Pillbug Behavior & Choice Chambers
Lab Overview. Pillbugs (woodlice, roly-polies) are terrestrial isopod crustaceans — descendants of marine ancestors that retained gill-like respiratory structures. Because their pleopods still need a moist film to exchange gases, pillbugs must continuously evaluate their environment and migrate toward conditions that preserve internal water balance. In this lab you will construct two-chamber arenas, place pillbugs at the boundary, and record their distribution at regular intervals. You will then test whether their behavior departs from a 50:50 random distribution using the chi-square goodness-of-fit test.
Learning Objectives. By the end of this lab you will be able to:
- Distinguish between taxis and kinesis and identify which is occurring in a data set.
- Design a controlled choice-chamber experiment with appropriate independent and dependent variables.
- Calculate a chi-square statistic, determine degrees of freedom, and interpret the result against a critical value.
- Connect arthropod cuticle and gas-exchange physiology to observed behavioral preferences.
- Construct and interpret bar graphs and contingency-style figures of behavioral data.
Vocabulary Cards
Tap any card to flip and reveal its definition. Only one card opens at a time, and each open card stays visible for 8 seconds before flipping back. You may reopen a card as many times as you need.
Tip: open every card at least once before attempting the matching exercise.
Vocabulary Matching
Click a term on the left, then click its matching definition on the right. Correct matches lock in green. Each correct match is worth 1 point.
Terms
Definitions
Reading: Behavioral Ecology of Terrestrial Isopods
Pillbugs occupy a curious evolutionary position. Although they belong to the crustacean order Isopoda — a group dominated by marine and freshwater species — the suborder Oniscidea abandoned aquatic life roughly 300 million years ago and colonized the leaf litter of every continent except Antarctica. The transition was incomplete. Unlike insects, which evolved tracheae lined with waterproof cuticle, isopods retained pleopods — flattened abdominal appendages that function as gills and require a thin film of water at all times. The dorsal cuticle of a pillbug is only weakly sclerotized and contains relatively little wax, so transpirational water loss across the body wall is among the highest of any terrestrial arthropod, often exceeding that of an earthworm of comparable mass. Survival on land therefore depends not on a physiological barrier to desiccation, but on a behavioral one: pillbugs must continuously seek microhabitats whose temperature, humidity, and substrate combine to slow water loss.
Behavior of this kind falls into two broad categories that AP Biology students are expected to distinguish. A taxis is a directional movement, oriented either toward (positive) or away from (negative) a stimulus gradient. A salmon swimming upstream toward the chemical signature of its natal river performs positive chemotaxis; a cockroach scuttling away from a flashlight performs negative phototaxis. A kinesis, by contrast, is non-directional: the organism changes the rate or turning frequency of its movement in response to a stimulus, without orienting along the gradient. The classic example is the woodlouse itself — in a dry petri dish a pillbug walks rapidly and turns infrequently, sweeping through space; in a humid dish it walks slowly and turns often, lingering in place. The end result resembles taxis: pillbugs accumulate in moist regions. But the underlying mechanism is mechanistically different and a careful experimenter must design controls capable of distinguishing the two.
The choice chamber as an experimental design
A choice chamber is among the simplest behavioral assays in biology. Two petri dishes are joined by a short bridge so that an animal can move freely between them; each dish is prepared with a different condition (for example, moist filter paper in one and dry filter paper in the other). A cohort of organisms — typically ten — is placed at the boundary and given several minutes to redistribute. The investigator then counts how many individuals occupy each side at fixed time intervals. Because the animal "chooses" repeatedly during the trial, this design generates a behavioral preference score that can be tested against a null hypothesis of random distribution.
The relevant null hypothesis (H₀) is that pillbugs distribute themselves 50:50 between the two chambers — that is, they show no preference. The alternative hypothesis (H₁) is that the observed distribution departs from 50:50 in a way unlikely to be due to chance. To decide between these, AP-level investigators apply the chi-square (χ²) goodness-of-fit test. The chi-square statistic is computed as Σ[(O − E)²/E], where O is the observed count in a category and E is the expected count under the null. With two categories (wet vs. dry, light vs. dark), there is one degree of freedom (df = k − 1 = 1), and the conventional critical value at α = 0.05 is 3.841. A computed χ² exceeding 3.841 lets us reject H₀: the pillbugs are not behaving randomly.
Why moisture matters mechanistically
The behavioral preferences pillbugs display are not arbitrary aesthetic choices; they are direct consequences of cuticular and respiratory physiology. Within minutes of exposure to dry air, an isopod begins losing water across its dorsal surface and through its still-wetted pleopods, and within an hour the cumulative loss may exceed 10% of body mass. Locomotor activity, which depends on hemolymph volume and ion balance, declines sharply. Selecting for moist substrate is therefore selecting for survival, and natural selection has favored individuals whose kineses and taxes reliably deliver them there. By contrast, the preference for darkness is partly secondary: dark sites tend to be moist and cool, so photonegative behavior is in part a proxy for hydrophilic behavior. Disentangling the two requires factorial designs — pairing wet/dry with light/dark in a 2 × 2 matrix — and is one of the analyses you will be asked to consider later in this lab.
Reading Comprehension
Answer each question in complete sentences using evidence from the reading. Each question is worth 1 point.
Sentence Reconstruction
The words below appear in scrambled order. Click each word in the order it belongs in the sentence. Correct positions will turn dark green. Each completed sentence is worth 1 point.
Simulation 1 — Wet vs. Dry Choice Chamber
Background. In this virtual choice chamber, ten pillbugs are released at the boundary between a dry chamber (left, sand-toned) and a wet chamber (right, moss-toned). Each pillbug performs a biased random walk in which its turning frequency and step size depend on local humidity. You will record the distribution at four time intervals and then test the distribution against the null hypothesis using chi-square.
Directions.
- Press Start Trial. The pillbugs will redistribute over four minutes of simulated time (the simulation runs faster than real time).
- At each prompt (every 30 simulated seconds), the counts for each side are auto-logged into your data table.
- After all four readings, complete the chi-square calculation in the table and answer the analysis questions.
- If you wish, you may press Reset to run a second trial — the second trial overwrites the first.
Hypothesis. If pillbugs prefer moist environments because their pleopods require a moisture film for gas exchange, then a significantly greater number of pillbugs should be observed in the wet chamber than the dry chamber after equilibration.
Choice Chamber Apparatus — REAL TIME · 2 MIN
Data Table 1 — Wet vs. Dry Distribution (4 points)
You count and record. The simulation auto-pauses at 30 s, 60 s, 90 s, and 120 s — count the bugs on each side of the dashed line at each pause and type the counts into the row below.
| Time (s) | Observed — Dry | Observed — Wet | Expected (50:50) |
|---|---|---|---|
| 30 | 5 / 5 | ||
| 60 | 5 / 5 | ||
| 90 | 5 / 5 | ||
| 120 | 5 / 5 | ||
| Final Total | — |
Chi-Square Calculation
The table below computes automatically once you enter your "Final Total" values above.
| Category | Observed (O) | Expected (E) | (O − E)² | (O − E)² / E |
|---|---|---|---|---|
| Dry | — | — | — | — |
| Wet | — | — | — | — |
| χ² Total = | — | |||
Critical value at df = 1, α = 0.05: 3.841 · Auto-conclusion: enter your data to compute
Analysis Questions
Simulation 2 — Light vs. Dark Choice Chamber
Background. In this trial both chambers contain identical moist filter paper, but one chamber is illuminated and the other is shaded. Because pillbugs may select dark sites partly because dark sites are typically moist (and partly because predators are visually oriented), removing the humidity variable lets us examine the independent contribution of light to the preference.
Directions. Same procedure as Simulation 1. Press Start Trial, allow the trial to run, and the counts will populate your data table at 30-second intervals.
Predicted outcome. If pillbugs respond independently to light intensity, the distribution will still depart significantly from 50:50 even with moisture controlled.
Choice Chamber Apparatus — REAL TIME · 2 MIN
Data Table 2 — Light vs. Dark Distribution (4 points)
You count and record. The simulation auto-pauses at 30 s, 60 s, 90 s, and 120 s — count the bugs on each side of the dashed line at each pause and type the counts into the row below.
| Time (s) | Observed — Light | Observed — Dark | Expected (50:50) |
|---|---|---|---|
| 30 | 5 / 5 | ||
| 60 | 5 / 5 | ||
| 90 | 5 / 5 | ||
| 120 | 5 / 5 | ||
| Final Total | — |
Chi-Square Calculation
| Category | Observed (O) | Expected (E) | (O − E)² | (O − E)² / E |
|---|---|---|---|---|
| Light | — | — | — | — |
| Dark | — | — | — | — |
| χ² Total = | — | |||
Critical value at df = 1, α = 0.05: 3.841 · Auto-conclusion: enter your data to compute
Analysis Questions
Case Study — The Vanishing Isopods of Pine Hollow
Dr. Marisol Vega, an ecologist studying invertebrate communities at Pine Hollow Preserve, has documented a 73% decline in the abundance of the native pillbug Armadillidium nasatum over a five-year period. The decline coincides with the conversion of two adjacent agricultural fields to soybean monoculture, but Dr. Vega is uncertain whether the cause is direct (pesticide exposure) or indirect (microclimate change in the leaf litter due to canopy thinning at the forest edge).
To investigate, she collects pillbugs from the disturbed forest edge (Population D) and from undisturbed interior forest (Population U) and runs both populations through four-chamber choice arenas at her field station. Each chamber contains one of four moisture levels: very dry (5% saturation), dry (25%), moist (75%), and saturated (95% — water film). She places 20 individuals per population in each trial and counts their distribution after 10 minutes. The results from her trials are summarized in the table below.
| Population | Very Dry | Dry | Moist | Saturated | Total |
|---|---|---|---|---|---|
| Undisturbed (U) | 1 | 3 | 11 | 5 | 20 |
| Disturbed (D) | 3 | 7 | 7 | 3 | 20 |
| Expected (random) | 5 | 5 | 5 | 5 | 20 |
Dr. Vega also samples the leaf litter from each population's source forest and finds that the disturbed forest has a litter moisture content of 28% by mass, while the undisturbed forest has 61%. She also assays both populations for the cytochrome P450 enzyme CYP6 — a detoxification enzyme that is typically upregulated in arthropods chronically exposed to insecticides — and finds CYP6 expression 4.2× higher in the disturbed population.
Graph & Data Analysis
The graph below shows mean pillbug count in the "preferred" chamber for four different environmental variables, replicated across 12 student lab groups (n = 12 trials per variable, 10 pillbugs per trial). Error bars represent ±1 standard error of the mean.
Figure 1. Mean pillbug count in the preferred chamber across four binary choice tests (n = 12 trials).
Free Response Questions (AP-Style)
Respond to each FRQ in clear, complete paragraphs. Each FRQ is worth 1 point. Use scientific terminology and reference specific evidence where required.
Question Bank — Mastery Quiz
Below are 8 randomly drawn multiple-choice questions from a bank of 24. Every student receives a different set of questions. Each question is worth 1 point. Select an answer for each question; correct answers will highlight green when you submit.
Final Grade Summary
Mr. Di's AP BIO · Lab Grade Report
Student: — · Period: — · Date: —