We present a novel laboratory activity to introduce students to experimental approaches often used by biologists to study orientation in animals. We first provide an overview of the current understanding of magnetoreception – the ability of some organisms to sense magnetic fields. We then outline an exercise that uses common pill bugs (Armadillidium vulgare) to examine whether a pulsed magnetic field affects their directional preference. The first part of the experiment includes the construction and visual testing of a pulse magnetizer built using low-cost and easily obtainable materials. Afterward, students examine the orientation of pill bugs both before and after being subjected to a magnetic pulse. Finally, students analyze their results with circular statistics using the open-source R coding platform, providing them experience in coding languages and statistical analysis. The interdisciplinary and biophysical nature of this experiment engages students in concepts of electromagnetic induction, magnetism, animal behavior, and statistics.

## Introduction

Magnetoreception, or the ability to sense magnetic fields, is possibly the most enigmatic and poorly understood of the animal senses. A growing amount of evidence, primarily from behavioral experiments, has demonstrated that a diverse array of species perceive the strength and/or direction of Earth's magnetic field and use it to guide their movements (Johnsen & Lohmann, 2005). Unlike many other senses, however, which involve specific exterior structures (e.g., eyes and ears) that amplify or focus the stimulus, Earth's weak magnetic field can, in theory, interact with every cell within the organism – making the search for a “magnetoreceptor” like finding a “needle in a needle stack” (Johnsen, 2017).

A majority of the behavioral evidence supporting magnetoreception is based on orientation experiments. Orientation, or a preferred direction of movement, is often compared in animals exposed to various magnetic fields or treatments in the absence of other cues. One common treatment is a strong, brief, magnetic pulse. This magnetic pulse is believed to alter the pattern of magnetization of iron oxide (magnetite) particles in the body, in turn disrupting a possible magnetic sense and orientation (Johnsen & Lohmann, 2005). Indeed, this effect has been observed in several species (e.g., sea turtles, birds, and lobsters; Wiltschko & Wiltschko, 1995; Irwin & Lohmann, 2005; Ernst & Lohmann, 2016). The recorded data are directions, or angles, and must be analyzed using methods intuitively familiar, but mathematically different, from canonical statistical techniques (reviewed by Lee, 2010). This type of analysis, called circular statistics, is often unfamiliar to most biologists, which suggests that an initial familiarity with circular statistics during secondary and/or postsecondary education is needed.

The goals of the following laboratory exercise are to determine whether pill bugs (isopod crustaceans in the genus Armadillidium) orient in a preferred direction and whether exposure to a magnetic pulse alters this orientation. Magnetoreception has been described in both crustaceans and other isopods (reviewed in Lohmann & Ernst, 2014), but it remains unknown if pill bugs can also sense Earth's magnetic field. Students will construct a miniature pulse magnetizer as part of the procedure and examine its effect on pill bug orientation; physical concepts of electromagnetism will thus be introduced or reinforced.

This activity will also introduce students to basic principles associated with designing and conducting studies of animal behavior. First, the orientation assay has no defined or expected result, but rather emphasizes the importance of observing and recording behavior as it arises, noting differences within and between individuals, and keeping notes of sufficient detail for use in later analysis – all crucial tools in the study of animal behavior (Altmann, 1974). Second, the activity emphasizes the discussion of issues that can arise from external confounding variables or influences – for example, hunger level, temperature, health, previous experience, and personality. Although it is difficult to control for all confounding factors in any experiment, promoting an awareness of how these variables may affect animal behavior will highlight good scientific practices. Third, the problem of pseudoreplication can also be incorporated into the activity's discussion (see below). Pseudoreplication, or incorrectly treating data points as independent, has to be properly addressed when performing statistical tests and has been a point of concern among scientists (Hurlbert, 1984). Therefore, it is important that students are familiar with pseudoreplication and its consequences.

Finally, the analysis of circular data will both complement existing and introduce new statistical principles. This exercise is designed to be completed within a single lab period (~2 hours) and is ideal for students ranging from high school to undergraduate levels. For high school instructors, relevant Next Generation Science Standards (NGSS) and AP Biology and Physics 2 topics are described in Tables 1 and 2. A detailed description of the procedures and tutorials can also be found in the Online Materials (see below).

Table 1.
Next Generation Science Standards (NGSS) relevant to our lesson plan.
Scientific PracticeNGSS StandardRelation to Our Lesson Plan
Planning and carrying out investigations:
Plan and conduct an investigation individually and collaboratively to produce data to serve as the basis for evidence and in the design of further experiments.
HS-LS1-3 Students can work together and/or independently to carry out an experiment, in which they build an apparatus and collect all the data, with opportunities to alter the experimental specifics to answer additional questions.
Analyzing and interpreting data:
Apply concepts of statistics and probability to scientific questions and problems, using digital tools when possible.
HS-LS4-3 Students collect their own data and then analyze it using circular statistics. Additionally, we provide and describe online resources for using the R coding platform during statistical analysis.
Using mathematics and computational thinking:
Use mathematical and/or computational representations of phenomena or design solutions to support explanations.
HS-LS2-1 Students are introduced to and can use circular statistics, introducing them to likely novel ideas that are related to more familiar statistical measures, such as the mean.
Engaging in argument from evidence:
Evaluate the evidence behind explanations or solutions to determine the merits of arguments.
HS-LS4-5 Students must evaluate the data that they and their classmates collected in order to evaluate the evidence for and against magnetoreception in pill bugs.
Scientific PracticeNGSS StandardRelation to Our Lesson Plan
Planning and carrying out investigations:
Plan and conduct an investigation individually and collaboratively to produce data to serve as the basis for evidence and in the design of further experiments.
HS-LS1-3 Students can work together and/or independently to carry out an experiment, in which they build an apparatus and collect all the data, with opportunities to alter the experimental specifics to answer additional questions.
Analyzing and interpreting data:
Apply concepts of statistics and probability to scientific questions and problems, using digital tools when possible.
HS-LS4-3 Students collect their own data and then analyze it using circular statistics. Additionally, we provide and describe online resources for using the R coding platform during statistical analysis.
Using mathematics and computational thinking:
Use mathematical and/or computational representations of phenomena or design solutions to support explanations.
HS-LS2-1 Students are introduced to and can use circular statistics, introducing them to likely novel ideas that are related to more familiar statistical measures, such as the mean.
Engaging in argument from evidence:
Evaluate the evidence behind explanations or solutions to determine the merits of arguments.
HS-LS4-5 Students must evaluate the data that they and their classmates collected in order to evaluate the evidence for and against magnetoreception in pill bugs.
Table 2.
AP Biology and AP Physics 2 topics relevant to our lesson plan.
Enduring UnderstandingEssential KnowledgeRelation to Our Lesson Plan
Biology 2.E:
Many biological processes involved in growth, reproduction and dynamic homeostasis include temporal regulation and coordination.
Biology 2.E.3:
Timing and coordination of behavior are also regulated by several means; individuals can act on information and communicate it to others, and responses to information are vital to natural selection.…Examples include behaviors in animals triggered by environmental cues.
Students learn about animal behavior, specifically how to observe and quantify it, the factors that confound studies of animal behavior, and how behavior can vary between individuals. They also learn about how information in the form of magnetic fields can influence animal behavior.
Biology 3.D:
For cells to function in a biological system, they must communicate with other cells and respond to their external environment.
Biology 3.D.3:
Signal transduction pathways link signal reception with cellular response.
Students learn about the cellular basis of magnetoreception and receive an introduction to this unfamiliar sensory modality.
Biology 3.E:
Organ systems have evolved that sense and process external information.…These include sensory systems that monitor and detect physical and chemical signals from the environment.…The nervous system interacts with sensory and internal body systems to coordinate responses and behaviors.
Biology 3.E.2:
Animals have nervous systems that detect external and internal signals, transmit and integrate information, and produce responses.
Students learn about magnetoreception in general, its potential functions as a navigation guide, and how animals with a magnetic sense can detect external signals and initiate a behavioral response.
Physics 2.C:
An electric field is caused by an object with an electric charge.
Physics 2.C.4.1:
The student is able to distinguish the characteristics that differ between monopole field…and dipole fields (electric dipole field and magnetic field) and make claims about the spatial behavior of the field.
Students in our lesson plan learn about the relationship between magnetic and electric fields and gain hands-on experience creating a device that exerts an electric (and magnetic) field.
Physics 2.D:
A magnetic field is caused by a magnet or a moving electrically charged object. Magnetic fields observed in nature always seem to be produced either by moving charged objects or by magnetic dipoles or combinations of dipoles.
Physics 2.D.3.1:
The student is able to describe the orientation of a magnetic dipole placed in a magnetic field in general and the particular case of a compass in the magnetic field of the Earth.
Students learn about Earth's magnetic field, conduct an activity that involves finding magnetic north on a compass, and are able to observe how the compass needle changes in response to an artificial electromagnetic field.
Physics 1.B:
Electric charge is a property of an object or system that affects its interactions with other objects or systems containing charge.
Physics 1.B.1:
Electric charge is conserved. An electrical current is a movement of charge through a conductor.
Students gain hands-on experience building a coil that carries electric current.
Enduring UnderstandingEssential KnowledgeRelation to Our Lesson Plan
Biology 2.E:
Many biological processes involved in growth, reproduction and dynamic homeostasis include temporal regulation and coordination.
Biology 2.E.3:
Timing and coordination of behavior are also regulated by several means; individuals can act on information and communicate it to others, and responses to information are vital to natural selection.…Examples include behaviors in animals triggered by environmental cues.
Students learn about animal behavior, specifically how to observe and quantify it, the factors that confound studies of animal behavior, and how behavior can vary between individuals. They also learn about how information in the form of magnetic fields can influence animal behavior.
Biology 3.D:
For cells to function in a biological system, they must communicate with other cells and respond to their external environment.
Biology 3.D.3:
Signal transduction pathways link signal reception with cellular response.
Students learn about the cellular basis of magnetoreception and receive an introduction to this unfamiliar sensory modality.
Biology 3.E:
Organ systems have evolved that sense and process external information.…These include sensory systems that monitor and detect physical and chemical signals from the environment.…The nervous system interacts with sensory and internal body systems to coordinate responses and behaviors.
Biology 3.E.2:
Animals have nervous systems that detect external and internal signals, transmit and integrate information, and produce responses.
Students learn about magnetoreception in general, its potential functions as a navigation guide, and how animals with a magnetic sense can detect external signals and initiate a behavioral response.
Physics 2.C:
An electric field is caused by an object with an electric charge.
Physics 2.C.4.1:
The student is able to distinguish the characteristics that differ between monopole field…and dipole fields (electric dipole field and magnetic field) and make claims about the spatial behavior of the field.
Students in our lesson plan learn about the relationship between magnetic and electric fields and gain hands-on experience creating a device that exerts an electric (and magnetic) field.
Physics 2.D:
A magnetic field is caused by a magnet or a moving electrically charged object. Magnetic fields observed in nature always seem to be produced either by moving charged objects or by magnetic dipoles or combinations of dipoles.
Physics 2.D.3.1:
The student is able to describe the orientation of a magnetic dipole placed in a magnetic field in general and the particular case of a compass in the magnetic field of the Earth.
Students learn about Earth's magnetic field, conduct an activity that involves finding magnetic north on a compass, and are able to observe how the compass needle changes in response to an artificial electromagnetic field.
Physics 1.B:
Electric charge is a property of an object or system that affects its interactions with other objects or systems containing charge.
Physics 1.B.1:
Electric charge is conserved. An electrical current is a movement of charge through a conductor.
Students gain hands-on experience building a coil that carries electric current.

## Materials

See Table 3 for a list of materials. These are listed per group, two to four students per group being optimal.

Table 3.
List of materials required for this activity.
Required ItemsExample
9-volt battery Amazon (no. B00MH4QM1S)
9-volt battery connector, ends stripped Amazon (no. B06X8YZJ64)
2 m insulated wire (24 gauge), ends stripped Mouser Electronics Inc. (no. 566-8538-100-09)
Push-button switch (normally open, or NOAmazon (no. B01IU898QA)
4 cm length × 2 cm diameter (3/4 in) PVC pipe Amazon (no. B003OSKZF0)
Two to four 1.5 cm rubber bands Amazon (no. B007PJ4Z82)
Compass Carolina Biological Supply Co. (no. 974291)
Ammeter/multimeter Carolina Biological Supply Co. (no. 757460)
Electrical tape Carolina Biological Supply Co. (no. 973464)
Pill bug (Armadillidium sp.Field collected or Carolina Biological Supply Co. (no. 143080)
76 × 20 mm plastic tube Sarstedt AG & Co. (no. 80.734, spatula removed)
Stopwatch Carolina Biological Supply Co. (no. 962106)
Ruler Carolina Biological Supply Co. (no. 702616)
Paper orientation arena See Online Materials for a printable copy
Optional Items Example
DC magnetometer AlphaLab Inc. (no. DCMHSACDC)
Alligator clips Carolina Biological Supply Co. (no. 756521)
470 Ohm resistor Amazon (no. B0185FIDF0)
5 mm LED Amazon (no. B005ONQ41W)
Required ItemsExample
9-volt battery Amazon (no. B00MH4QM1S)
9-volt battery connector, ends stripped Amazon (no. B06X8YZJ64)
2 m insulated wire (24 gauge), ends stripped Mouser Electronics Inc. (no. 566-8538-100-09)
Push-button switch (normally open, or NOAmazon (no. B01IU898QA)
4 cm length × 2 cm diameter (3/4 in) PVC pipe Amazon (no. B003OSKZF0)
Two to four 1.5 cm rubber bands Amazon (no. B007PJ4Z82)
Compass Carolina Biological Supply Co. (no. 974291)
Ammeter/multimeter Carolina Biological Supply Co. (no. 757460)
Electrical tape Carolina Biological Supply Co. (no. 973464)
Pill bug (Armadillidium sp.Field collected or Carolina Biological Supply Co. (no. 143080)
76 × 20 mm plastic tube Sarstedt AG & Co. (no. 80.734, spatula removed)
Stopwatch Carolina Biological Supply Co. (no. 962106)
Ruler Carolina Biological Supply Co. (no. 702616)
Paper orientation arena See Online Materials for a printable copy
Optional Items Example
DC magnetometer AlphaLab Inc. (no. DCMHSACDC)
Alligator clips Carolina Biological Supply Co. (no. 756521)
470 Ohm resistor Amazon (no. B0185FIDF0)
5 mm LED Amazon (no. B005ONQ41W)

## Preparation

### Species Collection

Pill bugs (Armadillidium spp.), also known as woodlice, sow bugs, potato bugs, or roly polies, are a terrestrial crustacean of the order Isopoda. They are harmless, easily handled, and known for their ability to roll into a ball in order to protect themselves. The common pill bug, Armadillidium vulgare, is native to Europe but has been introduced worldwide and is regularly found in North America. Instructors (or students) can collect pill bugs underneath loose leaf litter or debris on the ground, often in cool, dark places. Alternatively, they are readily available for order from biological supply companies (see Table 3). They can be kept on lightly damp soil or paper towels in any small container at room temperature with air circulation. Just prior to the beginning of the experiment, it is recommended to transfer each pill bug to a separate 76 × 20 mm tube (Table 3) containing a small, damp piece of paper towel.

### Wiring Preparation

Depending on the time available and student background, the instructor can perform several steps to facilitate the construction of the pulse magnetizer. We recommend having all wire leads previously stripped of insulation and pre-connecting (preferably with solder) the push-button switch to the positive (red) terminal of the 9-volt battery connector. Connections can also be made between two bare wires by twisting together tightly and covering with electrical tape.

## Procedure

### Part 1: The Mechanisms of Magnetoreception

We provided a handout ( Appendix) and lecture slides (see Online Materials) that the instructor can use to familiarize students with the proposed mechanisms of magnetoreception. The instructor can choose to present the material in the lecture or have the students learn independently from the handout. Comprehension questions are provided in the handout.

### Part 2: Circular vs. Linear Statistics

Because the data collected in orientation studies are often directions, or angles, they have to be analyzed differently from traditional linear data. For example, if we have two angles of 2° and 358° marked on a 360° circle, they are nearly identical (both are near 0°). However, if we calculate a traditional (or arithmetic) mean, one would mistakenly report it to be $2+3582=180°$. This would point in the entirely opposite direction! By using geometry, we can actually calculate the circular mean correctly as 0° (for more information, see the Online Materials). The handout ( Appendix) provides an additional example that uses simple math that students can work by hand to illustrate how using linear statistics, for example calculating a mean, is incorrect when applied to circular data.

### Part 3: Construct the Pulse Magnetizer

The pulse magnetizer circuit consists of a 9-volt battery, a switch, and a coil (Figure 1A, B and Figure 2A). Alternatively, a 470 Ω resistor and light-emitting diode (LED) can be added in parallel with the coil to illuminate when the switch is pressed (Figure 1C). The coil is constructed by tightly wrapping the electrical wire around the PVC pipe, leaving ~5 cm excess on either end available for connections (Figure 2B–D). The rubber bands can be applied to either end of the coil to secure the wire and prevent the coil from loosening. Connect one lead of the coil to one terminal of the switch, then connect the remaining terminal of the switch to the positive (red) lead of the battery connector (the instructor may prepare this ahead of time as described above). Connect the positive (red) probe of the ammeter/multimeter to the other lead from the coil and the negative probe (black) to the negative lead (black) from the battery connector. This connection is temporary – just to read current – and the use of alligator clips may help (Table 3). Set the multimeter to record current (in amperes) and connect the battery to the circuit. Press the switch, hold for two seconds, and record the maximum current in amps. Repeat three times and note the reading each time in the “Constructing the Pulse Magnetizer” portion of the lab handout. Disconnect the multimeter and connect the negative battery lead (black) to the coil. Cover all connections with electrical tape.

Figure 1.

Wiring diagrams demonstrating how to build the pulse magnetizer circuit (A) with the ammeter/multimeter and (B) without the ammeter/multimeter. (C) An example circuit with the addition of a light-emitting diode (LED) and 470 Ω resistor that illuminates when the pulse magnetizer is switched on. The directions of current (solid arrow) and electron flow (dashed arrow) are shown in A.

Figure 1.

Wiring diagrams demonstrating how to build the pulse magnetizer circuit (A) with the ammeter/multimeter and (B) without the ammeter/multimeter. (C) An example circuit with the addition of a light-emitting diode (LED) and 470 Ω resistor that illuminates when the pulse magnetizer is switched on. The directions of current (solid arrow) and electron flow (dashed arrow) are shown in A.

Figure 2.

(A) Materials necessary for building and testing the pulse magnetizer, (BD) steps to wrap the coil for it, and (EF) how to verify its function. The arrows emphasize the change in direction of magnetic north before (E) and after (F) the pulse magnetizer is activated.

Figure 2.

(A) Materials necessary for building and testing the pulse magnetizer, (BD) steps to wrap the coil for it, and (EF) how to verify its function. The arrows emphasize the change in direction of magnetic north before (E) and after (F) the pulse magnetizer is activated.

To estimate the strength of the magnetic field inside the coil, use the equation (from Purcell, 1985, section 6.5)

$B=u0NLI$

where $B$ is the field strength in Tesla (T), $u0$ is the permeability of free space , $N$ is the number of turns (wraps) in the coil, $L$ is the length of the coil in meters (length of PVC pipe covered with wire coils), and $I$ is the current measured in amperes (A; average of the three measurements taken). Both $N$ and $L$ can be measured by counting the number of turns of wire and measuring the length of the coil (in meters) with a ruler, respectively. Students can record the estimated strength of their magnetic field coil in the “Constructing the Pulse Magnetizer” portion of the lab handout ( Appendix).

### Part 4: Verify the Pulse Magnetizer's Function

The goal is to identify the north (N) and south (S) poles of the electromagnet created when electric current passes through the coil. First, allow the compass needle to align with magnetic north and place one end of the coil facing the north compass point (Figure 2E). Press the switch and hold for two seconds. If the compass needle reverses direction (Figure 2F), label this end of the coil S, and the opposite end N. If the compass needle does not reverse, rotate the coil 180° and repeat the previous step.

### Part 5: Orientation Trials

Use the compass to align the circular orientation arena (see Online Materials) so the 0° label is facing magnetic north. Secure to the table with tape. Magnetic north may vary depending on the amount and location of iron used to construct the building, and this variation can be used as a discussion point for the class. Gently place the pill bug in the center of the arena (circle) and cover it with the cap from the tube. Allow the pill bug to rest for one minute. Remove the cap, start the stopwatch, and monitor the pill bug as it walks in the arena (Figure 3A). Mark the point where the pill bug first crosses the circle's boundary and record the angle to the nearest 10° tick mark (Figure 3B). If the pill bug does not cross the edge within 60 seconds, repeat the trial. Repeat the orientation procedure 10 times (or more if time permits) with the same pill bug, recording the angle for each trial. This is the “control” set.

Set up the pulse magnetizer so the N pole of the coil is facing up (Figure 3C). Place the pill bug inside the plastic tube and insert the tube inside the coil so the pill bug is approximately in the center (Figure 3C). Press the switch to activate the pulse magnetizer and hold for two seconds. Perform an orientation trial as described above. Repeat for a total of at least 10 trials. This is the “pulsed” group. The lab handout provided has areas for students to record their orientation angles and the various behaviors they see.

Figure 3.

(A) Performing an orientation trial with the pill bug and (B) recording the direction. The dashed line indicates the pill bug's movement, and the arrow represents how to record the angle (an example of 315° is shown). (C) Diagram representing how to expose the pill bug to the magnetic pulse. The blue lines and arrows portray the direction of the magnetic field (north should point upward).

Figure 3.

(A) Performing an orientation trial with the pill bug and (B) recording the direction. The dashed line indicates the pill bug's movement, and the arrow represents how to record the angle (an example of 315° is shown). (C) Diagram representing how to expose the pill bug to the magnetic pulse. The blue lines and arrows portray the direction of the magnetic field (north should point upward).

### Part 6: Data Analysis

Each group should plot their results and calculate the mean direction in both the control and pulsed trials. Furthermore, students can test for a preferred direction (nonrandom orientation, P < 0.05) using the Rayleigh test and for a difference between the control and pulsed trials (P < 0.05) using the Watson two-sample test (for statistical details, see Batschelet, 1981). A tutorial in R is provided in the Online Materials, including an example data set drawn from the magnetoreception literature. The tutorial can be completed by the students in advance of the lab activity if desired. The lab handout ( Appendix) has questions for the students to answer regarding the results of their statistical tests.

### Part 7: Conclusion & Comprehension

Following statistical analyses, students should answer the “Conclusion and Comprehension” questions in their lab handout, in which we ask students to talk to each other about the observations of pill bug behavior that they made during the lab, brainstorm what confounding variables may have resulted in different results between different groups, and suggest further experiments that they could perform. It may also be useful for the groups to combine their data and calculate the statistics for the class as a whole using the mean direction for each individual pill bug to avoid issues of pseudoreplication (see below). Lastly, students brainstorm ways in which an understanding of the magnetic sense might be useful to society, for example in developing advanced navigation technologies.

## Assessment

At the end of the activity, students should be able to

• articulate a basic understanding of the proposed mechanisms of magnetoreception and the ways in which animals use a magnetic sense,

• assess whether the control and pulse groups were oriented,

• determine whether the control and pulsed groups were different from each other,

• suggest other cues that may be influencing the pill bug's orientation, and

• outline other orientation experiments to test other possible cues.

The assessment can take place through a variety of assignments. The simplest is to have students complete the handout ( Appendix), but other techniques such as oral presentations, posters, and journal entries submitted to the instructor can be included.

## Results & Discussion

### Example Results

We present a set of experimental results obtained from six groups (16 students) in a 100-level undergraduate sensory biology course in which the students were either advanced high school students or college freshmen. The mean current measured from the pulse magnetizers was 3.5 A, which corresponded with a mean magnetic field strength of ~3.0 × 10−3 T (3 mT). To put this number in perspective, this field strength is approximately 50–100× greater than Earth's magnetic field (0.03–0.06 mT) and approaching that of an ordinary refrigerator magnet (5 mT). Five of the six groups found no evidence of orientation in their pill bug both before and after the magnetic pulse, and the mean directions did not differ. However, one group found significant orientation in their control trials, which also differed from their pulse-magnetized trials (Figure 4). This activity thus illustrates the difficult nature of performing behavior experiments and should motivate discussion between instructor and students regarding potential confounding cues and methods to control for them (see below).

Figure 4.

Example orientation data from a single pill bug before (control; black) and after (pulsed; gray) exposure to a pulsed magnetic field. Some points overlap. Arrows indicate mean direction within each group. The control trials were significantly oriented (Rayleigh test, P = 7.0 × 10−4) and differed from the pulsed trials (Watson test, 0.01 < P < 0.05).

Figure 4.

Example orientation data from a single pill bug before (control; black) and after (pulsed; gray) exposure to a pulsed magnetic field. Some points overlap. Arrows indicate mean direction within each group. The control trials were significantly oriented (Rayleigh test, P = 7.0 × 10−4) and differed from the pulsed trials (Watson test, 0.01 < P < 0.05).

### Confounding Factors, Pseudoreplication & Experimental Design

After data collection, the students engaged in lively discussion based on the questions provided in the “Conclusion and Comprehension” section of the handout ( Appendix). They proposed that confounding factors such as magnetic anomalies in the room due to the building's metal structure and electronic equipment may have affected the pill bugs’ orientation. Other examples of confounding variables not controlled included hunger level (a reasonable prediction is that hungrier individuals may be more inclined to searching behavior), “personality” (in many species, some individuals are consistently bolder than others and thus more willing to move), lighting, and temperature.

Because each group performs their data analysis independently, teachers can lead the class in combining all groups’ data in a class data set. This should begin with a short discussion about pseudoreplication. For example, the class could brainstorm what “pseudoreplication” means and how it might apply to this activity. Teachers must emphasize that, when the class combines their data, they will have to account for pseudoreplication. In particular, if each group has performed 10 trials with their pill bug, they will not submit 10 different values to the class data set, but rather only one: the mean angle. To illustrate how failure to account for pseudoreplication can change results, teachers could have students submit (a) all of their data and (b) only the mean, and have students calculate the Rayleigh and Watson tests separately for each scenario and compare the inferences made.

When discussing improvements to the experimental design, the students suggested performing the experiment several times in different places throughout the room, standardizing the way in which students placed the pill bugs in the center of the arena, and controlling for size differences between individuals. For collecting further data, they suggested performing more trials per group, using multiple pill bugs per group, and swapping pill bugs between groups to see if the same individual acted differently when tested by different experimenters. Lastly, they discussed how increased understanding of the magnetic sense may assist society in developing navigation technologies.

### Student Feedback

The results generated in this class did not provide conclusive evidence for the presence or absence of a magnetic sense in pill bugs. Nonetheless, throughout the experiment the students were focused on the given tasks and excited to be generating their own data. For many students, this was their first opportunity to handle “bugs,” and although initially concerned, they quickly became accustomed to the handling procedure. When contributing anonymous feedback for the course, half (8/16) of the students ranked the pill bug lab as their favorite activity. Reasons for why it was their favorite activity included descriptions based on its interdisciplinary nature, for example, “It includes coding, using a compass, electric circuit making, and testing actual bugs, which was very novel and fun,” and “I loved using R by ourselves to do the pill bug lab. It put what we had learned to good use”; whereas others enjoyed the process of discovery and lack of a predefined answer, such as “It's the most serious lab and so exciting to do a lab where we didn't know the result!” and “Because we got to do a whole experiment from start to finish, and it is really great to get our own results.”

### Implications for STEM Curricula

Interdisciplinary and active-learning exercises that address a variety of NGSS and AP concepts (see Tables 1 and 2), such as this pill bug activity, are in increasing demand to promote the addition and retention of undergraduate students in the science, technology, engineering, and mathematics (STEM) disciplines (PCAST STEM Undergraduate Working Group, 2012; Freeman et al., 2014). Furthermore, introductory-level activities in biophysics that apply evidence-based inquiry spanning scientific disciplines are effective tools for improving scientific literacy among non-STEM students (Parthasarathy, 2015).

An important component of the pill bug activity is to introduce students to new concepts in statistics, such as circular data analysis, while reinforcing existing knowledge (e.g., statistical testing, means). Although all calculations could be made by hand, the tutorials (see Online Materials) include basic training in data analysis using the free and open-source statistical software R (http://www.r-project.org). Until recently, quantitative skills in the sciences, notably the life sciences, had been deemphasized, thus resulting in the growing concern among professional and academic organization for improved training (Berlin & Lee, 2005; Feser et al., 2013). Evaluations from students enrolled in the sensory biology course demonstrated their enthusiasm for data analysis through such remarks as “… my favorite thing: using R,” “This is the first time I learn [sic] about the data analyzing software. It's cool!” and “It was cool seeing how my statistics background actually is being used.” As a result, the exposure of undergraduate STEM students to basic computer programming and statistical analysis (e.g., using R) through active-learning exercises like the pill bug lab will greatly contribute to the improvement of quantitative skills.

The pill bug lab presented here provides an excellent opportunity for students to integrate concepts from multiple disciplines into an active-learning exercise. Furthermore, the materials required for this lab are inexpensive and readily available online or in most laboratories. Depending on the instructor's goals and the students’ background, the experiment can easily be modified to assess additional species or individuals, to include more complex electrical concepts (e.g., capacitance and micro-controlled circuits), and/or to examine other cues aside from magnetic effects. The incorporation of interdisciplinary exercises like the pill bug activity will be a valuable contribution to any science curriculum and will further prepare students for careers in the STEM fields.

## Online Materials

A complete repository of information, tutorials, and full description of procedures is available at GitHub (https://github.com/rfitak/Circular_Biology). These include the basis for calculating a statistical mean by hand (https://github.com/rfitak/Circular_Biology/blob/master/Circular_data_by_hand.pdf) and how to use the software R (https://github.com/rfitak/Circular_Biology/blob/master/Circular_ data_exercise.md).

The pill bug orientation data presented here were collected during two class periods of Duke University summer session course Bio190S (Sensory Biology – Sight, Smell, Taste, Touch, Sound, and Beyond) taught by E.M.C. (instructor of record) and R.R.F. (guest lecturer). The Duke Office of Continuing Studies and Summer Session provided funding for materials. We also thank the student participants in the Bio190S course for their useful input on improving the activity.

## References

References
Altmann, J. (
1974
).
Observational study of behavior: sampling methods
.
Behaviour
,
49
,
227
267
.
Batschelet, E. (
1981
).
Circular Statistics in Biology
.
New York
:
.
Berlin, D.F. & Lee, H. (
2005
).
Integrating science and mathematics education: historical analysis
.
School Science and Mathematics
,
105
,
15
24
.
Ernst, D.A. & Lohmann, K.J. (
2016
).
Effect of magnetic pulses on Caribbean spiny lobsters: implications for magnetoreception
.
Journal of Experimental Biology
,
219
,
1827
1832
.
Feser, J., Vasaly, H. & Herrera, J. (
2013
).
On the edge of mathematics and biology integration: improving quantitative skills in undergraduate biology education
.
CBE Life Sciences Education
,
12
,
124
128
.
Freeman, S., Eddy, S.L., McDonough, M., Smith, M.K., Okoroafor, N., Jordt, H. & Wenderoth, M.P. (
2014
).
Active learning increases student performance in science, engineering, and mathematics
.
Proceedings of the National Academy of Sciences USA
,
111
,
8410
8415
.
Hurlbert, S.H. (
1984
).
Pseudoreplication and the design of ecological field experiments
.
Ecological Monographs
,
54
,
187
211
.
Irwin, W.P. & Lohmann, K.J. (
2005
).
Disruption of magnetic orientation in hatchling loggerhead sea turtles by pulsed magnetic fields
.
Journal of Comparative Physiology A
,
191
,
475
480
.
Johnsen, S. (
2017
).
Open Questions: we don't really know anything, do we? Open questions in sensory biology
.
BMC Biology
,
15
,
43
.
Johnsen, S. & Lohmann, K.J. (
2005
).
The physics and neurobiology of magnetoreception
.
Nature Reviews Neuroscience
,
6
,
703
712
.
Lee, A. (
2010
).
Circular data
.
Wiley Interdisciplinary Reviews: Computational Statistics
,
2
,
477
486
.
Lohmann, K.J. & Ernst, D.A. (
2014
). The geomagnetic sense of crustaceans and its use in orientation and navigation. In C. Derby and M. Thiel (Eds.),
Nervous Systems and Their Control of Behavior
(pp.
564
).
New York, NY
:
Oxford University Press
.
Parthasarathy, R. (
2015
).
‘The physics of life,’ an undergraduate general education biophysics course
.
Physics Education
,
50
,
358
366
.
(
2012
).
Engage to Excel: Producing One Million Additional College Graduates with Degrees in Science, Technology, Engineering, and Mathematics
.
Washington, DC
:
Office of the President
.
Purcell, E.M. (
1985
).
Electricity and Magnetism
.
Boston, MA
:
McGraw-Hill
.
Wiltschko, W. & Wiltschko, R. (
1995
).
Migratory orientation of European robins is affected by the wavelength of light as well as by a magnetic pulse
.
Journal of Comparative Physiology A
,
177
,
363
369
.

### Appendix

Pill Bug Magnetoreception Lab: Handout                NAME___________________________

Magnetoreception: Potential Mechanisms and Uses of the Magnetic Sense

Numerous behavioral experiments have shown that a variety of organisms, from bacteria to mammals, can sense Earth's magnetic field. This ability, called magnetoreception, is similar to how humans use a compass to figure out the direction they are moving. In addition to identifying a direction, some animals like sea turtles, can actually sense changes in the magnetic field strength and angle relative to the Earth's surface to navigate to specific locations. This is like having a natural, built-in GPS sensor! So, how do these animals sense the magnetic field? Unfortunately, the mechanism is not well known, but scientists have come up with a few ideas that are supported by their data: (1) a magnetic particle-based mechanism and (2) a chemical mechanism.

The magnetic particle mechanism predicts that animals have tiny crystals of magnetite, a naturally occurring mineral, in contact with various cells. These magnetite crystals act like little compass needles, so when they attempt to turn or rotate in response to Earth's magnetic field, they mechanically stress sensory cells, like hair cells or stretch receptors, or open and close ion channels in neurons to initiate a signal. The chemical mechanism is a bit more complicated. Certain proteins, like the visual pigment cryptochrome, have a free pair of electrons, or radical pair, after exposure to light. Each electron in this pair is spinning in a particular direction, so taken together the two electrons can be spinning the same direction (parallel) or different directions (antiparallel) – with different chemical reactivity in each state. Because the amount of time spent spinning either parallel or antiparallel depends on the angle of the ambient magnetic field, different chemical reactions can occur as the magnetic field changes. These reactions are thought to occur in the eyes of many migrating bird species.

How do scientists study magnetoreception in animals? The most common method used is an orientation experiment. The goal of an orientation experiment is to identify the direction an individual wants to move. Hopefully, by controlling all the possible cues an animal can use to move and manipulating only the magnetic field, scientists can determine the role of the magnetic field. Scientists often compare the directions of control individuals with individuals exposed to some kind of magnetic field treatment. In today's laboratory, we are going to expose pill bugs (a terrestrial isopod crustacean) to a magnetic pulse to see if the pulse affects their movement. To date, it is unknown whether pill bugs can sense a magnetic field, so your experiments will provide some of the first data to answer this question! You can read more about pill bugs, the experimental procedure, and data analysis at the link above. The worksheet below should help guide your thinking along the way and includes space for recording your data. After completing the lab, please submit your answers to the questions below. Have fun!

1. Briefly describe each of the two proposed mechanisms of magnetoreception:

2. Describe two ways in which animals might use their magnetic sense:

Circular vs. Linear Statistics

Examine the orientation data displayed below, along with the angles of orientation:

1. Draw an arrow that you think approximates the average angle of orientation.

2. Using linear statistics, calculate the average value of the angles, and draw an arrow that represents that value.

Average Value: ___________

3. How do the arrows you drew in questions 1 and 2 compare with one another? Why might they be different?

4. Now use circular statistics in R to calculate the average angle from the above data. Draw a third arrow that represents this value. How does this arrow compare to the arrow you drew in question 1?

Constructing the Pulse Magnetizer

1. Once you have built your pulse magnetizer, use the multimeter to record the maximum current it produces, in Amps. Repeat the measurement three times; record each measure here and then calculate the average current.

• Measurement 1__________

• Measurement 2__________

• Measurement 3__________

• Average current__________

2. Now use the numbers you recorded above to estimate the strength of the magnetic field inside the coil using the equation

$B=u0NLI$

$B$ is the field strength in Tesla (T)

$u0$ is the permeability of free space ()

$N$ is the number of turns (wraps) in the coil

$L$ is the length of the coil in meters (measure with a ruler)

$I$ is the current measured in Amperes (A; average of the three measurements taken)

B = _______________

Orientation Experiments

One of the key skills in the study of animal behavior is the ability to take detailed notes about what you observe. Practicing on simple behaviors like the orientation of your pill bug is a great way to develop this skill. Practice below by making at least three observations about the “control” behaviors you observe, and three observations about the “pulsed” behaviors you observe. These observations can be anything you think is important – be creative and observant!

1. Control Behaviors:

• Observation 1:

• Observation 2:

• Observation 3:

2. Pulsed Behaviors:

• Observation 1:

• Observation 2:

• Observation 3:

Statistical Data Analysis

Here is a table you can use to record your data and results.

TrialControl AnglesPulsed Angles

10
Mean Angle
Rayleigh Test P-value
TrialControl AnglesPulsed Angles

10
Mean Angle
Rayleigh Test P-value

Watson two-sample test P-value: ____________________

1. What can you conclude about orientation from the results of your Rayleigh test?

2. What can you conclude about orientation from the results of your Watson test?

Conclusion and Comprehension

1. Once you have finished your orientation trials and statistical analyses, find at least two other groups and compare your results with theirs. Are your results similar or different? How?

2. What are some external factors that you think might contribute to variation between groups? Why might some bugs in some groups act differently than other bugs in other groups?

3. Describe three ways that you might make this experiment more controlled, meaning reduce the variation between groups. (Hint: you can use the external factors you outlined in Question 2 for inspiration!)

4. Propose at least two follow-up experiments that you might do to learn more about magnetoreception in pill bugs, or another animal.

5. Brainstorm and describe one way that you think an increased understanding of the magnetic sense might be useful to human society.