not very abundant.  So, in order to achieve these elevated body temperatures, pregnant females seek out rare, open habitat (known as gestation sites) that receive a lot of sunlight.  Sometimes, females may travel up to a mile from their den sites just to gain access to openings within the forest canopy.

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Yellow morph female timber rattlesnake sits coiled under a small rock overhang.

However, not all open gestation sites are created equally.  Last year, Mark Herr, Michaleia Mead, and I uncovered a trade-off at timber rattlesnake gestation sites of various sizes.  Gestation sites that were very open provided pregnant females with more sunlight and warmer body temperatures for a longer duration of the day.  But, these same sites also came with an increased risk of predation!

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At a timber rattlesnake gestation site, a red-tailed hawk swoops in on top of a foam timber rattlesnake model (not seen). A yellow-morph timber rattlesnake model is pictured behind the hawk.

More predators visited these more open sites, predators like bobcats, fishers, and hawks.  But, smaller, more enclosed gestation sites were so thermally poor, we observed females returning to their den at the end of the field season still pregnant!  The use of thermally unfavorable gestation sites may provide the rattlesnake with a sanctuary from potential predators, but there are still terrible repercussions for choosing to use these sites.  We have found that snakes using thermally unfavorable sites tend to give birth at a later date.  The timing that an animal gives birth is very important.  If rattlesnakes give birth to their offspring too late in the year, the small neonate offspring will have little, to no time to complete their first shed and then obtain a small meal before entering the den for hibernation.  It is believed that survival for neonates unable to do these two things is close to zero.  Further, some pregnant rattlesnakes that use thermally unfavorable gestation sites are known to abort their entire litter toward the end of the summer if she decides that they are developing too slowly.

So why would pregnant females continue to use small, more enclosed, thermally poor gestation sites?  Possibly because there was a decreased risk of predation?  Possibly, however, because of strong site fidelity?  Possibly these sites were, at one time, thermally favorable, but over the years vegetation has encroached upon these open areas and shaded out the once warm, sunny rocks.  Due to the rattlesnake’s fidelic response to locating a favorable gestation site, they now find themselves sitting among the shade throughout much of the day.

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Timber rattlesnake taking advantage of a small break in the canopy and some sunlight reaching the forest floor.

 

We’ve addressed the problem, now how do we fix it?

In order to manage timber rattlesnake populations better, forest and wildlife managers have begun to open up gestation sites, un-shading these areas from vegetation, in a process called “daylighting”.  However, recall that more open sites have an increased risk of predation.  So can we open up these sites just enough to let the sun in, but keep the hawks out?  In an attempt to suppress the increased risk of predation that we observed at more open sites, I have begun to direct daylighting techniques to target specific trees that would increase the amount of solar radiation a site would see, without greatly increasing the risk of predation.  To do this, I will use hemispherical photography (see picture below), observe the path of the sun throughout the gestation period, and then target those trees that overlap with the path of the sun.  This way I can open up each of the sites just enough, but keep those trees that do not overlap with the path of the sun and could perhaps maintain some decreased risk of predation.

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Hemispherical photograph at one of our gestation sites. The path of the sun throughout the entire gestation period is shown by the yellow arc. Trees that overlap with this arc can be targeted to improve the thermal quality of the site.

Of course I want to address this management idea with as much scientific rigor as possible for an ecological study.  This past summer, I visited six historic gestation sites within Pennsylvania.  Four out of the six sites are pretty shaded over, and I consider thermally poor.  Two of the gestation sites are fairly open and should provide pregnant females with a plethora of sunlight.  This summer, I surgically implanted nine pregnant timber rattlesnakes with temperature-sensitive radio transmitters, which allowed me to track these individuals throughout the summer. I followed each snake and noted their behaviors, body temperatures, and the date that they gave birth to their young.  Additionally, I measured available body temperatures and risk of predation just like we did in the previous experiment that Mark, Michaleia, and I completed last summer.  The catch is, this winter I will go into three of the six sites (along with US Forestry personnel), and we will remove specific trees blocking out the path of the sun.  Then next summer I will repeat everything and determine if Daylighting improved these thermally poor sites.  Will I see warmer available temperatures within the Daylighted sites?  Will snakes within these sites maintain warmer body temperatures and for a longer duration of the day?  Will these snakes give birth at an earlier date, allowing their young to shed and get a first meal before hibernation?  Will I still see an increased risk of predation?

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Biophysical models laid out at a gestation site and measuring the potential body temperatures that a snake could achieve at that site.

Many of these questions I won’t be able to answer until next year.  But, I am collecting some interesting data thus far.  As expected, the two sites that were more open did have warmer available temperatures.  Snakes occupying these sites maintained warmer body temperatures, moved less often, and were the first to give birth to adorable baby rattlesnakes!

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Neonate rattlesnake coiled up next its mother.

But, these sites were also visited by more potential predators.  In fact, in one instance we observed a red-tailed hawk swoop down and nab a garter snake that was basking alongside our pregnant rattlesnakes!

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Red-tailed hawk grabbing a quick dinner at a gestation site. I’m fairly certain this was a garter snake that got nailed.

The more enclosed sites were indeed cooler and snakes at these sites maintained cooler body temperatures and moved more often.  It appeared that some of these pregnant females shuttled between nearby sites in order to track the path of the sun.  In the morning the snake may be at one site, and in the afternoon that same snake would move to a nearby site.

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Female at a thermally poor area was typically found in the morning at a cut tree stump. In the afternoon she typically moved to another location.

Other snakes continued to move throughout the entire summer from one site to the next; constantly searching for a thermally suitable site where she could continue to develop her babies.  Unfortunately, all of these movements brought one of our mommas too close to a nearby road where someone swerved into the shoulder in order to run her over.  Although the loss of this mom was a little tough on me, it did show me just how important it is to improve these historic gestation sites.

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Our big momma rattlesnake could never find a suitable gestation site. She moved from potential site to potential site for about a month before coming too close to a nearby road.

Currently, as I am writing this blog, we are still waiting on some of our snakes from the thermally poor sites to give birth.  As we find neonates (babies) at each of our sites, I will also collect data on each of them so that we can compare body condition (health) among sites.

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Collecting data on newborn rattlesnakes is a fantastic way to start anyone’s day. I’m collecting data on body length and mass so that I can determine the body condition (or health) of each individual.

 

Next year, I will continue to track snakes throughout these same gestation sites.  However, following our daylighting management, we hope that all of our pregnant snakes will give birth at early dates, move less, and stay clear of predators.  To be continued…

 


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Solon Dixon!

Hey everyone!

Even though I am continuing the same project from last year (how maternal stress affects the offspring in fence lizards), there are still some striking differences. One of the biggest is that there are fellow grad students and a post doc this summer!

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From left to right: Cameron (PhD), myself (PhD), Kirsty (post-doc), and Dustin (PhD).

Also, last year we made the drive in one day, however this year we broke the drive up over 2 days. This gave us an excellent opportunity to experience different parts of the USA on our drive. For the night we stopped in Knoxville, TN and had dinner at an amazing place called Calhoun’s On The River. True to its namesake, it had a beautiful view of the Tennessee River!

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After all the driving, we finally made it back to Solon Dixon and started catching lizards. As usual, the lizards’ personalities were very evident.

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Male fence lizard unamused with our attempts to catch him

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Apparently the female lizards found that corner of the tub to be very interesting.

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As I went to put this female back in her tub, she refused to let go of my fingers!

On top of finding many fence lizards, we were also about to see many other reptiles and amphibians!

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A barking tree frog tightly hugging my finger.

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An American alligator, at a very reasonable size to handle.

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A yellow bellied slider who found a little bit of water to sit in.

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A glass lizard!

As I spend more time down here, I find it rubbing off on me more and more.

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Very tempted to get a cowboy hat.

After catching the females, our first trip came to an end. However, we were quickly back down to release the females and run experiments with the hatchlings. With us this time we had an undergraduate researcher, Jen!

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The Bayfront Park, overlooking Mobile Bay. Located right next to one of our field sites, Blakeley State Park.

As we wait for more hatchlings to emerge, we have been focusing on removing fire ants from some of the enclosures we built. As fire ants are highest in the mounds earlier in the day, this means some early mornings. On the up side, it also means we always get to see the sunrise.

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Sunrise right near the enclosures.

Most things have gone well, with only one piece of equipment starting to show signs of wear, but this just gave me an excuse to do some handywork!

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Used some steel epoxy to seal a leak in the pot we boil water in for fire ants.

Things have started to pick up in terms of hatchling, so soon you should be able to hear about how things are going with them. Until then, here is a pic from right here at Solon Dixon

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With the drier weather they are finally able to do prescribed burns.

Cheers,
David

 


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What’s all the fuss about Alabama?

Since joining the lab, I’ve heard nothing else more than field seasons in Alabama; from the heat, lizards, those darn fire ants, and the wonderful people they come in contact with. So of course, I was anxious as ever to finally embark on my first field season in the Langkilde lab. After months of preparation, designing projects and all the logistics involved, May 9th had finally arrived and it was time to head south.

The first round of people in the Alabama crew this year had 3 newbies to the lab; Dustin Owen (our personal herpetology specialist), Dr. Kirsty Macleod (our Scottish Post Doc who should’ve been born in the southern US) and Myself aka Frog stallion (long story). The last member of the crew was David Ensminger. This was his third year there, sort of making him our expert of all things and everything that we needed to know (in other words he was our ear to ask a million questions).

The wonderful staff of The Solon Dixon Forestry Education Center welcomed us to the 5 starred field station (in my opinion – I don’t know who could possibly refute that). There were also lots of Auburn University students that were super interested in our work and simply cool people to hang out with. Essentially, everyone we met made sure we left knowing that southern hospitality is 100% real. I now understand why everyone in the lab loves and talks about it so much.

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Some Auburn students were successful on their first lizard hunt. It was Herp week in their class, luckily for them, they had some experienced catchers around

Now on to the fun science! In case you’re reading this and don’t know, the Langkilde lab is well invested into the Fence Lizard and Fire Ant system, but I’m just going to focus on my specific part for now. In the broadest of terms, I am interested in the diet of the Fence Lizards, but of course I can’t help but wonder about other aspects of this creature. A former member of our lab (the undergrad king Mark Herr) published a paper that suggests fence lizards seemingly build an addiction to the fire ants. My first thought was the possibility that the fire ants may be more nutritious. In Alabama, I collected loads of ants to quantify carbs, lipids, and proteins in comparison to fire ants, hoping for something to support a risk-reward relationship. My next thought was, if the lizards are presented with a second option, what will they pick? To test this, The Lizard Queen (Dr. Tracy Langkilde) and I ran food preference trials. We used the fire ants and “Dory ants” (still waiting for a true identification, but we call them Dory ants) in tubs with one lizard to see the choices they made. Now back in State College, I’m going through and analyzing all the data. I won’t spoil the surprise, which will hopefully be published, so stay tuned in the near future.

 

 

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My first Catch of the trip. Gotta love when they just pose and hug your thumb!

 


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The Natural State

Happy Memorial Day!

I am happy to report that the “uninvaded” team has returned to PA, lizards in hand–or bag as it were. Our team, consisting of Braulio,  Caty, and myself, traveled to Tennessee and Arkansas. Arkansas prides itself on being “The Natural State” for its “natural scenic beauty, clear lakes and streams, and abundant wildlife.” I can’t speak to most of that, but it does have lizards!

Fence lizard with a regenerating tail.

Fence lizard with a regenerating tail.

Rainy and overcast days slowed us down a bit. As ectotherms, lizards rely on external sources of heat, which means they like to bask in sunny spots in order to warm up. The thick clouds didn’t provide many good basking opportunities, but thankfully a few lizards made an appearance in the brief moments of sun.

Sometimes fence lizards like rocky habitat.

Sometimes fence lizards like rocky habitat.

2blendinMany other lizards like to bask on trees.

We did see a few sunny days, which gave Braulio and Caty the opportunity to catch their first lizards.

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Braulio with a Tennessee lizard.

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One of Caty’s first catches!

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Lizard selfies are the best selfies.

We even managed some “expert” catches, on more than one occasion slowly driving by a basking lizard and noosing it through the open car window.

Because we were looking for females, we of course became experts at catching males. One male lizard really hoped I was a tree. We tried to return him to his log, but on two separate occasions he ran up my leg. Sorry little guy!

Nope, not a tree.

Nope, not a tree.

Another male, pictured below, really surprised me. Lizards vary in coloration, but not usually by much. I’ve never seen a fence lizard so dark!

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A very dark fence lizard. His chest badges were impressive as well!

After two weeks of catching, we headed back to the lab. Our females are now happily housed in their nesting boxes, and one has laid her first clutch of the season. We’re all excited to see the resulting hatchlings!

Check back soon for more stories and photos from the field as well as updates on the specific research projects happening this summer.

We even spotted a fence lizard on a fence. So satisfying. A fence lizard on a fence. So satisfying.


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All Good Things…

There is unrest in the Langkilde Lab. Several members have declared their intention to graduate and leave the Penn State University.

This graduatist movement, under the leadership of the mysterious Doctor Thawley, will make it difficult for the limited number of graduate students left to maintain scientific rigor and order in the fence lizard universe.

Principal Investigator Tracy, the Head of the Biology Department, returns to the field to orchestrate the critical succession of power and saurian expertise in the upcoming research season….

A More Dramatic Version…


The past few weeks have been eventful ones in the Langkilde Lab. Gail and I were both repping the lab at the inaugural Graduate Student Awards Luncheon. I received the Intercollege Graduate Student Outreach Achievement Award, an award close to my heart, for bringing research to the community and “commitment to advancing the welfare of the public through scholarly pursuits.” Gail received her highly prestigious Penn State Alumni Association Dissertation Award which included a very spiffy and heavy Distinguished Doctoral Scholar Medal. Previous lab members have received both of these awards, and we hope that our new lab members will continue this tradition of excellence and outreach!

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Gail receiving her totally legit medallion from Penn State President, Dr. Eric Barron.

Gail and I also both graduated this past Sunday. It was a great chance to hang out in the last row of the huge auditorium and crack jokes with Tracy, observe the diverse footwear of graduating graduate students, and attempt to determine the school with the most awesome PhD robes.

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We were totally serious throughout the entire two hour ceremony.

Unlike some other awards ceremonies (*cough* Awards Luncheon *cough*), we were allowed to sit and walk together which was great. None of us tripped while negotiating the steep, narrow stairs to the stage, and I even managed to shake the President’s hand with something approaching poise.

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I was the only graduate wearing formal Birkenstocks at the ceremony.

Afterwards, we gathered outside with our families and friends for a few pics to document our snazzy regalia and the fact that an institute of higher learning actually saw fit to give us advanced degrees.

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That’s a wrap!

Lab Undergraduate Extraordinaire, Mark Herr, graduated as well (though we have no photographic evidence of this). Mark has worked in the lab for four years and published several papers. In recognition of his work, Mark received several lab commendations, including a certificate of excellence for Use of the Word “Devastating” on a Daily Basis and the Award for Forcing the Creation of a Special SSAR Award for Undergraduates.

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Mark Herr is recognized for his efforts to reintroduce large mammalian predators to control overpopulation.

In spite of these losses, a new hope has arisen in the next generation of lab graduate students and post-doctoral scholars.

Tracy is leading the bulk of the lab force, including post-doc Kirsty and graduate students Cam, Dustin, David, and Michaleia, to Solon Dixon, where an old ally, Nicole Freidenfelds, will help them discover clues as to how maternal stress hormones influence behavior, stress physiology, and metabolism.

Gail has volunteered for one final mission in the field: training a team of lizard neophytes including Braulio and Caty to capture fence lizards, especially gravid females, at sites in Arkansas and Tennessee that remain uninvaded by fire ants. This team will return to the lab and allow these female lizards to lay their eggs. They hope to determine if ladies bearing typically male characteristics, in this case blue throat badges (“beards”), also have bearded offspring.

Stay tuned this summer for more exciting developments in the fence lizard universe!