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Exploring Predatory Nematode Chemotaxis Using Low-Cost and Easy-to-UseMicrofluidicsAuthor(s): Matthew D. Stilwell, Julia F. Nepper, Elizabeth D. Clawson, Val Blair, Travis Tangen andDouglas B. WeibelSource: The American Biology Teacher, 79(9):753-762.Published By: National Association of Biology TeachersURL: http://www.bioone.org/doi/full/10.1525/abt.2017.79.9.753

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ABSTRACT

Symbiosis is a fascinating and diverse phenomenon. The study of symbiosis isimportant to understanding ecology, as it helps us understand relationshipsbetween organisms and provides insight into co-evolution, mutualism,adaptation, and survival. Ecological studies are challenging to implement inK-12 classrooms because they often require multiple organisms (often verydifferent in size) and complex environments that are difficult to replicateaccurately (e.g., soil composition, temperature, pH, and humidity). Thesefactors can make it difficult to study quantitative changes in ecosystems. Wedeveloped an inexpensive, quantitative experiment for classrooms that can beused to explore important aspects of microbial symbiosis, pathogenesis, andecology, and that helps support more investigations in this area of education.The experiment is low-cost, designed for K-12 teachers and students, usescommon materials, and teaches students about the exciting relationshipsamong bacteria, worms, and insects.

Key Words: microfluidics; symbiosis; low-cost; inexpensive; nematodes; bacteria;chemotaxis; ecology; active learning.

IntroductionSymbiosis—the interaction between differ-ent species—is found in every ecosystemstudied to date (Saffo, 2014). To fullyunderstand an ecosystem, one must lookat all of the interacting partners to seehow the actions of one species affectanother. Most well-known examples ofthese interactions—e.g., sea anemones andclown fish (Fautin, 1991; Miller, 2016),or oxpecker and rhinoceros (Mengesha,1978)—are difficult to bring into a class-room. Microbes, on the other hand, are usually inexpensive and easyto acquire and grow. However, the small size of microbes (typically1–50 microns) can make them difficult to study, especially when

microscopes are not available. Nematodes (roundworms) are largeenough to see with the naked eye (~1 mm) and are inexpensiveand easy to grow. They are also involved in several interesting inter-actions with other organisms, making them ideal for studyingsymbiosis.

Nematodes have adapted to almost every ecological niche onthe planet, and are so abundant that four out of five animals onthe planet are nematodes (Chen et al., 2004). Nematodes formmutualistic (mutually beneficial) relationships with bacteria toacquire nutrients. Although numerous species are animal or plantparasites, many nematodes and their bacterial symbionts are usefulfor insect pest control in agriculture, making them excellent modelorganisms for academic studies of symbiosis (Dillman & Sternberg,2012; Ehlers, 2001). For example, the soil-dwelling, entomopatho-genic (insect-eating) nematode Steinernema feltiae forms a symbioticrelationship with the bacterium Xenorhabdus bovienii, which helpsit kill and digest its insect prey (Hirao & Ehlers, 2009). S. feltiaenavigates through soil toward its prey by responding to concentra-tion gradients of chemical signals, or chemoeffectors, released byits insect prey in a process called chemotaxis (chemo as in chemical,taxis as in movement) (Hui & Webster, 2000).

It is easiest to study nematodes when theirmovement is confined. We use small microflui-dic channels to accomplish this. These channelsare shallow enough that their motion is essen-tially limited to two dimensions, making themeasier to observe (San-Miguel & Lu, 2013).Microfluidics experiments use very small vol-umes of fluids (down to 10−18 liters) in channelsthat have a width or height that is on the scale ofmicrometers (1 micrometer is one millionth of ameter) (Whitesides, 2006). Scientists and engi-neers use microfluidic technologies to reduce

the size, cost, and materials needed for experiments ranging fromchemical synthesis to DNA sequencing (Abate et al., 2013; Elviraet al., 2013; Feng et al., 2015). The constraint imposed by

Nematodes formmutualistic

(mutually beneficial)relationships withbacteria to acquire

nutrients.

The American Biology Teacher, Vol. 79, No 9, pages. 753–762, ISSN 0002-7685, electronic ISSN 1938-4211. © 2017 National Association of Biology Teachers. All rightsreserved. Please direct all requests for permission to photocopy or reproduce article content through the University of California Press’s Reprints and Permissions web page,www.ucpress.edu/journals.php?p=reprints. DOI: https://doi.org/10.1525/abt.2017.79.9.753.

THE AMERICAN BIOLOGY TEACHER PREDATORY NEMATODE CHEMOTAXIS 753

I N Q U I R Y &I N V E S T I G A T I O N

Exploring Predatory NematodeChemotaxis Using Low-Cost andEasy-to-Use Microfluidics

• MATTHEW D. STILWELL, JULIA F. NEPPER,ELIZABETH D. CLAWSON, VAL BLAIR,TRAVIS TANGEN, DOUGLAS B. WEIBEL

microfluidic channels creates a phenomenon referred to as laminarflow (see goo.gl/BxIHNf for a demonstration of this property). Fluidsexhibiting laminar flow move in “sheets,” which differs from theturbulent flow that is often observed in mixing fluids (Figure 1)(Falkovich, 2011). Laminar flow can be used to manipulate fluidsvery precisely (Whitesides, 2006).

Microfluidics is a powerful method to study chemotaxis, but tra-ditional methods for creating microfluidic channels are often veryexpensive and require resources that are not commonly available(Xia & Whitesides, 1998). We use a technique that requires onlyinexpensive office supplies (double-sided adhesive and transparencysheets) and a craft cutter (Clawson et al., 2018). Using these micro-fluidic channels, we demonstrate an easy way to measure the chemo-taxis of nematodes in response to various chemicals. Using thisapproach, students can ask whether nematodes swim toward oraway from different chemicals—including those released by insects.This gives them the tools to answer fundamental ecological questionsabout nematode behavior—in this case, how the nematode S. feltiaeidentifies and hunts insect prey.

The Nematode LifecycleNematodes can infect a wide range of hosts, including insects,plants, and mammals (Lee, 2002). Entomopathogenic nematodes,such as S. feltiae, can infect and consume a wide range of insects(Lacey & Georgis, 2012). S. feltiae has a mutualistic relationshipwith the bacterium Xenorhabdus bovienii, which they carry in spe-cialized pockets in their intestines (Hirao & Ehlers, 2009; Kimet al., 2012). S. feltiae senses chemical compounds produced andreleased by the insect prey and uses this information to track prey(Figure 2) (Hui & Webster, 2000). After catching its prey, the nem-atode burrows into natural openings in the insect such as itsmouth, anus, or spiracles (breathing openings) (Dowds & Peters,2002). Once inside, nematodes release their symbiotic bacteria,which multiply and produce toxins and other compounds that killand help digest the insect, which is then eaten by the nematodeand the bacteria to provide energy for survival and reproduction.

After the insect nutrients are depleted, the nematodes re-associatewith the bacteria, exit the insect cadaver, and traverse the soil insearch of their next insect prey (Figure 3) (Dowds & Peters, 2002).

Method DevelopmentMicroscopic organisms move differently from larger ones (Purcell,1977). Steinernema nematodes move by pushing their body againstsolid objects, which makes it difficult to observe nematodes swim-ming in water (Park et al., 2016). We filled channels with an agarsolution (agar powder is available online and at brick-and-mortarstores), which produces a transparent gel in which the worms canswim (Hida et al., 2015). To form the gel, the agar solution mustbe boiled, then allowed to cool. We found the nematodes couldsurvive brief exposure to the high temperature of the solution ofmelted agar, and after the gel set the nematodes could easily swimthrough the gel. The agar solution cools and solidifies rapidly, sofilling the channels may require a level of coordination that is toochallenging for elementary school students.

We designed a microfluidic system consisting of two straightinlet channels (with inlet holes) intersecting with a single centralchannel and ending in an outlet hole; the channel system looked likethe letter Y. We used three layers of material to form the channels(Figure 4): (1) a layer of double-sided adhesive tape with the chan-nels cut into it; (2) a top layer of transparency film forming the chan-nel ceiling with the shape and dimensions of the adhesive tape (thislayer contained the inlet and outlet holes); and (3) a bottom layer oftransparency film forming the channel floor with the shape of theadhesive tape but slightly longer to provide a handle. We used a craftcutter to cut the channels out of the double-sided adhesive and tocut the top and bottom layers out of transparency sheets. We assem-bled the system by pressing a transparency film layer on each side ofthe adhesive, then hot-gluing a small piece of Tygon tubing to oneinlet (Figure 4). A microscope slide can be used instead of transpar-ency film for the bottom layer of the channel if it makes visualizationeasier. For a very detailed description of the process for making themicrofluidic system, see our recently submitted paper in The Journalof Microbiology and Biology Education (Clawson et al.). A step-by-stepvideo can be found at https://youtu.be/BDFWlELvzJo.

When using the channels, we treated the tubing as a reservoir forthe chemoeffector, to ensure the channels would not become filledwith air. We filled the tubing reservoir with our chemoeffectorof interest while preparing for subsequent steps. (Note: If the tubingprevents the channel from fitting onto a microscope, the tubingcan be removed after filling the channels by pulling it off orusing scissors.) After soaking the nematode-containing sponge inwater, we mixed the nematode solution with the warm agar solution,pipetted it on the other inlet hole (without tubing), and drew thetwo solutions through the channel using a syringe positioned atthe outlet (Figure 5). To create a better seal, we put a small pieceof tubing on the tip of the syringe. After a few minutes, the channelwas moved to the microscope to visualize nematodes.

Our microfluidic chemotaxis system differs from those reportedpreviously in a seemingly small but important way: we do not loadchannels entirely with a solution of agar. Instead, we dissolve che-moeffectors in water (rather than in an agar solution) and use lami-nar flow to fill one half of the main channel with chemoeffector andone half with agar and nematodes. After the agar sets, half of thechannel contains the agar gel, and the other half of the channel

Figure 1. A depiction of laminar flow and turbulent flow.(A) Laminar flow causes fluids to move in sheets such thatthe fluids only mix by diffusion. (B) Turbulent flow causes fluidsto readily mix and is the state we commonly observe whenwatching liquids. A convenient way to adjust between laminarand turbulent flow is by adjusting the diameter of a channel, d.

THE AMERICAN BIOLOGY TEACHER VOLUME. 79, NO. 9, NOVEMBER/DECEMBER 2017754

contains the chemoeffector solution. There are two primary advan-tages to this approach. First, the channels are much easier for stu-dents to load when agar is added to only one of the two inletsbecause it cools and solidifies rapidly. We found this task to be diffi-cult, so younger students would certainly have trouble correctlyloading the channels. Our second reason for loading chemoeffectorsin water is that the nematodes swim well in the agar gel but not inthe liquid, so chemotaxing nematodes are essentially trapped if theymove to the chemoeffector side. This makes counting the wormseasier. We have not observed any nematodes swimming into thechemoeffector area (liquid side) when a repellant is present, so webelieve false positives are unlikely.

To test our system, we compared the nematodes’ responses toan attractant (waxworm extract), a repellant (acetic acid; i.e., white

vinegar), and water (a control). In each experiment, we observed4–12 nematodes in the main channel. (Note: The amount of waterused to soak the nematode sponge can be adjusted to increase ordecrease the number of worms suspended in liquid in each chan-nel.) To quantify the nematode response to each chemical, we cal-culated the chemotaxis index (CI) using the following equation:

CI ¼ # nematodes on chemoeffector side

total # of nematodes in channel

According to this equation, a perfect repellant will create a CI of 0,and a perfect attractant will yield a CI of 1. After performing theexperiment in triplicate, we found that the waxworm extractyielded a CI of ~0.71, the acetic acid yielded a CI of ~0.03, andthe water yielded a CI of ~0.14. (See Table 1 for individual

Figure 2. A cartoon depicting nematodes responding to chemical signals released by prey insects. Each prey insect producesand secretes multiple organic compounds that nematodes chemotax toward and use to hunt the insect. Wavy lines indicateproduction of volatile compounds. Examples of compounds shown are (from top left, clockwise): α-pinene, C10H16 (waxworm),dimethylsulfone, C2H6O2S (house crickets), furan, C4H8O (earwig), acetone, and C3H6O (pillbug). Note that although we depict onlyone compound for simplicity, insects release many organic compounds.

THE AMERICAN BIOLOGY TEACHER PREDATORY NEMATODE CHEMOTAXIS 755

Figure 3. A cartoon depicting the life cycle of nematodes. Nematodes traverse soil in search of insect prey, infect the insect, andsubsequently release their symbiotic bacteria to kill the insect. The nematodes and bacteria feed on the dead insect until thenutrients are depleted, at which point they re-associate and exit the insect cadaver in search of new prey.

Figure 4. The structure of the microfluidic channels and their assembly. (A) The components of a single channel. (B) Carefullyaligning the adhesive with the smaller piece of transparency. (C) After attaching the second transparency, the plastic tubing is hot-glued onto only one inlet. (D) The finished channel.

THE AMERICAN BIOLOGY TEACHER VOLUME. 79, NO. 9, NOVEMBER/DECEMBER 2017756

experimental values, or Figure 6 for a photo of the channels andquantification of the nematode response). (Note that ideally, as acontrol, water would yield a CI of 0.5). We found that adding foodcoloring to the chemoeffector solution greatly helped us visualizethe boundary between the agar and the chemoeffector, and madeit easier for us to see that both solutions were loaded into the

channel. We also observed that the laminar flow profile in channelscontaining both the warm agar solution and the waxworm extractwas not always evenly distributed, most likely due to the evapora-tion of the isopropanol in the waxworm extract. An occasionalwavy boundary formed between the two solutions, but will notaffect the outcome of the experiment.

Figure 5. How the experiments are performed. (A) An image depicting all of the materials required for this experiment.(B) The tubing at one inlet is filled with chemoeffector. (C) The agar is loaded into the other inlet. (D) The fluids are drawn intothe microfluidic channels using a tubing-tipped syringe. (E) An image of the laminar flow profile.

THE AMERICAN BIOLOGY TEACHER PREDATORY NEMATODE CHEMOTAXIS 757

Materials

• craft cutter (e.g., Cricut Explore One, $184.51; Silhouette Portrait,$149.99; or comparable craft cutter that allows uploading ofdesigns)

• Steinernema feltiae nematodes (e.g., Nemaglobe Fungus GnatControl Nematodes, available from Amazon.com)

• fresh waxworms (available at bait and pet stores or Amazon.com; can be frozen to maintain freshness)

• mortar and pestle (available at most cooking stores or Amazon.com)

• 70% isopropanol (available at drug stores and grocery stores)

• 5% acetic acid (white vinegar)

• agar powder (available on Amazon.com or at ethnic food stores)

• transparency film (available at office supply stores) (Write-onsheets are less expensive, but laser-compatible sheets are neces-sary for lined transparency guides.)

• double-sided adhesive (e.g., Elizabeth Craft Designs Clear Double-Sided Adhesive, 8.5 by 11 inch, available on Amazon.com)

• 3-mm hole punch (e.g., Cmxsevenday No.97C3 Metal Handheld1-Hole Metal Punch, 1/8" Hole Size, available on Amazon.com)(Note that most hole punches are 6 mm and thus are too large.)

• plastic tubing (3/16" inside diameter) (e.g., Tygon B-44-3 PVCBeverage Tubing)

• hot glue gun (available at craft stores)

• 1 mL syringes, Luer slip tip (e.g., Easy Glide 1cc Luer Slip TBSyringe) (Larger syringes will work, as well as Luer-Lok tips,but smaller syringes will be easier to work with.)

• food coloring (optional, but highly recommended)

• transfer pipettes with gradations (200 μL-1000 μL range) (e.g.,Karter Scientific Plastic Transfer Pipettes 1ml, Graduated), orvariable volume micropipette(s)

• water bath at 50–55°C (122–130°F) (A beaker of water on a hotplate will work.)

Table 1. Nematode chemotaxis results using the microfluidic channels.

Number of nematodes inagar side

Number of nematodes inchemoeffector side Chemotaxis Index

Waxworm extract

3 1 0.25

1 8 0.89

0 4 1.00

Acetic acid, 5%(white vinegar)

10 0 0

10 1 0.09

7 0 0

Water

9 2 0.18

6 0 0

10 3 0.23

Figure 6. (A) Five nematodes in a channel loaded withattractant (waxworm extract); the image was taken ~5 minutesafter loading the chemoattractant using an LG® G3 smartphoneand a platform microscope outfitted with the lens from alaser pointer (Yoshino, 2015). Arrows indicate live nematodes.(B) Chemotaxis indices calculated from three separateexperiments.

THE AMERICAN BIOLOGY TEACHER VOLUME. 79, NO. 9, NOVEMBER/DECEMBER 2017758

Methods

1. Assemble the microfluidic channels as described in Clawsonet al. (2018) and shown at https://youtu.be/BDFWlELvzJo.Cut the channels out of the double-sided adhesive, and cutthe top and bottom out of the transparency sheet. Removethe paper backing from the double-sided adhesive and attachto the smaller transparency. Ensure holes are cut out of thetransparency either by the craft cutter or manually using the3-mm hole punch. Remove the paper backing from the otherside of the adhesive and adhere to the larger transparency(Figure 4). Push all layers together from end to end to avoidcreases.

2. To create the chemoeffector reservoir and syringe adapter, cuttwo 0.5-0.75 cm pieces of tubing (we use Tygon PVC tubing,SAE, 3/16"; inside diameter, ¼" outside diameter, 1/32" wall).Hot glue one piece of tubing to one inlet to create a reservoirto hold the chemoeffector (Figure 4). Attach the other piece oftubing to the tip of the syringe to make pulling the solutions intothe channel easier. Use caution with hot glue to avoid burns.Avoid sealing the access holes to the microfluidic channel.

3. Prepare the chemoeffectors as outlined below:

• Attractant: Waxworm extract. Use the mortar and pestleto grind 5 frozen waxworms (freeze waxworms whilethey are fresh). Suspend the waxworm mash in 3 mL of70% isopropanol. Prepare a twenty-fold dilution (e.g.,1 mL mash + 19 mL water) from this solution usingwater. Add food coloring to visualize the liquid. (Note:If sacrificing insects is not acceptable for the experiment,chemically pure chemoeffectors can be purchased fromscientific supply companies.)

• Repellant: Acetic acid (vinegar). Make a solution of 5%acetic acid in water (or use white vinegar, which is ~5%acetic acid in water), and add a second color of food coloringto visualize the liquid.

• Control: Water. Add a third color of food coloring tovisualize the liquid.

Adding different colors to the solutions will make it easier tosee the laminar flow profile and clarify that the channel isworking as expected.

4. Suspend the nematode sponge in water. We resuspendedone sponge of nematodes in ~100 mL water, or half of asponge in ~50 mL water.

5. Melt a 2% agar solution and keep warm in a water bath(approximately 55°C). For ease of viewing, add a fourthcolor of food coloring.

6. For each group of students, prepare three microfluidicchannels—one each for attractant, repellant, and control—asoutlined above and in Clawson et al. (2018). Provide eachgroup of students with assembled microfluidic channels (orhave them assemble their own), along with the 1-mL syringeconnected to the extra piece of tubing and some sort of pipet.

7. One student loads a chemoeffector solution into the inlettubing (as in Figure 5b). Meanwhile, a second student mixesthe nematode suspension with the 2% agar solution in a3:1 ratio, so that the final agar concentration is 0.5% (e.g.,

0.5 mL of the nematode suspension with 1.5 mL of the 2%agar solution). Immediately after mixing, the second studentpipets the nematode-agar mixture onto the other inlet hole(as in Figure 5c). As soon as the nematode solution coversthe inlet hole, the first student quickly and gently draws thetwo solutions into the channel using the tubing-tipped syringeplaced at the outlet hole (as in Figure 5d). Note that the agarsolution cools quickly, so this step needs to be performedquickly.

8. Repeat step 7 for the other two microfluidic channels andconditions.

9. Image the channels using a microscope (Yoshino, 2015).Record the number of nematodes in each side of the channelover time (e.g., at 5, 10, 15 minutes). To quantify the nematoderesponse to the chemoeffector, use the following equation tocalculate the chemotaxis index (CI):

CI ¼ # nematodes on chemoeffector side

total # of nematodes in device

A chemotaxis index of 1 indicates that the chemical was astrong chemoattractant, and a value of 0 indicates a strongchemorepellant.

Note: The nematodes can be disposed of by rinsing them down asink or releasing them into soil outside. The used channels can beflushed with hot water for reuse or discarded.

In the FieldThe experiment was completed by 76 middle school students lackingformal biology training. The students experienced this investigation ina field trip setting at a science center near our lab. The teachersselected this workshop for their students from a number of other fieldtrip options. To gain insight into the activity’s impact on the students’enjoyment and engagement, we asked them to complete a shortsurvey after the workshop. The survey was based on the learning acti-vation surveys developed by Activation Lab (activationlab.org). Theframe of reference for this survey was on looking at evaluative meas-ures that would indicate how the activity impacted participants. Thesurveys were anonymous and optional. No identifying informationwas collected from the students, and all responses were aggregated.Based on these attributes, Institutional Review Board staff deemedthese evaluative measures to be exempt from IRB process.

We found this activity to be a rich platform for collaboration anda reflection of the practices we do as scientists. For brevity, we aggre-gated responses from several questions addressing a similar concept(collaboration with others) into a single measure (Figure 7a). Whenasked in anonymous surveys what they enjoyed most about theactivity, more than half the students said “everything” or “we learnedby doing things instead of listening the whole time,” while the rest“enjoyed being able to see the ways the nematodes reacted.” Weobserved students talking among themselves about what they weredoing and working together to do the activity successfully, in muchthe same way that academic scientists collaborate in the research lab.Additionally, over 90 percent of the students reported that theyasked questions, tried out new ideas, and discussed the experimentwith their mentors and peers to aid their comprehension (Figure 7a).

THE AMERICAN BIOLOGY TEACHER PREDATORY NEMATODE CHEMOTAXIS 759

These are all hallmarks of a collaborative process that the activity’sframework provided.

The activity provided opportunities for high levels of engagement.Of all student responses, 80–90 percent indicated that they wereactively engaged and self-reported that they had learned somethingabout science (Figure 7b). This activity can span engagement acrossage bands. We found this activity to be appropriate for students asyoung as 6th grade and can be adapted for students in high schooland college by exploring the key concepts more deeply.

The Role of Mutualistic Bacteria in NematodePredationEntomopathogenic nematodes form relationships with bacteria thatenable nematodes to infect and digest their insect prey. S. feltiaenematodes work with X. bovienii bacteria to obtain nutrients fromprey insects. To learn more about the mutualistic relationshipbetween X. bovienii and S. feltiae, students can create and study

the behavior of aposymbiotic nematodes (ASN)—worms that lackbacterial symbionts. For a detailed protocol on making ASNs, seeMcMullen and Stock (2014). To confirm that the ASNs do not con-tain Xenorhabdus, students can grind up nematodes and inoculatethem onto Luria-Bertani (LB) media plates as outlined by Sanders(2012) and observe bacterial growth (Sicard et al., 2006). Afterobtaining ASNs, students can compare the ability of aposymbioticworms to infect and kill prey insects with the ability of bacteria-colonized nematodes (as outlined in McMullen & Stock, 2014).

ConclusionsUsing simple, inexpensive, easy-to-make microfluidic channels, wedemonstrated how students can identify and study chemicals thatalter the behavior of predatory nematodes, such as the chemicalsthat nematodes use in nature to track their prey and avoid danger.An important aspect of this activity is the engagement of students

Figure 7. Student responses to survey questions regarding the activity. (A) Student responses to questions concerningcollaboration and exploration (N = 76). Students reported that they were actively checking their own understanding of theexperiment (N = 22). (B) Students were actively engaged in the experiment. The majority of students participated in the activitybecause they wanted to, not because they were instructed to (N = 76). Most of the students felt they learned something aboutscience (N = 22).

THE AMERICAN BIOLOGY TEACHER VOLUME. 79, NO. 9, NOVEMBER/DECEMBER 2017760

in the design of authentic scientific investigations. We often thinkof ecology on the scale of large animals and ecosystems; however,this activity reinforces the importance of ecosystems of small ani-mals and bacteria, and enables teachers to investigate a complexand important ecological web using tools that are widely accessibleand quantitative.

This activity can be expanded to fit within existing biology andenvironmental science curricula, as different aspects of symbiosisare reflected in this experiment: e.g., the mutualistic relationshipbetween the worms and their symbiotic bacteria, as well as the path-ogenic relationship between the worms and their insect prey. Theseconcepts could fit in a curriculum studying bacteria, pathogens,chemical ecology, soil science, and insects. This activity can be mod-ified to use different chemicals, temperatures, microfluidic channelshapes/designs, or species of nematode, and enables students tomake predictions about the response of nematodes to those condi-tions. We have provided a worksheet for students to fill out as theyperform the activity (Supplemental Material 1), as well as a work-sheet (Supplemental Material 2) and key with provided data forpractice analyzing data from the activity (Supplemental Material 3).

Microfluidics makes possible the investigations described herebecause it imposes dimensions on liquids that bring out the uniquelaminar behavior of fluids. Traditional methods for making micro-fluidic channels are tricky to incorporate into school activitiesbecause they require materials and facilities that are expensive anddifficult to access. Our method uses inexpensive office materials, acraft cutter to pattern the double-sided adhesive and transparencysheets, and simple methods for assembly and introducing fluids.The ease of this method enables students to create and test newchannel designs, and makes it possible to incorporate perspectiveson engineering into lessons.

Educators will be able to incorporate the Educators Evaluating theQuality of Instructional Products (EQuIP) Rubric to measure thealignment to the Next Generation Science Standards (NGSS) to helpdetermine which elements from these investigations connect to thescience and engineering practices (SEP), disciplinary core ideas(DCI), and/or crosscutting concepts (CCC) of NGSS. The embeddedtasks expected of students in these activities may demonstrate theirproficiency of one or more performance expectations. For example,students develop and use SEPs, DCIs, and CCCs that fit within theEQuIP three-dimensional framework. Teachers will need to identifythe key CCCs relevant to their instructional framework so thatstudents can translate specific information to general principles.

Looking deeper into the EQuIP (v3.0) rubric, there are inten-tional opportunities for students to explain phenomena and designsolutions that are integrated into life science DCIs (MS-LS2-1,2,4 &HS-LS2-8) and SEPs (MS/HS-ETS1). In particular, the evidencestatements reflected in NGSS HS-LS2-8 connect very well to thepotential for students to develop a causal explanation on the groupbehavior dynamics of chemotaxing nematodes in microfluidicchannels (NGSS, 2015). Students will use science and engineeringdesign practices in this activity and other potential investigationsof ecosystems and inter-organism interactions.

AcknowledgmentsWe acknowledge funding from the Dreyfus Foundation (SG-10-032), National Science Foundation (DMR-1121288, AISL 1241429,

and pre-doctoral fellowship DGE-1256259 to J.F.N.), WisconsinAlumni Research Foundaton (MSN193090), and Madison Commu-nity Foundation (5947) that made this research possible. We aregrateful to Heidi Goodrich-Blair and Mengyi Cao for introducingus to this fascinating area of biology.

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MATTHEW D. STILWELL is a Graduate Student in the Department ofBiochemistry, University of Wisconsin–Madison, 440 Henry Mall, Room6424, Madison, WI 53706; email: mstilwell@wisc.edu. JULIA F. NEPPER is aGraduate Student in the Department of Biochemistry, University ofWisconsin–Madison, 440 Henry Mall, Room 6424, Madison, WI 53706;email: jnepper@wisc.edu. ELIZABETH D. CLAWSON is a Discovery OutreachIntern at the Morgridge Institute for Research, 330 N. Orchard St., Madison,WI 53715; email: eclawson@morgridge.org. VAL BLAIR is an Education andOutreach Manager at the Morgridge Institute for Research, 330 N. OrchardSt., Madison, WI 53715; email: vblair@morgridge.org. TRAVIS TANGEN is anEducation and Outreach Manager at the Wisconsin Alumni ResearchFoundation, 614 Walnut St., Madison, WI 53726; email: ttangen@warf.org.DOUGLAS B. WEIBEL is a Professor in the Department of Biochemistry,University of Wisconsin–Madison, 440 Henry Mall, Room 6424, Madison,WI 53706; email: douglas.weibel@wisc.edu.

THE AMERICAN BIOLOGY TEACHER VOLUME. 79, NO. 9, NOVEMBER/DECEMBER 2017762