Adaptations to introduced ants with lizard data

Methods

Activity

3. Take several minutes to draw a picture, however crude, of natural selection—the process whereby organisms better adapted to their environments tend to survive longer and produce more offspring.

Background concepts: Adaptation

In ecology, adaptation can mean different things. Sometimes the word refers to the traits that organisms evolve through the process of natural selection that allow them to survive in their particular environments. These evolved characteristics can be structural (physical), behavioral, or physiological (internal body processes). 

Additionally, “adaptation” can refer to how the form or behavior of an organism changes in response to environmental stimuli. It can also refer to the way a sense organ can become more or less sensitive as a result of prolonged stimulation. In this lab activity, we will focus more on the first two definitions of adaptation. 

Species information: Eastern fence lizard

The eastern fence lizard (Sceloporus undulatus) is a medium-sized lizard (10–18.5 cm) native to eastern North America and occurs from the Atlantic coast to as far west as Texas (Conant and Collins 1998). Belonging to the family of North American spiny lizards, eastern fence lizards have rough, pointed scales on their backs. They are gray to brown in color with a chevron pattern on their backs that can help them blend into their background. Most mature males, and some females, have patches of bright blue scales on their bellies and throats. Male eastern fence lizards establish and defend their territory by doing ‘push-ups’ to flash their blue scales and scare off other males.

Eastern fence lizards are sit-and-wait predators that eat a wide variety of insects, spiders, and other invertebrates, including ants. They are common in many habitats, particularly open forests and field edges. Their cryptic coloration (i.e., camouflage) can reduce visual detection by some of their main predators including birds and domestic cats.

Species information: Red imported fire ant

Solenopsis invicta, commonly known as the red imported fire ant in the United States, is a species of fire ant native to tropical and subtropical South America. These ants thrive in ecologically disturbed areas and live in a wide variety of habitats such as rain forests, deserts, grasslands, alongside roads and buildings, and in electrical equipment (Callcott and Collins 1996). The red imported fire ant is one of the most successful nuisance species in parts of the world where humans accidentally introduced it: Australia, New Zealand, several Asian and Caribbean countries and the United States. Red imported fire ants have spread considerably throughout the southeastern United States since accidentally introduced into the port of Mobile, Alabama via cargo ship during the 1930s. They currently inhabit nearly 370 million acres in the U.S. and Puerto Rico. Counties throughout Texas, Arkansas, Oklahoma, Mississippi, Louisiana, Tennessee, Alabama, Georgia, Florida, South Carolina, and North Carolina are under Federal Quarantine, which restricts the interstate movement of regulated articles (e.g., hay, plants, soil) to prevent the human-assisted spread of fire ants. 

Solenopsis invicta, like other ant species, perform valuable ecosystem functions such as nutrient turnover and soil modification, and serve as prey and detritivores within the system. As omnivores their diet consists of dead mammals, invertebrates (e.g., arthropods, earthworms), vertebrates, seeds, and sweet liquid substances from plants or honeydew-producing insects. In their native South American range, their abundance is moderated by competition with other ant species (Buren et al. 1974) and the presence of co-evolved predators (Porter et al. 1997). They often become the dominant ant species in introduced areas outside of their native range due to their aggressive foraging behavior, high reproductive capability and lack of predators and competitors (Allen et al. 2004). 

Introduced Species

Introduced species are organisms non-indigenous to a specific region. While many introduced species are not destructive, some cause substantial damage to the ecosystem, economy, and/or public health in their new environment. There are several mechanisms through which introduced species can spread and establish themselves: 

Introduction through human activities: This can happen accidentally through international trade, tourism, escape from captivity, or transportation. Organisms can “hitchhike” on goods, vehicles, or in ballast water from ships, or be introduced to an area by release of unwanted pets or live bait.

Intentional introduction: In some cases, species are deliberately introduced for specific purposes, such as agriculture, hunting, or ornament. However, these introduced species can become problematic if they have no natural predators, competitors, or pathogens in their new environment. 

Climate change: As the climate changes, certain areas may become more hospitable to species that were previously limited by temperature or other factors. This can lead to species expanding their ranges and becoming problematic in new areas. 

Lack of natural predators: Introduced species often lack natural predators or diseases that would normally keep their populations in check in their native ecosystems. This allows them to reproduce and spread rapidly in their new environment. 

Rapid reproduction and adaptation: Introduced species often have high reproductive rates and adapt well to different conditions. This gives them a competitive advantage over native species in their new habitat. 

Altered ecosystem dynamics: In some cases, introduced species can alter the ecosystem dynamics of their new habitat by outcompeting native species for resources like food, water, and shelter.

Where introduced, red imported fire ants can negatively impact people, agriculture, natural resources, and native organisms. They can damage crops by feeding on the buds and fruits of plants, especially corn, soybean, okra, and citrus. They sometimes chew through irrigation tubing and cause physical damage to other farm equipment from their mounds or through electrical shorts. A single fire ant can sting repeatedly, causing injury to, or killing livestock either intentionally to obtain prey or in fierce defense of their mound; young and newborn animals are especially susceptible to the ants’ venom. All these actions can result in substantial economic losses. They can also harm wildlife by reducing native ant and other invertebrate biodiversity and injuring or killing native birds, reptiles, amphibians, and mammals (Allen et al. 2004). Because of their painful, venomous sting, red imported fire ants are a nuisance introduced species, particularly in urban areas, and can even cause allergic reactions including rare instances of anaphylactic shock in humans (Potiwat and Sitcharungsi 2015).

Researchers are studying how fence lizards and other native species respond to the threat of red imported fire ants. When fire ants and fence lizards come into contact, the lizards’ behavior can determine whether or not they survive. Scientists discovered lizards that exhibit a behavioral response such as body twitching or fleeing can reduce their venom exposure and survive the attack. While it is possible that there exists a genetic basis to these behaviors and that lizard populations may adapt to exhibit higher frequencies of these behaviors, it is important to point out that not all behavioral responses are inherited and contribute to a population’s evolution. As seen in our second definition of adaptation, some adaptive behaviors that improve an individual’s longevity are environmentally induced and learned throughout an organism’s lifetime. 

4. Use the following chart of evolution misconceptions and realities to answer these questions. Read carefully and focus more on the “Reality” column.

a. Does evolution occur as a result of gradual or sudden environmental events, or both?

b. What role does reproduction play in evolution? This question will require you to 

c. What are the three primary paths through which evolution operates?

d. Does evolution occur through conscious processes of organisms?

Making predictions & analyzing results

Penn State biologist Dr. Tracy Langkilde conducts research on native fence lizards and their adaptive responses to the presence of red imported fire ants. The fire ants’ well-documented spread through the southern U.S., and the fact that they are currently restricted to just a portion of the lizards’ range, allows Dr. Langkilde’s team to compare how lizards from ecologically similar areas differ behaviorally and physically based on the presence or absence of these fire ants. 

Below, you will find Dr. Langkilde’s research study notes and data. Answer the questions that follow.

5. How many different locations were the fence lizards in this study from and where were they?

6. How were Time to Flee data recorded for lizards that did not flee (run) off the fire ant mount in 60 seconds?

7. Replace the underlined bracketed words based on background information that you have been given: “Lizards from [location] twitch [more/less] than lizards from [location] when exposed to red imported fire ants.”

You will now test your prediction. Open the spreadsheet, click the Data tab and familiarize yourself with the information provided.

To compare the two groups of lizards, those from Alabama versus those from Arkansas, you need to first calculate the average values for each. In column C, under Average # Twitches, type the following to calculate the average twitch times for Alabama: =AVERAGE(C2:C41). Note, we are only going cells C2 through C41 because cells C42 through C81 contain data for Arkansas.

Alternatively, you can use a pivot table to compare these values and more. Refer to this link or your previous lab on animal activity for more information. 

8. What is the average number of times that lizards from Alabama twitched in response to fire ants?

9. Now, make the same calculation for Arkansas. What is the average number of times that lizards from Arkansas twitched in response to fire ants?

Now what? Are these averages considered different from each other, or are they pretty much the same? Scientists will use statistical tests to support these types of determinations. You will now analyze the data using what is called an unpaired t-test. The unpaired t- test compares the averages of two groups. 

10. What are the two groups you are comparing?

11. What variable did you calculate the average for?

T-test and p-values

To perform a t-test, we will use the T.TEST function in Google Sheets. Find any cell you like and insert the following function: =T.TEST(x, y, 2,3). Replace x with the range of the first group and y with the range of your second group. Note the last 2 arguments for the T.TEST function (2,3) are pre-specified in this exercise because their meanings are beyond the scope of this lesson. 

The t-test will yield a p-value. A p-value is a metric that allows us to determine whether the signal we are observing in our data is significant or not. If we find a difference between two groups, we want to make sure that that difference is not due to random chance variations, but rather a result of an actual difference between the groups. 

A p-value can fall between 0 and 1, as it is reflective of a percent chance. For example, a p-value of 0.12 indicates that there is a 12% chance that the statistical signal we see is caused by random sampling error. In other words, it is the probability of our signal being a fluke. For most applications, scientists determine a cutoff of 0.05 for an acceptable p-value. A p-value below 0.05 leads us to reject the null hypothesis. A p-value above 0.05 does not lead us to accept the null hypothesis, but instead we fail to reject the null hypothesis. In short, the null hypothesis is the assumption of no difference between our groups.

When testing data, we must develop a null hypothesis (H0) and an alternative hypothesis (HA). The null hypothesis is the assumption that there will be no statistically significant groups, and the alternative hypothesis is the assumption that there will be a statistically significant difference. You will learn more about null and alternative hypotheses and formulate your own in a future lab activity.

12. What p-value did your t-test yield?

13. What does this p-value lead you to believe about how significant the difference is between the average number of twitches of the two groups of fence lizards?

14. Was your earlier prediction from question 7 correct? 

Now, you will make a prediction and analyze different data (Time to Flee) from the same set.  

15. Replace the underlined bracketed words based on background information that you have been given: “Lizards from [location] flee [quicker/slower] than lizards from [location] when exposed to red imported fire ants.”

16. What is the average time to flee for lizards from the Alabama site when exposed to fire ants?

17. What is the average time to flee for lizards from the Arkansas site when exposed to fire ants?

18. What unit is Time to Flee reported in?

19. What p-value did your t-test yield?

20.  What does this p-value lead you to believe about how significant the difference is between the time to flee of the two groups of fence lizards?

21. Was your earlier prediction from question 15 correct? 

22. How many seconds faster, on average, did lizards from Alabama flee from fire ants than lizards from Arkansas? 

23. Why do you think the lizards from Alabama fled faster than those from Arkansas? 

24. What can happen if a lizard does not twitch or flee (run away) in response to fire ants? 

25. Based on the data you analyzed, which population of lizards, those from Alabama or Arkansas, do you think would survive better in the presence of fire ants? Explain why.

This [twitching/fleeing] behavior is common in baby fence lizards, which are vulnerable even to native ants, but is usually lost in adults as they outgrow threats from native ants by getting larger. However, in areas with fire ants, adult lizards retain this behavior that better enables them to survive fire ant attack. 

Dr. Langkilde’s research suggest that the higher percentage of behaviorally responsive adult lizards in fire ant areas could be the result of selection acting against unresponsive adults (e.g., if you don’t twitch, you have a higher chance of dying from fire ant attack, and hence reproducing) and/or it could also be due to lifetime exposure that leads lizards to benefit from continuing some of their juvenile behaviors into adulthood.

What this means is that the fence lizard’s twitching behavior is complicated! It may be that natural selection acts on a heritable twitching trait across generations for all lizards, even those that haven’t been exposed to fire ants, because babies that twitch are more likely to survive into adulthood and reproduce. And, as lizards that live where fire ants have been introduced grow into adults, they learn to “keep twitching” because that behavioral response removes attacking fire ants. 

26. Do you think adult lizards from Alabama twitched and fled in response to fire ants when they were first introduced to the area in the 1930s? Explain why or why not.

Beyond Behavior 

Watch the lizard video again. Speak to your neighbor about the physical characteristic you think is most important in helping lizards remove fire ants? Focus on physical (body) traits that are easily measurable.

Below is a description of more of Dr. Langkilde’s data from the spreadsheet that you will be using in this section.

27. All the data you have analyzed up until now have been behavioral data—how the lizards from different populations responded behaviorally to fire ants. Think back to the video you just watched. What physical (body) trait would best help a lizard remove fire ants crawling on it? You may want to watch the video again.

28. There are two types of physical trait data that Dr. Langkilde measured for each lizard, BL and HLL. What do these abbreviations stand for?

29. Why was it important to calculate a relative hind limb length?

In cell C2 of the Data tab, write out a formula that will automatically calculate the REL HLL from the BL and HLL. The formula will be “=F2/E2”, because those are the respective cells that contain HLL and BL. At this point, Google Sheets pay prompt you to simply autofill the remaining cells, but if this does not happen, click the first cell, then hold CTRL or CMD and click the last cell in column G. Then, with all of the column G data cells selected, press CTRL+D or CMD+D. 

Using formulas in this way is extremely powerful, as even if you add new data or previous data is updated, the calculated output will change to update automatically. The possibilities for using equations in Sheets can scale very well, as it is possible to string together many complex functions in even a single cell.

30. What unit of measurement is REL HLL recorded in?

31. Make a prediction based on the background information that you have previously been provided: Lizards from [location] have [shorter/longer] hind limbs than lizards from [location].

Now, calculate the averages of REL HLL for each location using methods from earlier.

32. What is the average relative hind limb length of lizards from Alabama?

33. What is the average relative hind limb length of lizards from Arkansas?

34. Do these two values seem very different, yes or no? What is the mathematical (i.e., numerical) difference between them?

Now, you will once more perform a t-test on the REL HLL of the two groups of lizards. 

35. What p-value did your t-test yield?

36. What does this p-value lead you to believe about how significant the difference is between the REL HLL of the two groups of fence lizards?

How can what looks to be such a small difference yield a statistically significant result? You must note that the statistical significance has little to do with how much difference there is numerically, but rather with how different the groups are from one another and how the data is clustered around the mean of each set. 

It is hypothesized that there is a genetic basis to these differences in relative hind limb length. Read another excerpt from Tylan and Langkilde (2023, The Conversation): 

We find lizard populations that have been living with fire ants have adapted [over generations] to have longer legs, which are better at removing fire ants when a lizard twitches and flees. This is a big shift for this species, reversing the latitudinal pattern we see in museum specimens—lizards tend to have shorter limbs the closer the population is to the equator.

Putting the pieces together

Watch a brief video from Penn State Eberly College of Science for insights on additional research from Dr. Langkilde’s team: “Lizards adapt to invasive fire ants, reversing geographical patterns in traits”

References

Adapted from:

• Freidenfelds, Nicole A, Jennifer M Deitloff, and Tracy Langkilde. “Lessons from Lizards: Adaptation to Introduced Ants.” Lessons in Conservation 13, no. 1 (2023): 44–60.

• Conant, R., and J.T. Collins. 1998. A Field Guide to Reptiles and Amphibians of Eastern and Central North America. 3rd ed. Houghton Mifflin Company. Boston, MA, USA.

• Callcott, A.A., and H.L. Collins. 1996. Invasion and range expansion of imported fire ants Hymenoptera: Formicidae in North America from 1918–1995. Florida Entomologist 79:240–251.

• Buren, W.F., G.E. Allen, W.H. Whitcomb, F.E. Lennartz, and R.N. Williams. 1974. Zoogeography of the imported fire ants. Journal of the New York Entomological Society 82:113–124.

• Porter, S.D., D.F. Williams, R.S. Patterson, and H.G. Fowler. 1997. Intercontinental differences in the abundance of Solenopsis fire ants (Hymenoptera: Formicidae): Escape from natural enemies? Environmental Entomology 26:373–384.

• Allen, C.R., D.M. Epperson, and A.S. Garmestani. 2004. Red imported fire ant impacts on wildlife: A decade of research. American Midland Naturalist 152:88–103.

• Potiwat, R., and R. Sitcharungsi. 2015. Ant allergens and hypersensitivity reactions in response to ant stings. Asian Pacific Journal of Allergy and Immunology 33:267–275.

• Langkilde, T., and N.A. Freidenfelds. 2010. Consequences of envenomation: Red imported fire ants have delayed effects on survival but not growth of native fence lizards. Wildlife Research 37:566–573.

• Tylan, C., and T. Langkilde. 2023. Native eastern fence lizards changed their bodies and behavior in response to invasive red imported fire ants. The Conversation. Available from: https://theconversation.com/native-eastern-fencelizards-changed-their-bodies-and-behavior-in-response-to-invasive-red-imported-fire-ants-186275 (accessed December 2, 2023).