DTIC ADA569798: Aspirator Gun for High-Throughput Mosquito Bioassays

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by  Defense Technical Information Center

Aspirator Gun for High-Throughput Mosquito Bioassays
Author(s): Robert L. Aldridge, W. Wayne Wynn, Seth C. Britch, and Kenneth J.
Linthicum
Source: Journal of the American Mosquito Control Association, 28(1):65-68.
2012.
Published By: The American Mosquito Control Association
DOI: http://dx.doi.org/10.2987/11-6195.1
URL: http://www.bioone.org/doi/full/10.2987/11-6195.1
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1. REPORT DATE 3. DATES COVERED
2012 2. REPORT TYPE 00-00-2012 to 00-00-2012
4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER
Aspirator Gun for High-Throughput Mosquito Bioassays
5b. GRANT NUMBER
5c. PROGRAM ELEMENT NUMBER
6. AUTHOR(S) 5d. PROJECT NUMBER
5e. TASK NUMBER
5f. WORK UNIT NUMBER
7. PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) 8. PERFORMING ORGANIZATION
US Department of Agriculture,Agricultural Research Service,1600 SW REPORT NUMBER
23rd Drive,Gainesville,FL,32608
9. SPONSORING/MONITORING AGENCY NAME(S) AND ADDRESS(ES) 10. SPONSOR/MONITOR’S ACRONYM(S)
11. SPONSOR/MONITOR’S REPORT
NUMBER(S)
12. DISTRIBUTION/AVAILABILITY STATEMENT
Approved for public release; distribution unlimited
13. SUPPLEMENTARY NOTES
14. ABSTRACT
We describe an innovative aspirator gun designed to transfer individual anesthetized mosquitoes directly
into glass bioassay tubes. The gun has been used for thousands of transfers with extremely low associated
mortality and is the central component of a high-throughput bioassay system. The gun is constructed using
readily obtainable materials and can be modified for a range of insects.
15. SUBJECT TERMS
16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF 18. NUMBER 19a. NAME OF
ABSTRACT OF PAGES RESPONSIBLE PERSON
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Prescribed by ANSI Std Z39-18
JournaloftheAmericanMosquitoControlAssociation,28(1):65–68,2012
CopyrightE2012byTheAmericanMosquitoControlAssociation,Inc.
OPERATIONAL NOTE
ASPIRATOR GUN FOR HIGH-THROUGHPUT MOSQUITO BIOASSAYS
ROBERTL.ALDRIDGE,W.WAYNEWYNN,SETHC.BRITCHANDKENNETHJ.LINTHICUM
USDepartmentofAgriculture,AgriculturalResearchService,CenterforMedical,Agricultural,andVeterinary
Entomology,1600SW23rdDrive,Gainesville,FL32608
ABSTRACT. We describe an innovative aspirator gun designed to transfer individual anesthetized
mosquitoes directly into glass bioassay tubes. The gun has been used for thousands of transfers with
extremelylowassociatedmortalityandisthecentralcomponentofahigh-throughputbioassaysystem.The
gunisconstructedusingreadilyobtainablematerialsandcanbemodifiedforarangeofinsects.
KEYWORDS Mosquito,arthropod,insect,aspirator,suction
Aspiratorsareindispensabletoolsforconduct- Recent work using fine insect forceps to
ingfieldandlaboratoryresearchwithmosquitoes transfer anesthetized adult female mosquitoes
(Peterson 1953, Fergusson 1982, Arnold 1994, fromcoloniestobioassaychambers(Britchetal.
Tang1996,Toth2000).Aspiratorswereoriginal- 2009,2010)highlightedtheneedforanimproved
ly developed to use mouth suction (Psota 1916); aspirator design. Having observed damage to
however, aspirators using powered suction, such mosquitoes after transfer, and having found that
as with bulbs or bellows (Wishart 1930, Brophy forceps transfer was slow and not keeping up
et al. 1982, Toth 2002), electric- or gas-powered with the period of anesthesia, we specifically
fans (Nelson and Chamberlain 1955, Dietrick designed an aspirator to rapidly and gently pick
1961, Murphey and Darsie 1962, Clark et al. up and deposit females directly into bioassay
1994), or laboratory bench vacuum (Schwartz tubes.WebuiltuponChaoandPeterson’s(1952)
1964), help to protect workers from harmful T-aspirator model to develop an aspirator gun
inhalation of dust, insect parts, or pathogens (the ‘‘Wynn Gun’’; Fig. 1) with modern, easily
(Hurd1954,Douglas1984,KimandHong2007). procuredmaterials,andadditionalinnovationsto
Powered aspirators also substantially reduce overcome challenges imposed by the mosquito
worker fatigue and error, increase speed and body plan.
efficiency, and minimize harm to living insect Mosquitoes are soft bodied with fine wings
samples. Early aspirator designs placed the and scales that are subject to sticky electrostatic
collecting tube in-line with the suction (e.g., forces. Mosquito bodies are also sensitive to
Kunkel 1926), but this required the unit to be air flow because their mass is diffused over air-
takenaparttoaccesssamples—apoordesignfor catching structures such as long legs, antennae,
flyinginsects.Also,asinsectsweredrawnintothe wings,andproboscis.Ininitialtrialswithabasic
collecting tube they would strike and be forced T-aspirator, we learned that electrostatic adhe-
against the protective mesh at the opposite end, sion frequently prevented mosquitoes from com-
often causing injuries and damage. pleting the path to the bottom of the bioassay
These shortcomings were soon resolved with vial. We also learned that turbulence in the
thedevelopment ofthe‘‘pooter’’-styleaspirator bioassay tube could cause some mosquitoes to
(Poos1929),wherethe collectingvialwasoffset flushstraightthroughthedeviceandoutintothe
atarightangletotheflowofairsuchthatinsects vacuummotor.Asaresultweaddedanantistatic
were deposited directly into alcohol (Singer grounding system to the pick-up tube and an
1964),orintoasmallcage(Peterson1953:Plate airflow baffle to the rear of the expansion
166) or vial (Matteson 1965) that could be re- chamberwithinthemainbodyassembly(Fig. 1).
movedandquicklycapped.Thislastfeaturewas The main body of the Wynn Gun is impact-
especially attractive for use in bioassay opera- resistant, rigid polyvinyl chloride (PVC) that can
tions if the removable vial was in fact the be easily modified with couplers to hold a range
bioassay chamber. Using an innovative bent of bioassay collection tubes. Similarly, the pick-
pick-up tube, Chao and Peterson (1952) de- up tube can be replaced with a range of nozzle
signed a T-shaped aspirator that efficiently diameterstoaccommodatearangeofinsectsizes.
deposited insects into removable bioassay vials. The separate vacuum blower can be changed to
Ristich and Lockard (1953) quickly followed produce a range of suction forces, and the air
with a set of quick-change adapters that would exhaustcanbefilteredtoprotectairqualityinthe
permittheT-aspiratortobeusedwitharangeof lab. The Wynn Gun was used to aspirate Culex
collectingvial sizes. quinquefasciatus Say mosquitoes (1995 USDA
65
66 JOURNALOFTHEAMERICANMOSQUITOCONTROLASSOCIATION VOL.28,NO.1
Fig.1. Cross-sectionaldetail ofWynnGunaspira-
torbodyandpick-uptube.Numbersarekeyedtoparts
listedinTable1.
Fig.2. ExteriorphotographofWynnGunaspirator
with bioassay tube and vacuum hose attached. Num-
strain, Gainesville, FL) at approximately 60 bersarekeyedtopartslistedinTable1.
transfers/min for 360 bioassays of 10 females
each in Britch et al. (2011), with a mean of 1% theWynnGunisdividedinto4maincomponents
mortality in controls, demonstrating that both for simplicity and efficient assembly. These com-
the device and the bioassay system are safe for ponents are: the pick-up tube, the main body
adult mosquito transfer. assembly, the electrostatic groundingsystem, and
Identificationofpartsandsupplies(inTable 1) theindependentvacuumsource.Thepick-uptube
is keyed to part numbers in the Wynn Gun aspi- component is constructed first. It is made up of
ratorwithpick-uptube(Fig. 1)andwithbioassay brass and acrylic tubing and allows the operator
tubes and vacuum hose (Fig. 2). Construction of to collect individual insects from a surface. Thin-
Table1. PartslistofWynnGunaspiratorassemblyaskeyedtopartnumbersinFigs.1and2.
Partno. Product/part1 Dimensions2 Quantity Source3
1 Tubing,thin-walledbrass 8-mmOD 2.7in.(6.75cm) H,O
2 Fisherbrandrubberpipettebulb4 J-in.ID,3-mlcapacity;1Hin.long 1 S,O
3 Straightcoupling,CPVC5 S/S,Kin.(Kin.) 2 H,O
4 Tubing,clearacrylic K-in.ID,H-in.OD 5.5in.(13.75cm) H,O
5 L-coupling,CPVC45uangle S/S,Kin.(Kin.) 1 H,O
6 Tubing,clearacrylic K-in.ID,H-in.OD 4.15in.(10.37cm) H,O
7 Reducerbushing6 Sch.40,1Jin.3Kin. 1 H,O
8 Tubing,clearacrylic7 K-in.ID,H-in.OD 1.75in.(4.37cm) H,O
9 L-coupling,CPVC90uangle8 S/S,Kin.(Kin.) 1 H,O
10 Pipe,PVCthin-walled 1in. 2in.(5cm) H,O
11 Reducerbushing9 Sch.40,1Jin.3Lin. 1 H,O
12 T-coupling,PVC10 Sch.40,S/S/S,1Jin.31Jin.31Jin. 1 H,O
13 Coppertape,adhesiveback Kin.wide 12in.(0.3m) H,O
14 Wire,noninsulated 24AWGstranded 24in.(0.6m) H,O
15 Roundheadmachinescrew #6–3238mm 2 H,O
16 Roundheadmachinescrew #6–3235mm 2 H,O
17 Testtube(Corning-PyrexH9850-40) 40mm3130mm 1 S,O
18 CardstockwithHDpackingtape 2in.32in.square 1 H,O
onaside
19 Flexiblehose 1in. 4ft(1.2m) H,O
20 Pipecoupling11 S/S,Sch.40,1Jin. 1 H,O
21 VelcroHself-grabbingstrap Kin.38in. 1strip H,O
1CPVC,chlorinatedpolyvinylchloride;PVC,polyvinylchloride;HD,heavyduty.
2OD,outerdiam;ID,innerdiam;S,slip;Sch.,schedule;AWG,Americanwiregauge.
3H,hardwarestore;S,scientificsupply;O,online.
4Roundedendofbulbremovedand2opposingJ-in.(6.2-mm)holesdrilledhalfwayupsides.
5Insidestopflangeremovedfromonecoupling.
6Modifiedtoacceptthecouplingfrompartno.3.
7K-in.(12.5-mm)‘‘V’’cutfromthebottom.
8Jin.(6.2mm)ofthedownwardendremoved.
9Modifiedtoacceptthin-walledPVCpipe.
10Mouthofverticaltubeof‘‘T’’honedouttoaccommodatethicknessofVelcro-strapgasket.
11Usedasastandtoholdcompleteddevice.
MARCH2012 OPERATIONALNOTE 67
walledbrasstubing(Table 1:partno.1)isinserted of an insect being drawn back out of the glass
into a rubber pipette bulb (Table 1: part no. 2) bioassay tube into the vacuum source. The baffle
withtheroundedendremovedand2opposingJ- andthevacuumpressureareoptimizedbyleaving
in. (6.2 mm) holes drilled halfway up the sides of a J-in. (6.2 mm) gap at the top between the
the bulb. The drilled holes reduce eddies in the cardstockpaper and interior upper surfaceof the
pipette bulb and smooth the flow of air through ‘‘T.’’Thebushingofthepick-uptubeassemblyis
the pick-uptube assembly. The cut section ofthe then attached to the other lateral opening of the
pipette bulb is inserted up to the stop flange in a PVC‘‘T’’andalsosecuredwithascrew(Table 1:
straight coupling (Table 1: part no. 3). A stop partno.16).Theverticalopeningofthe‘‘T’’(i.e.,
flangeisaraisedportionwithinPVCfittingsthat thehandgrip)isslightlyboredouttoaccommodate
prevents joined parts from sliding through. A a length of the loop material side of a VelcroH
lengthofclearacrylictubing(Table 1:partno.4) strap adhered with double-sided sealing tape or
isinserteduptothestopflangefromtheopposite room-temperature vulcanization (RTV) adhesive
endofthesamestraightcoupling.Theotherendof (Table 1:partno.21;notshown).Thiscushioned
the clear acrylic tubing is attached to an L- gasket creates a vacuum seal and allows easy
coupling (Table 1: part no. 5), which is then insertion and removal of the glass bioassay tubes
attached to a 2nd length of clear acrylic tubing (Table 1: part no. 17; PyrexH glass culture tube,
(Table 1:partno.6).This2ndclearacrylictubeis 40 mm 3 130 mm, rimless, flat-bottom; part
theninsertedcompletelythroughanotherstraight no. 9850-40; Corning, Corning, NY). Screws
coupling(Table 1:partno.3)thathashadthestop (Table 1: part no. 15) are inserted into the
flangegroundoffsothat,1Lin.(43.7mm)ofthe handgrip at 1J in. (31.2 mm) from the opening,
tube is protruding through the coupling. The asstopsforthebioassaytubes(Fig. 2:partno.15).
coupling and acrylic tubing assembly is inserted The electrostatic grounding system is con-
into a reducer bushing (Table 1: part no. 7) that structed of grounding (noninsulated) wire, RTV
hasbeenboredouttoaccepttheoutsidediameter adhesive,andcoppertape.Thegroundingsystem
of the coupling. An L-coupling (Table 1: part isconstructedtoutilizetheoperatorasthebridge
no.9)withJ in. (6.2mm) of the downwardend betweentheWynnGunandtheground.Oneend
removed is attached to the protruding end of the of the grounding wire (Table 1: part no. 14) is
acrylic tubing. Another length of clear acrylic wrapped 3 times around the exposed part of the
tubing (Table 1: part no. 8) is attached to the brass tubing (Table 1: part no. 1) and anchored
trimmedendoftheL-coupling,andaK-in.(12.5- with solder. The grounding wire is then wrapped
mm) ‘‘V’’ is cut from the opposite end of the tightlyaroundthepick-uptubeandroutedtothe
tubing. The entire series of fittings in the pick-up handgrip as shown in Figs. 1 and 2. The wire
tube assembly should be cemented using PVC/ must be tightened flush against all surfaces it
chlorinated polyvinyl chloride (CPVC) adhesive contacts and to help prevent unraveling, dots
(e.g., All-Purpose PVC/CPVC cement; product of RTV adhesive (e.g., INSTAbond 1645 RTV
no.30818;Oatley,Cleveland,OH),exceptforthe silicone; National Stock No. 8040-00-145-0020;
proximaljointinthe45uL-coupling(Table 1:part ACCRAbond, Inc., Olive Branch, MS) can be
no.5), which should beallowed torotatetoseta applied intermittently. Adhesive copper tape
working angle comfortable for the operator, and (Table 1: part no. 13) is applied to the handgrip
thelastlengthofclearacrylictubing(Table 1:part to anchor the other end of the grounding wire
no. 8), which should be attached and cemented and provide a good contact to the operator and
after the tube assembly has been affixed to the completing the circuit to the ground.
mainbodyassembly. Finally, the vacuum source is fitted to the
Themainbodyassemblyisattachedtoboththe system. The thin-walled PVC tube projecting
vacuumsourceandthepick-uptube.Theelectro- fromtherear of thegun (Table 1: part no.10)is
staticgroundingsystemcannotbecompletedand attachedviaflexibleairhose(Table 1:partno.19)
thepick-uptubecannotbeattachedandcemented to the intake aperture of an electric blower fan
in place without the main body assembly. Thin- (e.g.,a60-ft3/minDaytonblower;partno.4C441;
walledPVCpipe(Table 1:partno.10)iscutand Grainger Industrial Supply, Lake Forest, IL).
cemented to a reducer bushing (Table 1: part For aspirating mosquitoes, the vacuum source
no. 11) that has been bored out to accept the shouldbeadjustedtoproduceanoptimalairflow
outsidediameterofthepipe.Thereducerbushing of 1.5 cm3/sec through the tip of the pick-up
isattachedtooneofthelateralopeningsofaPVC tube. Proper airflow should be checked with
‘‘T’’ (Table 1: part no. 12) and secured lightly an anemometer. A pipe coupling (Table 1: part
withpressure from a screw (Table 1: part no. 16) no. 20; not shown) can be fastened to the lab
insertedthroughtheupperwallofthe‘‘T’’.Apiece benchtoserveasastandfortheWynnGunwith
of cardstock paper (Table 1: part no. 18) is bioassay tube attached.
attached with heavy-duty packing tape to the The Wynn Gun is a novel modification of
inside rear surface of the ‘‘T’’ to act as a flow established aspirator designs that allows delicate
restrictorbaffle.Thisbafflereducesthepossibility testorganismstobeaddedrapidlyandefficiently
68 JOURNALOFTHEAMERICANMOSQUITOCONTROLASSOCIATION VOL.28,NO.1
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