I would like to thank Meredith Cenzer for purchasing all flight mill materials and providing continuous feedback from the construction to the write-up of the project. I also thank Ana Silberg for her contributions to standardize_troughs.py. Finally, I thank the Media Arts, Data, and Design Center (MADD) at the University of Chicago for permission to use its communal makerspace equipment, technology, and supplies free of charge.

<ol>
	<li>Krogh, A., Weis-Fogh, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Roundabout+for+studying+sustained+flight+of+locusts.">Roundabout for studying sustained flight of locusts.</a> Journal of Experimental Biology. 29, 211-219 (1952).</li><li>Hocking, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+intrinsic+range+and+speed+of+flight+of+insects.">The intrinsic range and speed of flight of insects.</a> Transactions of the Royal Entomological Society of London. 104 (8), 223 (1953).</li><li>Chambers, D. L., O'Connell, T. 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The author has nothing to disclose.

The simple, modern flight mill provides a range of advantages for researchers interested in studying tethered insect flight&#160;by delivering a reliable and automated design that tests multiple insects efficiently and cost-effectively13,31,35. Likewise, there is a strong incentive for researchers to adopt&#160;fast-emerging technologies and techniques from industry and other scientific fields as a means to build experimental to...

Given how intractable the dispersal of insects is in the field, the flight mill has become a common laboratory tool to address an important ecological phenomenon - how insects move. As a consequence, since the pioneers of the flight mill1,2,3,4 ushered in six decades of flight mill design and construction, there have been noticeable design shifts as technologies improved and became more integrated into scientific communities. Over time, automated data-collecting software replaced chart recorders, and flight mill arms transitioned from glass rods to carbon rods and steel tubing5. In the last decade alone, magnetic bearings replaced Teflon or glass bearings as optimally frictionless, and pairs between flight mill machinery and versatile technology have been proliferating as audio, visual, and layer fabrication technology become increasingly integrated into researchers&#39; workflows. These pairings have included high-speed video cameras to measure wing aerodynamics6, digital-to-analog boards to mimic sensory cues for studying auditory flight responses7, and 3D printing to make a calibration rig to track wing deformation during flight8. With the recent rise of emerging technologies at makerspaces, particularly at institutions with digital media centers run by knowledgeable staff9, there are greater possibilities to enhance the flight mill to test a larger range of insects and to transport the device to the field. There is also a high potential for researchers to cross disciplinary boundaries and accelerate technical learning through production-based work9,10,11,12. The flight mill presented here (adapted from Attisano and colleagues13) takes advantage of emerging technologies found in makerspaces to not only 1) create flight mill components whose scales and dimensions are fine-tuned to the project at hand but also 2) offer researchers an accessible protocol in laser cutting and 3D printing without demanding a high-budget or any specialized knowledge in computer-aided design (CAD).
The benefits of coupling new technologies and methods with the flight mill are substantial, but flight mills are also valuable stand-alone machines. Flight mills measure insect flight performance and are used to determine how flight speed, distance, or periodicity relates to environmental or ecological factors, such as temperature, relative humidity, season, host plant, body mass, morphological traits, age, and reproductive activity. Distinct from alternative methods like actographs, treadmills, and the video recording of flight movement in wind tunnels and indoor arenas14, the flight mill is notable for its ability to collect various flight performance statistics under laboratory conditions. This helps ecologists address important questions on flight dispersal, and it helps them progress in their discipline - whether that be integrated pest management15,16,17, population dynamics, genetics, biogeography, life-history strategies18, or phenotypic plasticity19,20,21,22. On the other hand, devices like high-speed cameras and actographs can require a strict, complicated, and expensive setup, but they can also lead to more fine-tuned movement parameters, such as wing-beat frequencies and insect photophase activity23,24. Thus, the flight mill presented here serves as a flexible, affordable, and customizable option for researchers to investigate flight behavior.
Likewise, the incentive to integrate emerging technologies into ecologists&#39; workflow continues to rise as questions and approaches to studying dispersal become more creative and complex. As locations that promote innovation, makerspaces draw in multiple levels of expertise and offer a low learning curve for users of any age to acquire new technical skills10,12. The iterative and collaborative nature of prototyping scientific devices in the makerspace and through online open sources can accelerate the application of theory11 and facilitate product development in the ecological sciences. Furthermore, increasing the reproducibility of scientific tools will encourage wider data collection and open science. This can help researchers standardize equipment or methods for measuring dispersal. Standardizing tools could further allow ecologists to unify dispersal data across populations in order to test metapopulation models that develop from dispersal kernels25 or source-sink colonization dynamics26. Much like how the medical community is adopting 3D printing for patient care and anatomy education27, ecologists can use&#160;laser cutters and 3D printers to redesign ecological tools and education and, within the scope of this study, can design additional flight mill components, such as landing platforms or a flight mill arm that can move vertically. In turn, the customization, cost-effectiveness, and increased productivity offered by makerspace technology can help start up dispersal projects with a relatively low barrier for researchers who intend to develop their own tools and devices.
To construct this flight mill, there are also mechanical and instrumental limitations that can be considered by the maker. Magnets and 3D printed enhancements allow the flight mill to be essentially glueless, except for the construction of the cross brackets, and to be accommodable to insects of different sizes. However, as the mass and the strength of insects increase, insects may be more likely to dismount themselves while tethered. Strong magnets can be used at the cost of increased torsional drag, or ball bearings can replace magnetic bearings as a robust solution for flight testing insects that weigh several grams28,29. Nevertheless, ball bearings can also present some problems, mainly that running prolonged experiments with high speeds and high temperatures can degrade the lubrication of ball bearings, which increases friction30. Thus, users will have to discern which flight mill mechanics would best suit their insect(s) of study and experimental design.
Similarly, there are several ways to instrument a flight mill that is beyond this paper&#39;s considerations. The flight mill presented here uses IR sensors to detect revolutions, WinDAQ software to record revolutions, and programming scripts to process the raw data. Although it is easy-to-use, the WinDAQ software has a limited array of tools available.&#160;Users cannot attach comments to their corresponding channel, and they cannot be alerted if any component of the circuitry fails. These cases are solved by detecting and correcting them through code but only after data collection. Alternatively, users can adopt more than one software that offers customizable data collection features28 or sensors that take direct speed and distance statistics, like bike milometers29. However, these alternatives can bypass valuable raw data or diffuse functionality across too many software applications, which can make data processing inefficient. Ultimately, rather than refashioning flight mill instrumentation, this protocol offers robust programming solutions to present-day software limitations.
In this paper, a design for an enhanced simple flight mill is described to aid researchers in their dispersal studies and to encourage the incorporation of emerging technologies in the field of behavioral ecology. This flight mill fits within the constraints of an incubator, holds up to eight insects simultaneously, and automates data collection and processing. Notably, its 3D printed enhancements allow the user to adjust the mill arm and IR sensor heights to test insects of various sizes and to disassemble the device for quick storage or transportation. Thanks to institutional access to a communal makerspace, all enhancements were free, and no additional costs were accrued compared to the simple, modern flight mill. All software needed are free, the electronic circuitry is simple, and all scripts can be modified to follow the specific needs of the experimental design. Moreover, coded diagnostics allow the user to check the integrity and precision of their recordings. Lastly, this protocol minimizes the stress sustained by an insect by magnetically painting and tethering&#160;insects to the mill arm. With the assembly of the simple flight mill being already accessible, affordable, and flexible, the use of makerspace technologies to enhance the simple flight mill can grant researchers the space to overcome their own specific flight study needs and can inspire creative flight mill designs beyond this paper&#39;s considerations.

<table><tbody><tr><td>180&amp;nbsp;&amp;Omega; Resistor</td><td>E-Projects</td><td>10EP514180R</td><td>Carbon film; stiff 24 gauge lead.</td></tr><tr><td>19 Gauge Non-Magnetic Hypodermic Steel Tubing</td><td>MicroGroup</td><td>304H19RW&amp;nbsp;</td><td></td></tr><tr><td>2.2 k&amp;Omega; Resistor</td><td>Adafruit</td><td>2782</td><td>Carbon film; stiff 24 gauge lead.</td></tr><tr><td>3D Printer</td><td>FlashForge</td><td>700355100638</td><td></td></tr><tr><td>3D Printer Filament</td><td>FlashForge</td><td>700355100638</td><td>Diameter 1.75 mm; 1kg/roll.</td></tr><tr><td>3D Printing Slicing Software</td><td>FlashPrint</td><td>4.4.0</td><td></td></tr><tr><td>Acrylic Plastic Sheets</td><td>Blick Art Supplies</td><td>28945-1006</td><td></td></tr><tr><td>Aluminum Foil</td><td>Target</td><td>253-01-0860</td><td></td></tr><tr><td>Breadboard Power Supply</td><td>HandsOn Tech</td><td>MDU1025</td><td>Can take 6.5V to 12V input and can produce 3.3V and 5V.</td></tr><tr><td>DI-1100 USB Data Logger</td><td>DATAQ Instruments</td><td>DI-1100</td><td>Has 4 differential armored analog inputs.</td></tr><tr><td>Electrical Wires</td><td>Striveday</td><td>B077HWS5XV</td><td>24 gauge solid wire.</td></tr><tr><td>Entomological Pins</td><td>BioQuip</td><td>1208S2</td><td>Size 2; diameter 0.45 mm.</td></tr><tr><td>Filtered 20 uL Pipette Tip</td><td>Fisher Scientific</td><td>21-402-550</td><td></td></tr><tr><td>Hot Glue Gun with Hot Glue</td><td>Joann Fabrics</td><td>17366956</td><td></td></tr><tr><td>IR Sensor</td><td>Adafruit</td><td>2167</td><td>This is the&amp;nbsp;3 mm IR&amp;nbsp;version; works up to 25 cm.</td></tr><tr><td>Large Clear Vinyl Tubing</td><td>Home Depot</td><td>T10007008</td><td>Inner diameter 3/8 in; outer diameter 1/2 in; length 20 ft.</td></tr><tr><td>Large Magnets</td><td>Bunting</td><td>EP654</td><td>Low-friction N42 neodymium; diameter 0.394 in; length 0.157 in; holding force 4.9 lb.&amp;nbsp;</td></tr><tr><td>Laser Cutter&amp;nbsp;</td><td>Universal Laser Systems&amp;nbsp;</td><td>PLS6.75</td><td></td></tr><tr><td>M5 Hex Nut</td><td>Home Depot</td><td>204274112</td><td>Thread pitch 0.8 mm; screw length 20 mm; diameter 5 mm.</td></tr><tr><td>M5 Long Iron Screws</td><td>Home Depot</td><td>204283784</td><td>Philips pan head; thread pitch 0.8 mm; screw length 20 mm; diameter 5 mm.</td></tr><tr><td>M5 Short Iron Screws</td><td>Home Depot</td><td>203540129</td><td>Philips pan head; thread pitch 0.8 mm; screw length 10 mm; diameter 5 mm.</td></tr><tr><td>Neoprene Rubber Sheet</td><td>Grainger</td><td>60DC16</td><td>Length 12 in; width 12 in; depth 1/8in.</td></tr><tr><td>Online 3D Modeling Software</td><td>Autodesk</td><td>2019_10_14</td><td>Tinkercad.com offers a free account.</td></tr><tr><td>Power Adaptor</td><td>Adafruit</td><td>63</td><td>9 VDC 1000mA regulated switching; input voltage DC 3.3V 5V.</td></tr><tr><td>Small Clear Vinyl Tubing</td><td>Home Depot</td><td>T10007005</td><td>Inner diameter 1/4 in; outer diameter 3/8 in; 20 ft long.</td></tr><tr><td>Small Magnets</td><td>Bunting</td><td>N42P120060</td><td>Low-friction N42 neodymium; diameter 0.120 in; length 0.060 in; holding force 0.5 lb.</td></tr><tr><td>Solderless MB-102 Breadboard&amp;nbsp;</td><td>Adafruit</td><td>239</td><td>830 tie points; length 17 cm; width 5.5 cm; input voltage, DC 3.3 V 5 V.</td></tr><tr><td>Sophisticated Finishes Iron Metallic Surfacer</td><td>Blick Art Supplies</td><td>27105-2584</td><td></td></tr><tr><td>Wire Cutters</td><td>Target</td><td>&amp;nbsp;84-031W</td><td></td></tr></tbody></table>

building an enhanced flight mill for the study of tethered insect flight

Makerspaces have a high potential of enabling researchers to develop new techniques and to work with novel species in ecological research. This protocol demonstrates how to take advantage of the technology found in makerspaces in order to build a more versatile flight mill for a relatively low cost. Given that this study extracted its&#160;prototype from flight mills built in the last decade, this protocol focuses more on outlining divergences made from the simple, modern flight mill. Previous studies have already shown how advantageous flight mills are to measuring flight parameters such as speed, distance, or periodicity. Such mills have allowed researchers to associate these parameters with morphological, physiological, or genetic factors. In addition to these advantages, this study discusses the benefits of using the technology in makerspaces, like 3D printers and laser cutters, in order to build a more flexible, sturdy, and collapsible flight mill design. Most notably, the 3D printed components of this design allow the user to test insects of various sizes by making the heights of the mill arm and infrared (IR) sensors adjustable. The 3D prints also enable the user to easily disassemble the machine for quick storage or transportation to the field. Moreover, this study makes greater use of magnets and magnetic paint to tether insects with minimal stress. Lastly, this protocol details a versatile analysis of flight data through computer scripts that efficiently separate and analyze differentiable flight trials within a single recording. Although more labor-intensive, applying the tools available in makerspaces and on online 3D modeling programs facilitates multidisciplinary and process-orientated practices and helps researchers avoid costly, premade products with narrowly adjustable dimensions. By taking advantage of the flexibility and reproducibility of technology in makerspaces, this protocol promotes creative flight mill design and inspires open science.

Flight data were obtained experimentally during Winter 2020 using field collected J. haematoloma from Florida as the model insects (Bernat, A. V. and Cenzer, M. L. , 2020, unpublished data). Representative flight trials were conducted in the Department of Ecology and Evolution at the University of Chicago, as shown below in Figure&#160;6, Figure 7, Figure 8, and&#160;Figure&#160;9. The flight m...

Watch this Scientific Journal Video about Building an Enhanced Flight Mill for the Study of Tethered Insect Flight at JoVE.com

Building an Enhanced Flight Mill for the Study of Tethered Insect Flight

This protocol uses three-dimensional (3D) printers and laser cutters found in makerspaces in order to create a more flexible flight mill design. By using this technology, researchers can reduce costs, enhance design flexibility, and generate reproducible work when constructing their flight mills for tethered insect flight studies.

Makerspaces have a high potential of enabling researchers to develop new techniques and to work with novel species in ecological research. This protocol ...

building-an-enhanced-flight-mill-for-study-tethered-insect

Universidad de Cartagena UDEC

Research

JoVE Journal

Engineering

2.8K Views.  University of Chicago. This protocol uses three-dimensional (3D) printers and laser cutters found in makerspaces in order to create a more flexible flight mill design. By using this technology, researchers can reduce costs, enhance design flexibility, and generate reproducible work when constructing their flight mills for tethered insect flight studies.

Video: Building an Enhanced Flight Mill for the Study of Tethered Insect Flight

Early Metamorphic Insertion Technology for Insect Flight Behavior Monitoring

Early Metamorphosis Insertion Technology (EMIT) is a novel methodology for integrating microfabricated neuromuscular recording and actuation platforms on insects during their metamorphic development. Here, the implants are fused within the structure and function of the neuromuscular system as a result of metamorphic tissue remaking. The implants emerge with the insect where the development of tissue around the electronics during pupal development results in a bioelectrically and biomechanically enhanced tissue interface. This relatively more reliable and stable interface would be beneficial for many researchers exploring the neural basis of the insect locomotion with alleviated traumatic effects caused during adult stage insertions. In this article, we implant our electrodes into the indirect flight muscles of Manduca sexta. Located in the dorsal-thorax, these main flight powering dorsoventral and dorsolongitudinal muscles actuate the wings and supply the mechanical power for up and down strokes. Relative contraction of these two muscle groups has been under investigation to explore how the yaw maneuver is neurophysiologically coordinated. To characterize the flight dynamics, insects are often tethered with wires and their flight is recorded with digital cameras. We also developed a novel way to tether Manduca sexta on a magnetically levitating frame where the insect is connected to a commercially available wireless neural amplifier. This set up can be used to limit the degree of freedom to yawing &#8220;only&#8221; while transmitting the related electromyography signals from dorsoventral and dorsolongitudinal muscle groups.

We present a novel surgical procedure to implant electrodes in Manduca sexta during its early metamorphic stages. This technique allows mechanically stable and electrically reliable coupling with the neuromuscular tissue to study flight neurophysiology dynamics. We also present a novel magnetic levitation platform for tethered studies of insect yaw.

Early Metamorphosis Insertion Technology (EMIT) is a novel methodology for integrating microfabricated neuromuscular recording and actuation platforms on ...

Behavior

Fabrication of Uniform Nanoscale Cavities via Silicon Direct Wafer Bonding

Measurements of the heat capacity and superfluid fraction of confined 4He have been performed near the lambda transition using lithographically patterned and bonded silicon wafers. Unlike confinements in porous materials often used for these types of experiments3, bonded wafers provide predesigned uniform spaces for confinement. The geometry of each cell is well known, which removes a large source of ambiguity in the interpretation of data.
Exceptionally flat, 5 cm diameter, 375 &#181;m thick Si wafers with about 1 &#181;m variation over the entire wafer can be obtained commercially (from Semiconductor Processing Company, for example). Thermal oxide is grown on the wafers to define the confinement dimension in the z-direction. A pattern is then etched in the oxide using lithographic techniques so as to create a desired enclosure upon bonding. A hole is drilled in one of the wafers (the top) to allow for the introduction of the liquid to be measured. The wafers are cleaned2 in RCA solutions and then put in a microclean chamber where they are rinsed with deionized water4. The wafers are bonded at RT and then annealed at ~1,100 &#176;C. This forms a strong and permanent bond. This process can be used to make uniform enclosures for measuring thermal and hydrodynamic properties of confined liquids from the nanometer to the micrometer scale.

A method for permanently bonding two silicon wafers so as to realize a uniform enclosure is described. This includes wafer preparation, cleaning, RT bonding, and annealing processes. The resulting bonded wafers (cells) have uniformity of enclosure ~1%1,2. The resulting geometry allows for measurements of confined liquids and gasses.

Measurements of the heat capacity and superfluid fraction of confined 4He have been performed near the lambda transition using lithographically patterned ...

Fast Imaging Technique to Study Drop Impact Dynamics of Non-Newtonian Fluids

In the field of fluid mechanics, many dynamical processes not only occur over a very short time interval but also require high spatial resolution for detailed observation, scenarios that make it challenging to observe with conventional imaging systems. One of these is the drop impact of liquids, which usually happens within one tenth of millisecond. To tackle this challenge, a fast imaging technique is introduced that combines a high-speed camera (capable of up to one million frames per second) with a macro lens with long working distance to bring the spatial resolution of the image down to 10 &#181;m/pixel. The imaging technique enables precise measurement of relevant fluid dynamic quantities, such as the flow field, the spreading distance and the splashing speed, from analysis of the recorded video. To demonstrate the capabilities of this visualization system, the impact dynamics when droplets of non-Newtonian fluids impinge on a flat hard surface are characterized. Two situations are considered: for oxidized liquid metal droplets we focus on the spreading behavior, and for densely packed suspensions we determine the onset of splashing. More generally, the combination of high temporal and spatial imaging resolution introduced here offers advantages for studying fast dynamics across a wide range of microscale phenomena.

Drop impact of non-Newtonian fluids is a complex process since different physical parameters influence the dynamics over a very short time (less than one tenth of a millisecond). A fast imaging technique is introduced in order to characterize the impact behaviors of different non-Newtonian fluids.

In the field of fluid mechanics, many dynamical processes not only occur over a very short time interval but also require high spatial resolution for ...

Scalable Nanohelices for Predictive Studies and Enhanced 3D Visualization

Spring-like materials are ubiquitous in nature and of interest in nanotechnology for energy harvesting, hydrogen storage, and biological sensing applications.&#160; For predictive simulations, it has become increasingly important to be able to model the structure of nanohelices accurately.&#160; To study the effect of local structure on the properties of these complex geometries one must develop realistic models.&#160; To date, software packages are rather limited in creating atomistic helical models.&#160; This work focuses on producing atomistic models of silica glass (SiO2) nanoribbons and nanosprings for molecular dynamics (MD) simulations. Using an MD model of &#8220;bulk&#8221; silica glass, two computational procedures to precisely create the shape of nanoribbons and nanosprings are presented.&#160; The first method employs the AWK programming language and open-source software to effectively carve various shapes of silica nanoribbons from the initial bulk model, using desired dimensions and parametric equations to define a helix.&#160; With this method, accurate atomistic silica nanoribbons can be generated for a range of pitch values and dimensions.&#160; The second method involves a more robust code which allows flexibility in modeling nanohelical structures.&#160; This approach utilizes a C++ code particularly written to implement pre-screening methods as well as the mathematical equations for a helix, resulting in greater precision and efficiency when creating nanospring models.&#160; Using these codes, well-defined and scalable nanoribbons and nanosprings suited for atomistic simulations can be effectively created.&#160; An added value in both open-source codes is that they can be adapted to reproduce different helical structures, independent of material.&#160; In addition, a MATLAB graphical user interface (GUI) is used to enhance learning through visualization and interaction for a general user with the atomistic helical structures.&#160; One application of these methods is the recent study of nanohelices via MD simulations for mechanical energy harvesting purposes.

Accurate modeling of nanohelical structures is important for predictive simulation studies leading to novel nanotechnology applications.&#160; Currently, software packages and codes are limited in creating atomistic helical models.&#160; We present two procedures designed to create atomistic nanohelical models for simulations, and a graphical interface to enhance research through visualization.

Spring-like materials are ubiquitous in nature and of interest in nanotechnology for energy harvesting, hydrogen storage, and biological sensing ...

Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus (Murayama), Using a Low-Cost, Small, and Easily Constructed Flight Mill

The ambrosia beetle, Platypus quercivorus (Murayama), is the vector of a fungal pathogen that causes mass mortality of Fagaceae trees (Japanese oak wilt). Therefore, knowing the dispersal capacity may help inform trapping/tree removal efforts to prevent this disease more effectively. In this study, we measured the flight velocity and duration and estimated the flight distance of the beetle using a newly developed flight mill. The flight mill is low cost, small, and constructed using commonly available items. Both the flight mill arm and its vertical axis comprise a thin needle. A beetle specimen is glued to one tip of the arm using instant glue. The other tip is thick due to being covered with plastic, thus it facilitates the detection of rotations of the arm. The revolution of the arm is detected by a photo sensor mounted on an infrared LED, and is indicated by a change in the output voltage when the arm passed above the LED. The photo sensor is connected to a personal computer and the output voltage data are stored at a sampling rate of 1 kHz. By conducting experiments using this flight mill, we found that P. quercivorus can fly at least 27 km. Because our flight mill comprises cheap and small ordinary items, many flight mills can be prepared and used simultaneously in a small laboratory space. This enables experimenters to obtain a sufficient amount of data within a short period.

Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus Murayama, Using a Low-Cost, Small, and Easily Constructed Flight Mill

We developed a low cost and small flight mill, constructed with commonly available items and easily used in experimentation. Using this apparatus, we measured the flight ability of an ambrosia beetle, Platypus quercivorus.

The ambrosia beetle, Platypus quercivorus (Murayama), is the vector of a fungal pathogen that causes mass mortality of Fagaceae trees (Japanese oak wilt). ...

Environment

A Magnetic Tether System to Investigate Visual and Olfactory Mediated Flight Control in Drosophila

It has been clear for many years that insects use visual cues to stabilize their heading in a wind stream. Many animals track odors carried in the wind. As such, visual stabilization of upwind tracking directly aids in odor tracking. But do olfactory signals directly influence visual tracking behavior independently from wind cues? Also, the recent deluge of research on the neurophysiology and neurobehavioral genetics of olfaction in Drosophila has motivated ever more technically sophisticated and quantitative behavioral assays. Here, we modified a magnetic tether system originally devised for vision experiments by equipping the arena with narrow laminar flow odor plumes. A fly is glued to a small steel pin and suspended in a magnetic field that enables it to yaw freely. Small diameter food odor plumes are directed downward over the fly s head, eliciting stable tracking by a hungry fly. Here we focus on the critical mechanics of tethering, aligning the magnets, devising the odor plume, and confirming stable odor tracking.

Here we describe how to tether a fly in an olfactory magnetic-tether (OMT) apparatus. We describe how to align the rare-earth magnets and odor ports, and how to set mass flow rates for both the stimulus delivery and vacuum suction to achieve optimal odor tracking.

It has been clear for many years that insects use visual cues to stabilize their heading in a wind stream. Many animals track odors carried in the wind. ...

Biology

A Wind Tunnel for Odor Mediated Insect Behavioural Assays

Olfaction is the most important sensory mechanism by which many insects interact with their environment and a wind tunnel is an excellent tool to study insect chemical ecology. Insects can locate point sources in a three-dimensional environment through the sensory interaction and sophisticated behavior. The quantification of this behavior is a key element in the development of new tools for pest control and decision support. A wind tunnel with a suitable flight section with laminar air flow, visual cues for in-flight feedback and a variety of options for the application of odors can be used to measure complex behaviour which subsequently may allow the identification of attractive or repellent odors, insect flight characteristics, visual-odor interactions and interactions between attractants and odors lingering as background odors in the environment. A wind tunnel holds the advantage of studying the odor mediated behavioural repertoire of an insect in a laboratory setting. Behavioural measures in a controlled setting provide the link between the insect physiology and field application. A wind tunnel must be a flexible tool and should easily support the changes to setup and hardware to fit different research questions. The major disadvantage to the wind tunnel setup described here, is the clean odor background which necessitates special attention when developing a synthetic volatile blend for field application.

Here, we describe the construction and use of a wind tunnel for odor mediated behavioural assays with insects. The wind tunnel design facilitates the release of odor sources by several methods, with and without visual stimuli. Wind tunnel experiments are important methods to identify behaviorally active volatile chemicals.

Olfaction is the most important sensory mechanism by which many insects interact with their environment and a wind tunnel is an excellent tool to study ...

Using Flight Mills to Measure Flight Propensity and Performance of Western Corn Rootworm, Diabrotica virgifera virgifera (LeConte)

The western corn rootworm, Diabrotica virgifera virgifera (LeConte) (Coleoptera: Chrysomelidae), is an economically important pest of corn in the northern United States. Some populations have developed resistance to management strategies including transgenic corn that produces insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt). Knowledge of western corn rootworm dispersal is of critical importance for models of resistance evolution, spread, and mitigation. Flight behavior of an insect, especially over a long distance, is inherently difficult to observe and characterize. Flight mills provide a means to directly test developmental and physiological impacts and consequences of flight in the laboratory that cannot be obtained in field studies. In this study, flight mills were used to measure the timing of flight activity, total number of flights, and the distance, duration, and speed of flights taken by female rootworms during a 22-h test period. Sixteen flight mills were housed in an environmental chamber with programmable lighting, temperature, and humidity control. The flight mill described is of a typical design, where a flight arm is free to rotate about a central pivot. Rotation is caused by flight of an insect tethered to one end of the flight arm, and each rotation is recorded by a sensor with a time-stamp. Raw data are compiled by software, which are subsequently processed to provide summary statistics for flight parameters of interest. The most difficult task for any flight mill study is attachment of the tether to the insect with an adhesive, and the method used must be tailored to each species. The attachment must be strong enough to hold the insect in a rigid orientation and to prevent detachment during movement, while not interfering with natural wing motion during flight. The attachment process requires dexterity, finesse, and speed, making video footage of the process for rootworms of value.

Using Flight Mills to Measure Flight Propensity and Performance of Western Corn Rootworm, Diabrotica virgifera virgifera LeConte

Flight mills are important tools for comparing how age, sex, mating status, temperature, or various other factors may influence an insect&#8217;s flight behavior. Here we describe protocols to tether and measure the flight propensity and performance of western corn rootworm under different treatments.

The western corn rootworm, Diabrotica virgifera virgifera (LeConte) (Coleoptera: Chrysomelidae), is an economically important pest of corn in the northern ...

A Simple Flight Mill for the Study of Tethered Flight in Insects

Flight in insects can be long-range migratory flights, intermediate-range dispersal flights, or short-range host-seeking flights. Previous studies have shown that flight mills are valuable tools for the experimental study of insect flight behavior, allowing researchers to examine how factors such as age, host plants, or population source can influence an insects' propensity to disperse. Flight mills allow researchers to measure components of flight such as speed and distance flown. Lack of detailed information about how to build such a device can make their construction appear to be prohibitively complex. We present a simple and relatively inexpensive flight mill for the study of tethered flight in insects. Experimental insects can be tethered with non-toxic adhesives and revolve around an axis by means of a very low friction magnetic bearing. The mill is designed for the study of flight in controlled conditions as it can be used inside an incubator or environmental chamber. The strongest points are the very simple electronic circuitry, the design that allows sixteen insects to fly simultaneously allowing the collection and analysis of a large number of samples in a short time and the potential to use the device in a very limited workspace. This design is extremely flexible, and we have adjusted the mill to accommodate different species of insects of various sizes.

Flight in insects is influenced by a number of factors and the propensity to disperse is an important variable in understanding insect ecology and biological control strategies. We describe the construction and use of a simple, relatively inexpensive, and flexible flight mill for measuring parameters of tethered flight in insects.

Flight in insects can be long-range migratory flights, intermediate-range dispersal flights, or short-range host-seeking flights. Previous studies have ...

Neuroscience

Insect-machine Hybrid System: Remote Radio Control of a Freely Flying Beetle (Mercynorrhina torquata)

The rise of radio-enabled digital electronic devices has prompted the use of small wireless neuromuscular recorders and stimulators for studying in-flight insect behavior. This technology enables the development of an insect-machine hybrid system using a living insect platform described in this protocol. Moreover, this protocol presents the system configuration and free flight experimental procedures for evaluating the function of the flight muscles in an untethered insect. For demonstration, we targeted the third axillary sclerite (3Ax) muscle to control and achieve left or right turning of a flying beetle. A thin silver wire electrode was implanted on the 3Ax muscle on each side of the beetle. These were connected to the outputs of a wireless backpack (i.e., a neuromuscular electrical stimulator) mounted on the pronotum of the beetle. The muscle was stimulated in free flight by alternating the stimulation side (left or right) or varying the stimulation frequency. The beetle turned to the ipsilateral side when the muscle was stimulated and exhibited a graded response to an increasing frequency. The implantation process and volume calibration of the 3 dimensional motion capture camera system need to be carried out with care to avoid damaging the muscle and losing track of the marker, respectively. This method is highly beneficial to study insect flight, as it helps to reveal the functions of the flight muscle of interest in free flight.

Insect-machine Hybrid System: Remote Radio Control of a Freely Flying Beetle Mercynorrhina torquata

This protocol describes the process of constructing an insect-machine hybrid system and carrying out wireless electrical stimulation of the flight muscles required to control the turning motion of a flying insect.

The rise of radio-enabled digital electronic devices has prompted the use of small wireless neuromuscular recorders and stimulators for studying in-flight ...