Lesson 3: The impact of environmental changes on animal behavior: the butterfly’s potato chips

LESSON PLAN: HIGH SCHOOL
Authors: Dr. Emilie Snell-Rood & Dr. Johanna T. Ohlmeyer
Ecology, Evolution and Behavior
[ezcol_1quarter]Research context:[/ezcol_1quarter] [ezcol_3quarter_end]Snell-Rood et al. (2014, PNAS) found that road salt runoff can have significant effects on roadside herbivores. Salt leads to increased sodium in roadside plants which, at moderate levels, can lead to an increase in brain or muscle size in butterflies that grew up on these plants as caterpillars. However, we have no idea how this salt runoff might affect the foraging behavior of butterflies. Many animals are attracted to salty foods because sodium is important in development, but limited in availability. Are roadside plants perceived as “potato chips” to roadside feeding animals like deer and butterflies? Here, we will design and conduct an experiment to test this idea.[/ezcol_3quarter_end]

[ezcol_1quarter]Objectives:[/ezcol_1quarter] [ezcol_3quarter_end] 1. To demonstrate the process of science– students will be engaged in asking questions, designing projects, collecting and interpreting data.
2. To show that through citizen science projects, students can answer completely open scientific questions!
3. To verbally present and write a report based on the results of the study.
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[ezcol_1quarter]Students’ Skills:[/ezcol_1quarter] [ezcol_3quarter_end] Observation, application of background knowledge, analysis, making use of knowledge, evaluation of results, and synthesis. These skills were drawn from Bloom’s Taxonomy and the constructivist list. NGSS connections:  HS-LS1 From Molecules to Organisms: Structures and Processes, HS-LS2 Ecosystems: Interactions, Energy, and Dynamics, HS-LS3 Heredity: Inheritance and Variation of Traits, HS-LS4 Biological Evolution: Unity and Diversity[/ezcol_3quarter_end]

[ezcol_1quarter]Materials:[/ezcol_1quarter] [ezcol_3quarter_end] 1. For the background exploration: computers to access the literature, textbooks, and material reference.
2. For the experiment: cabbage seeds or small plants, NaCl to make salt water, distilled water, Cabbage white butterflies (Pieris rapae) caught or from Carolina Biological Supply (CBS) company, butterfly eggs from CBS company, cages or bug dorm cage from BioQuip, graph paper. [/ezcol_3quarter_end]

[ezcol_1quarter]Lesson:[/ezcol_1quarter] [ezcol_3quarter_end]1. This is a long-term experiment (about 4-6 weeks, 1-5 hours/week). Students may work in small groups of two or tree but each student will produce their own report.

2. The class starts with a question for initial class exploration. Entry card:  Does road salt runoff affect foraging behavior of animals? How? Write down three ideas. The students should ponder the answer for a few minutes alone or in pairs, write the answer on an index card, and finally offer their ideas to the class for open discussion.”

3. Asking questions and designing experiments: (see notes to the teacher) Students should first spend some time with the background information (Research context) formulating a research question and hypothesis. Armed with this, can they design an experiment (or two) to test this idea. What would the experiment look like? If the hypothesis is true, what would you predict would happen in your experiment, can you draw a graph? Of the proposed experiments, which ones seem like they might have problems or be difficult to conduct?

4. Example Experimental approach:

Question: Does road salt runoff affect foraging behavior of animals?

Hypothesis: Because sodium is an important limited micronutrient, and animals actively seek out sodium, animals will be attracted to salty roadside plants.

Basic experimental design: Through watering with salt water, students will create plants that differ in salt levels. Herbivore choice can be measured in two ways – preference of egg-laying females and/or preference of foraging caterpillars.

Prediction: If animals are attracted to salty roadside plants, adult female butterflies will lay more eggs on the salt-watered plant than control plants; caterpillars will eat more of the salt-watered plant than control plants.

Possible alternative explanations or confounding variables to consider (solution):

  • are animals attracted to a salty plant, or salty soil? (present leaves)
  • high salt levels increase mortality – at some point eating high salt leaves may start to result in higher caterpillar death (use several salt manipulations)
  • different plants store salt differently (use plants we know have salt effects)

 

Experimental protocol:

  • Plant cabbage seeds in pots. Once seedlings are several weeks old (robust looking plants), randomly divide them into control or treatment groups. Treatment groups will be watered with salt water. Students can experiment with different levels. We’ve found that watering with 1% NaCl solution for 2-3 weeks significantly increases sodium levels in plant tissue (2000-10000 ppm). Control plants should be watered with regular tap or distilled water. The salt treatment should not be continued indefinitely because NaCl can be stressful for plants.
  • Cabbage white butterflies (Pieris rapae) feed on plants in the mustard family, including cabbage and kale. They can be caught most anywhere in the US, or ordered from Carolina Biological Supply company if the experiment is being performed in the winter. For the caterpillar test, it would be best to obtain butterfly eggs (e.g., on parafilm strips from Carolina) and let them hatch on control cabbage plants.
  • Once they have grown into larger caterpillars (after about 2 weeks) would be a good time to do the feeding test. The easiest way to run a feeding test is to setup leaves from control and treatment plants in plant water tubes, rubber banded together so the plants are touching and the caterpillar can crawl between them. You can take a picture of leaves before and after the test to see how much the caterpillar ate. For a fifth instar caterpillar, a 3-4 hour feeding test is generally enough time for them to wander around and eat some.
  • Questions to ask for the students to think about: how many caterpillars should be used? If they grew up feeding on cabbage, might that feeding experience have had an effect on the results? If so, what might be a way to control for that? The caterpillars start out really small – think their food preferences might be different then? How would you measure their preference if they can’t move around much?
  • For measuring adult butterfly plant preference, you will need mated female cabbage whites. These can be obtained by rearing them in the lab and letting males and females mate in cages put in full sunlight, or by collecting females from the wild (they are generally mated and ready to go). Place 2-3 mated adult females in a cage (e.g., bug dorm cage from BioQuip) with a control and treatment plant. The cage should be in full sunlight (not under fluorescent bulbs) – windowsills work well. Give the butterflies 24 hours to lay eggs, then remove the plants and count the number of eggs on each plant.
  • Questions to ask for the students to think about: how many groups of butterflies should be run (how many trials)? Female cabbage whites are often more cooperative if you put them into cages in groups – think there would be different results if you had groups of 1, 2, 5 or more females?

 

Data analysis:

There are multiple ways to analyze a data set that would come out of an experiment like this. The easiest way might be to, for a given trial, take the difference between the treatment plant and the control plant (e.g., # eggs on treatment plant – # eggs on control plant or area of treatment leaf eaten – area of control leaf eaten). Is the average of this value greater than zero? That’s what one would predict if there is a preference for the salty plant. More advanced classes may be able to delve into statistical analyses of the data set that account for variance and sample size.

 

5.  Report: Each group of students will present their findings and interpretation of the results to the class. The rest of the class should provide feedback to the presenters. The question to answer in the small presentation is:  Does road salt runoff affect foraging behavior of animals? In addition each student will write a report.

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[ezcol_1quarter]Class Closing:[/ezcol_1quarter] [ezcol_3quarter_end]
1. Collect entry card.
2. Homework: a) After each class, think about what have you learned, go over your lab notebook, and add comments, ideas, and question. b) Complete Part 1 experimental proposal handout. c) Complete part two of your handout and submit your written report along with your notebook. [/ezcol_3quarter_end]

[ezcol_1quarter]Assesment:[/ezcol_1quarter] [ezcol_3quarter_end] 1. Participation in class, project presentation, and discussions (peer review).
2. Grading: Homework  and completeness and thoroughness in filling out the lab notebook and index card. Written report.
3. Use a general rubric to evaluate the students’ overall performance.[/ezcol_3quarter_end]

[ezcol_1quarter]Teacher’s Reflections[/ezcol_1quarter] [ezcol_3quarter_end]1. Things that I did not cover.
2. Did I meet the lesson objectives?
3. Comments, conclusions, and modifications.
4. Pedagogical value of the lesson. Did my students learn the concepts and ideas explored in this class? Did the assessments provide enough evidence of understanding?[/ezcol_3quarter_end]

[ezcol_1quarter]Notes to the Teacher[/ezcol_1quarter] [ezcol_3quarter_end]
1. This experiment may need preparation, for example, in the interest of time the cabbage pants could be already growing.

2. It would be great if students have the opportunity to catch some Cabbage white butterflies (Pieris rapae) in the wild.

3. It is a good idea to hand out the student’s handout activity parts II and I separately.

4. Make sure the students are interacting in the groups. It is important to circulate from group to group to make sure the students are on the right track.

5. See the Snell-Rod PNAS paper for background and update on these concepts. Anthropogenic changes in sodium affect neural and muscle development in butterflies. PNAS 2014 Jul15; 111(28): 10221-6. http://www.ncbi.nlm.nih.gov/pubmed/24927579

6. During “Asking questions and designing experiments” students should be encouraged to be creative and present their own ideas. However they may need scaffolding and enough background to design the experiments. Thus we recommend leaving enough time to discuss the research context, and depending on the students’ abilities and interest, read the PNAS paper. They should have available additional background resources.

7. The teacher can have the students come up with their own range of salt concentrations. However ending up with one protocol will be a desirable in order to pull together all the groups’ results and to come up with a class report.

8. It is important to encourage the students to use a notebook or a lab book. Teachers can use this to follow progress and leave “questions to think about” as homework.

9. This class requires scaffolding and constant interaction with the students. When possible use prompts and questions to make sure they understand the concepts. Encourage the students to write in their lab notebooks.

10. If time allows in each class, there should be a discussion about the on-going work.

11. Exploring other citizen science projects: http://en.wikipedia.org/wiki/List_of_citizen_science_projects

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ACTIVITY: HIGH SCHOOL

cropped-owlet_logo.gifSalt is an important nutrient for herbivores like butterflies. In this experiment we are going to find out whether they prefer to feed on plants that contain more salt. Are salty plants the butterfly “potato chips”?

.

 

YOUR ACTIVITY: In this experiment you will formulate questions, make a hypothesis, design an experiment, collect data, interpret and present your results. Your findings will help us understand whether butterflies actually prefer salty plants, and if salt due to road runoff is changing their feeding behavior.

Part I:

We need to be familiar with the background information:

1. Your teacher will make groups of two or three students. Read the research context below:

“Snell-Rood et al. (2014, PNAS) found that road salt runoff can have significant effects on roadside herbivores. Salt leads to increased sodium in roadside plants which, at moderate levels, can lead to an increase in brain or muscle size in butterflies that grew up on these plants as caterpillars. However, we have no idea how this salt runoff might affect the foraging behavior of butterflies. Many animals are attracted to salty foods because sodium is important in development, but limited in availability. Are roadside plants perceived as “potato chips” to roadside feeding animals like deer and butterflies? Here, we will design and conduct an experiment to test this idea”.

 

2. Spent some time thinking and talking with your peers about the research context, and produce two or three ideas to discuss with the rest of the class. Your teacher will make available additional reference material for you to use, including the 2014, PNAS paper from Dr. Snell-Rood.

 

3. Also, you will need your lab book to record information, observations, questions, etc. Write as much as you can, be reflective and creative.

 

4. Consider the following points when answering (5) below.

  • What would the experiment look like?
  • If the hypothesis is true, what would you predict would happen in your experiment, can you draw a graph?
  • Of the proposed experiments, which ones seem like they might have problems or be difficult to conduct?

 

5. As a group use the format below to formulate your experiment:

Question:

Hypothesis:

Experimental Design:

Predictions:

6. Make sure you are ready for class discussion and that your ideas are backed-up by evidence.

 7. When your teacher calls, join the class and participate. This is the time for you to contribute your observations, ask questions, and listen to your peers. Use all this information to complete your experiment proposal (homework).

 

 

Part II:

Class experimental approach:

1. Question: Does road salt runoff affect foraging behavior of animals?

 

 2. Hypothesis: Because sodium is an important limited micronutrient, and animals actively seek out sodium, animals will be attracted to salty roadside plants.

 

3. Basic experimental design 1: Through watering with salt water, you will create plants that differ in salt levels. Herbivore choice can be measured in two ways – preference of egg-laying females and/or preference of foraging caterpillars.

 

4. Prediction: If animals are attracted to salty roadside plants:

  •  adult female butterflies will…

 

  •   caterpillars will…

 

 5. Possible alternative explanations or confounding variables to consider. Please write a possible solution:

  • are animals attracted to a salty plant, or salty soil?

 

  •  high salt levels increase mortality – at some point eating high salt leaves may start to result in higher caterpillar death.

 

  • different plants store salt differently

 

6. Experimental protocol:

Preference of foraging caterpillars.

1. Your teacher will provide you with plant cabbage in pots. Divide them into control or treatment groups.

Treatment groups will be watered with salt water for 2-3 weeks. Experiment with different levels of Salt _________________

Control plants should be watered with regular tap or distilled water.

 

2. Cabbage white butterflies (Pieris rapae) feed on plants in the mustard family, including cabbage and kale.

Obtain from your teacher butterfly eggs on parafilm strips and let them hatch on control cabbage plants. Once they have grown into larger caterpillars (after about 2 weeks) would be a good time to do the feeding test.

The easiest way to run a feeding test is to setup leaves from control and treatment plants in plant water tubes, rubber banded together so the plants are touching and the caterpillar can crawl between them.

You can take a picture of leaves before and after the test to see how much the caterpillar ate. For a fifth instar caterpillar, a 3-4 hour feeding test is generally enough time for them to wander around and eat some.

 

Think about and answer the following questions:

  • how many caterpillars should be used?

 

  • If they grew up feeding on cabbage, might that feeding experience have had an effect on the results?

 

  • If so, what might be a way to control for that?

 

  •  The caterpillars start out really small – think their food preferences might be different then?

 

  • How would you measure their preference if they can’t move around much?

 

 

Adult butterfly plant preference

1. You will need mated female cabbage whites. Your teacher will decide whether to obtain them by rearing them in the lab and letting males and females mate in cages put in full sunlight, or by collecting females from the wild (they are generally mated and ready to go).

 2. Place 2-3 mated adult females in a cage (bug dorm cage) with a control and treatment plant. The cage should be in full sunlight (not under fluorescent bulbs) – windowsills work well.

 3. Give the butterflies 24 hours to lay eggs, then remove the plants and count the number of eggs on each plant.

 

 

Think about and answer the following questions:

  • How many groups of butterflies should be run (how many trials)?

 

 

  • Female cabbage whites are often more cooperative if you put them into cages in groups – think there would be different results if you had groups of 1, 2, 5 or more females?

 

 

Data analysis:

For a given trial, take the difference between the treatment plant and the control plant:

 

# eggs on treatment plant – # eggs on control plant

trial 1

trial 2

trial 3

Average:

 

area of treatment leaf eaten – area of control leaf eaten

trial 1

trial 2

trial 3

Average:

 

 

  • Is the average of this value greater than zero?

 

 

  • What does your result tells you about the preference for the salty plant?

 

 Report:

1. As a group prepare a small 5 minutes presentation of your results to present to the rest of the class. You will receive and give feedback from/to your peers.

2. Each group will also enter the results into a class database.

3. Be ready to comment and participate in the discussion, take note of other students’ ideas and comments. This will be useful for you to write up your individual report.

 4. Write a report using your data, the literature, your notes, and the class results (Is your data consistent with the rest of the class? If not why do you think there are different results?).