Name: measuring the flight ability of the ambrosia beetle, platypus quercivorus (murayama), using a low-cost, small, and easily constructed flight mill
Uploaded: 2018-08-06T13:00:00
Description: Watch this Scientific Journal Video about Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus Murayama, Using a Low-Cost, Small, and Easily Constructed Flight Mill at JoVE.com

We thank Mr. S. Fukaya, Mr. N. Okuda, and Mr. T. Ishino for helping with the experiments. This study was supported by Grants-in-Aid for Scientific Research from the Japan Society for Promotion of Science (No. 15K14755).

1. Construction of a Flight Mill
<ol>
	<li>Construction of a flight mill apparatus
	<ol>
		<li>Cut off the plastic part from a needle (metal part: 40 mm in length and 0.25 mm in diameter; plastic part: 22 mm in length and 2 mm in diameter) with nippers (Figure 3).</li>
		<li>Fix this needle with an untreated needle in the shape of a cross with epoxy resin adhesive (Figure 3), referring them as a flight mill arm and an axial needle. 
		NOT...

<ol>
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D., Dorn, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diel+flight+pattern+and+flight+performance+of+Cactoblastis+castorum+(Lepidoptera:+Pyralidae)+Measured+on+a+flight+mill:+influence+of+age,+gender,+mating+status,+and+body+size.">Diel flight pattern and flight performance of Cactoblastis castorum (Lepidoptera: Pyralidae) Measured on a flight mill: influence of age, gender, mating status, and body size.</a> J Econ Entomol. 101 (2), 314-324 (2008).</li><li>Zhang, Y., Wyckhuys, K. A. G., Asplen, M. K., Heinpel, G. E., Wu, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+Binodoxys+Communis+parasitism+on+flight+behavior+of+the+soybean+aphid,+Aphis+glycines.">Effect of Binodoxys Communis parasitism on flight behavior of the soybean aphid, Aphis glycines.</a> Biol Control. 62, 10-15 (2012).</li><li>Sappington, T. W., Burks, C. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Patterns+of+flight+behavior+and+capacity+of+unmated+navel+orangeworm+(Lepidoptera:+Pyralidae)+Adults+related+to+age,+gender,+and+wing+size.">Patterns of flight behavior and capacity of unmated navel orangeworm (Lepidoptera: Pyralidae) Adults related to age, gender, and wing size.</a> Environ Entomol. 43, 696-705 (2014).</li><li>Attisano, A., Murphy, J. T., Vickers, A., Moore, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+flight+mill+for+the+study+of+tethered+flight+in+insects.">A simple flight mill for the study of tethered flight in insects.</a> J. Vis. Exp. (106), e53377 (2015).</li><li>Evenden, M., Whitehouse, L., C, M., Sykes, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+influencing+flight+capacity+of+the+mountain+pine+beetle+(Coleoptera:+Curculionidae:+Scolytinae).">Factors influencing flight capacity of the mountain pine beetle (Coleoptera: Curculionidae: Scolytinae).</a> EnvironEntomol. 43, 187-196 (2014).</li><li>Fahrner, S. J., Lelito, J. P., Blaedow, K., Heimpel, G. E., Aukema, B. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+affecting+the+flight+capacity+of+Tetrastichus+planipennisi+(Hymenoptera:+Eulophidae),+a+classical+biological+control+agent+of+Agrilus+Planipennis+(Coleoptera:+Buprestidae).">Factors affecting the flight capacity of Tetrastichus planipennisi (Hymenoptera: Eulophidae), a classical biological control agent of Agrilus Planipennis (Coleoptera: Buprestidae).</a> Environ Entomol. 43, 1603-1612 (2014).</li><li>Pham, D. L., Ito, Y., Okada, R., Ikeno, H., Isagi, Y., Yamasaki, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phototactic+behavior+of+the+ambrosia+beetle+Phatypusquercirorus+(Murayama)+(Coleoptera:+Platypodidae)+before+and+after+flight.">Phototactic behavior of the ambrosia beetle Phatypusquercirorus (Murayama) (Coleoptera: Platypodidae) before and after flight.</a> J Insect Behav. 30, 318-330 (2017).</li><li>Wanner, H., Gu, H., Dorn, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nutritional+value+of+floral+nectar+sources+for+flight+in+the+parasitoid+wasp,+Cotesia+glomerata.">Nutritional value of floral nectar sources for flight in the parasitoid wasp, Cotesia glomerata.</a> Physiol Entomol. 31, 127-133 (2006).</li><li>Rowley, W. A., Graham, C. 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We developed a low-cost, easy-to-build, and compact flight mill for small insects such as P. quercivorus (4-5 mm in body length and 4-6 mg in body weight). Our flight mill comprised only ordinary items such as a needle, an IR LED, a photo sensor, instantaneous glue, etc., and did not require any sophisticated, expensive, or rare items such as computer-controlled electric devices. This enabled the easy and quick collection of necessary items and reduced experimental costs. Indeed, it cost only 1,000 JPY ...

Animals migrate long distances in search of food and mates. Migrating animals might sometimes carry undesirable companions. The female ambrosia beetle, Platypus quercivorus (Murayama), is a known vector of the fungal pathogen, Raffaelea quercivora Kubono et Shin-Ito. This pathogen causes mass mortality of Fagaceae trees (Japanese oak wilt) and a high level of mortality1. Since 1980, this disease has been expanding throughout Japan, and has become a serious problem2.
P. quercivorus is a small insect (4-5 mm in body length and 4-6 mg in body weight), and yearly expansion of the disease suggests that they are capable of flying up to several km3,4. The male P. quercivorus locates a host tree and releases an aggregation pheromone that attracts both males and females5. Consequently, the host tree is mass attacked by conspecifics, and eventually dies. The male bores a tunnel inside the tree after landing and a pheromone-attracted female enters the tunnel and lays eggs. The hatched P. quercivours grow in the tunnel until they become adults. Adults emerge and disperse to locate new hosts. Thus, expansion of the disease is possibly related to the migratory ability of this beetle. However, the extent to which the beetle can fly is still unclear. In addition, females are bigger than males6 (female: 4.6 mm, and male: 4.5 mm) and male beetles search for a target tree, enter the tunnel inside the tree, and then attract the female. Considering these sexual differences in the body size and role of flight in their life, sexual differences may exist in flight ability, but the difference in ability remains unclear.
In general, it is extremely difficult to measure migratory ability in the field, especially flight ability, due to the wide range of the migratory area. Migratory ability has been measured in laboratories under tethered conditions, such as a flight mill system, for over 60 years7,8,9,10,11,12,13. Flight mill systems have shown that some insects have the ability for long distance flight. For example, the longest flight distance of the mountain pine beetle in a flight mill was over 24 km14, and Tetrastichus planipennisi Yang flew maximally over 7 km15. Although the flight mill is a commonly available tool, biological assays with a living animal often result in considerably large individual differences. To overcome this, many measurements, repeated multiple times, are required to obtain reliable estimates of mean dispersal capacity. Therefore, multiple individuals should be used at the same time for the quick collection of a sufficient amount of data. However, simultaneous experiments require a larger space, multiple experimental setups, and are costlier when compared to a single measuring system. Hence, the flight mill must be low cost, should be easily built with commonly available items, and compact in size. Furthermore, the experimental procedure should not be complicated or need a skillful operator.
In this study, we assembled a small, low-cost flight mill (Figure 1 and Figure 2) that could be easily used in experimentation, and measured the flight ability of the ambrosia beetle, P. quercivorus.

<table>
	<tbody>
		<tr>
			<td>needle</td>
			<td>Seirin</td>
			<td>J type No. 5 x 40 mm</td>
			<td></td>
		</tr>
		<tr>
			<td>epoxy resin adhesive</td>
			<td>Konishi</td>
			<td>#16113</td>
			<td></td>
		</tr>
		<tr>
			<td>metal plate</td>
			<td></td>
			<td></td>
			<td>from a home improvement store</td>
		</tr>
		<tr>
			<td>disposable plastic pipette</td>
			<td></td>
			<td></td>
			<td>from a home improvement store</td>
		</tr>
		<tr>
			<td>snap button</td>
			<td></td>
			<td></td>
			<td>from a craft store</td>
		</tr>
		<tr>
			<td>IR sensor</td>
			<td>Hamamatsu Photonics</td>
			<td>S7136</td>
			<td></td>
		</tr>
		<tr>
			<td>IR LED</td>
			<td>OptoSupply</td>
			<td>OSIR5113A</td>
			<td>150 mW</td>
		</tr>
		<tr>
			<td>custom-made program</td>
			<td></td>
			<td></td>
			<td>downloadable from Github. 
			URL: https://github.com/HidetoshiIkeno/FlightMill</td>
		</tr>
		<tr>
			<td>instant glue</td>
			<td>Toagosei</td>
			<td>31204</td>
			<td></td>
		</tr>
		<tr>
			<td>A/D converter</td>
			<td>LabJack Co.</td>
			<td>U3-HV</td>
			<td></td>
		</tr>
		<tr>
			<td>DAQ software</td>
			<td>AzeoTech</td>
			<td>DAQFactoryExpress</td>
			<td>download from AzeoTech Web page.</td>
		</tr>
	</tbody>
</table>

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.

In these experiments, about 50% of the beetles applied to the flight mill showed one or more revolutions. When the plastic part passed a virtual line between the sensor and the LED, the recorded voltage changed from about 0 V to about 6.5 V, and the duration of a passing was within 10-20 ms, depending on the flight velocity. Therefore, a spike-like voltage change is observed as one revolution (Figure 7B). We defined flight as when the flight mill arm revolved...

Watch this Scientific Journal Video about Measuring the Flight Ability of the Ambrosia Beetle, Platypus Quercivorus Murayama, Using a Low-Cost, Small, and Easily Constructed Flight Mill at JoVE.com

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.

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). ...

This method can help answer key questions in the entomology and forensic fields such as control of the migration of pests. The main advantage of this technique is to easily measure migratory distance and speed of a small insect in a laboratory under a controlled condition. Begin the construction of the spinning portion of the flight mill.First, remove the plastic portion of an acupuncture needle. A 40 millimeter long metal needle with a 0.25 millimeter diameter. Then, using an epoxy adhesive, make a cross with the modified needle and an unmodified needle of the same size.This makes the flight mill arm and an axial needle. Next, prepare a small metal plate. Hammer a dimple into the center of the plate where the axial needle will attach.Then, secure the plate to a wooden base using adhesive tape. Next, make a plastic cylinder to hold the spinning axis of the mill. Cut a plastic pipette to make it the same length with the double-L plate, which has an outside diameter of 4 millimeter and an interior diameter of 3 millimeter.Then attach these parts to the base plate with the cylinder over the dimple and with the double-L plate stabilizing the cylinder. Now make a holder from an L-shaped piece of metal and screw the metal to a vertical wooden board to stabilize the assembly. Into the hole of another L plate, fit a small metal cap that is about 10 millimeters long and one millimeter in diameter.A snap button can work for this purpose. Now add an infrared photo sensor to count the revolutions. First, incorporate the IR sensor into the circuit board, and then, screw the circuit board into the L plate.The path of the plastic part of the crossed needles must pass directly below the sensor for the assembly to function. Next, secure a 150 milliwatt infrared LED attached to circuit substrate to a small magnet using glue. Then place the LED on the base plate beneath the photo sensor.Finally, connect the photo sensor to an analog input channel of an AD convertor. Next bundle all the cabling and fix the cables to the plate. Lastly, connect the AD convertor to a personal computer via a USB cable.On the morning of the experiment collect all of the freshly emerged P.Quercivorus adults from a dead Qercus crispula bloom tree. More than 100 beetles are commonly harvested. Next, put a beetle on ice for anesthetization.Avoid getting the beetle wet, or it will be difficult to complete the procedure. Now, using two component glue, put a dab of the jelly consistency component on the tip of the mill arm. Then attach the mill arm to the beetle's pronotum.Next, using a fine needle, mix a small amount of the liquid glue component into a jelly glue component. Do not get any glue on the beetle's wings. Once the glue is hardened, use a magnet on the top plate to slide the plates apart and easily reinsert the flight mill into the apparatus.Next, tweak the position of the infrared LED so that it is directly beneath the sensor. When the beetle awakens it will begin to fly. Then start recording the amplified output from the photo sensor at a sampling rate of 1000 hertz.As the mill spins with the beetle's flight the IR beam gets disrupted by the plastic piece leading to a voltage jump on the sensor from 0 to 3.7 volts. If the recording threshold exceeds 0.5 volts, the data is good for analysis. Using the software controls start the recording by clicking on the plus symbol on the logging icon in the workspace window.Then right-click on the log name and select Begin Logging Set"To stop the recording, right-click the log name and select End Logging Set"The data will be saved to a csv file. In these experiment about half of the beetles attached to the flight mill showed one or more revolutions. Sometime after being attached most beetles started flying for a certain period and then flew again after a rest interval.Each beetle was recorded for at least one hour. The average beetle flew at 0.75 to 1.5 meters per second. Any beetle that flew less than a kilometer was omitted from the analysis.For 16 beetles that met this criterion the minimum flight time was 1.26 hours without energy intake, and the longest time and distance was 7.5 hours and 27.1 kilometers. In this cohort, no significant difference was found between males and females. After watching this video you will quickly collect plenty of data because this flight mill is low-cost, small, and easily constructed.And not to much many flight mill. Since its development, this method has paved the way for research in the field of basic biology as well as pest management to explore new avenues of the physiology and ecology of small flying insects. By modifying this method, for example by installing an odor delivery system or illumination system, we can explore how sensory information influence the flying capacity.

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