CTCFAQGuide(16July2018)

TABLEOFCONTENTS

1 CANETOADCHALLENGE(CTC)1.1. WhatistheCaneToadChallenge(CTC)?1.2. HowistheCTCfunded?1.3. HowcanIjoinasaCTCMember?1.4. HowcanIjoinasaCTCAffiliate?

2 CANETOADTADPOLETRAPPING2.1 Howdoestadpoletrappingwork?2.2 Whatistheecologybehindtadpoletrapping?2.3 Whatisthechemistrybehindtadpoletrapping?2.4 HowdoestheCTCharvesttadpoleattractant?2.5 HowdoestheCTCexciseparotoidglandsfromdeadtoads?2.6 HowdoestheCTCshipexcisedparotoidglands?2.7 HowdoestheCTCformulatetadpoleattractant?2.8 WhydoestheCTCaddgreenfooddyetoBufoTabs?2.9 HowcanIgetsomeBufoTabs?2.10 AreBufoTabssafefortheuser?2.11 AreBufoTabssafefortheenvironment?2.12 Isthereariskofby-catch?2.13 AretherelimitationsontheuseofBufoTabs?2.14 DoBufoTabshavealongshelflife?2.15 HowlongdoBufoTabsworkinatadpoletrap?2.16 HowdoIbuildatadpoletrap?2.17 WhenshouldIuseatadpoletrap?2.18 WhereshouldIuseatadpoletrap?2.19 I’vecaughttadpoles,sowhatshouldIdowiththem?2.20 WhatdoIdowithausedBufoTab?2.21 Dofrogtadpolesgetattractedtotraps?2.22 Willtadpoletrappingdeliveranenvironmentalbenefit?2.21 WilltadpoletrappingeradicatecanetoadsinAustralia?

3 ADULTCANETOADS3.1 Whatisthemosthumanewaytokilladulttoads?3.2 Aretheremethodsforkillingadulttoadsthatarenotrecommended?3.3 DoestheCTCneeddeadtoads?3.4 WheredoestheCTCsourceitsdeadtoads?

4IMAGESOFSUCCESSFULTRAPSINACTION

CTCFAQGuide(16July2018)

1.CANETOADCHALLENGE(CTC)

1.1. What is the CaneToadChallenge (CTC)?The CTC is a University of Queensland (Institute for MolecularBioscience) community engagement and citizen science initiative, designed to fast track public access toinnovative cane toad tadpole trapping technology co-invented by researchers from The University ofQueenslandandTheUniversityofSydney.

1.2. HowistheCTCfunded?TheCTCisactivelyseekingphilanthropicandcorporatesponsorship,researchgrantfunding,andpublicdonations.

1.3. HowcanIjoinasaCTCMember?Individualsover18yearsoldmayregisteronlineasCTCMembers,freeofcharge.CTCMemberswillreceiveperiodicemailCTCUpdates.

1.4. HowcanIjoinasaCTCAffiliate?CTCAffiliatemembership isby invitation,and isopentoawiderangeoforganizations.Theseincludevolunteerandnot-for-profitgroups,societies,philanthropicorganizationsandcommercial businesses, as well local, regional, state and federal government agencies. Interestedorganisation should contact theCTC.CTCAffiliates co-signamemorandumofunderstanding (MoU)withUQ,validfor3years,withthefollowingbenefits;

• RecognitionasasupporteroftheCTCcommunityengagementandcitizenscienceinitiative.• RecognitionforadvancingtheprotectionoftheAustralianenvironmentfromthecanetoad.• Fast-trackpathwayforcontributingto,andparticipatinginCTCactivities. • PriorityaccesstoCTCtechnicaladviceandBufoTabs(subjecttoavailabilityandapprovals).• Optiontoleveragemembershipinsupportofenvironmentrelatedgrantapplications.• Optiontoleveragemembershipinonline,on-airandinprintmediaenvironmentprofiles.

2.CANETOADTADPOLETRAPPING

2.1 How does tadpole trapping work? This technology uses natural cane toad attractant chemistry inconjunctionwithtraps,toachievethecaptureandremovalofcanetoadtadpolesfrommanagedwaterways(e.g.dams, ponds, streams, creeks). Tadpole trapping is an environmentally sustainable and intuitive technology,readily transferable to thepublic.Coordinated implementationof tadpole trappinghas thepotential toreducecanetoadtadpole(andadult?)populations,andalleviatetheenvironmentalimpactofthistoxicinvasivepest.

2.2 Whatistheecologybehindtadpoletrapping?Fieldobservationsrevealedthattadpolesactivelysearchoutandconsumetheeggsofotherfemalecanetoadswithinthesamebodyofwater.Possibleexplanationsforthisbehaviour include(i)providingtadpoleswithavaluablesourceofnutrition, (ii)allowingtadpoles torechargetheirtoxinreserves,tohelpdefendagainstpotentialpredators,and(iii)toeliminategeneticcompetition.Asthisbehaviour is successfuleven in lowtozerovisibility, itwasspeculated thatcane toadeggsreleaseachemicalattractantthatrevealstheirlocationtocanetoadtadpoles.Thisattractantisatthecoreoftadpoletrapping.

2.3 Whatisthechemistrybehindtadpoletrapping?Chemicalanalysisidentifiedtheattractantreleasedbyeggsasverycloselyrelatedtoadultcanetoadtoxin.Femaletoadsdepositamodifiedversionofadulttoxintoprotecteggs from potential predators, unaware that this chemistry exposes the eggs to cannibalism by tadpoles.Presumably the threat posed by non-cane toad related predators exceeds that posed by cane toad tadpoles,ensuringthatco-depositingofeggtoxinwitheggsdeliversanetsurvivaladvantage.Asfarasweknow,nootherAustralianaquaticspeciesproduce,orareattractedtocanetoadtoxin,ensuringspecies-specificity.

2.4 HowdoestheCTCharvesttadpoleattractant?As it is impractical tosourceattractantchemistry fromeggmasses, we harvest related chemistry from dead adult cane toads.Weworkwith approved CTC Affiliates toredirect and recycle dead toads acquired during toad busting activities, from landfill to our laboratory.Shipmentsofdeadadultcanetoads(orexcisedparotoidglands)mustbepre-approvedandaccompaniedbya"CaneToadShipmentForm".Torecoverattractantinvolvesexcisingtheparotoidglands,followedbybatch-wiseblending and enzymatic transformation, extraction, fractionation, purification and chemical analysis.Importantly, this process removesnon-attractant chemicals, aswell as all tracesof animal tissue, todeliver aproductthatcanbeformulatedastadpoleattractantbaits.

CTCFAQGuide(16July2018)

2.5 HowdoestheCTCexciseparotoidglandsfromdeadtoads?Dead frozenadult toadsandare thawedandasharpknifeisusedtocutofftheparotoidglands,positionedoneachfrontshoulder.Theattachedpicture(Figure1)showsthelocationofparotoidglandsonthetoad,plusdorsalandventralviewsofanexcisedgland,andanimage of a compressed gland illustrating toxin secretion. The ventral view reveals yellow vesicles inside thegland thathold the toxin.Eachvesiclehas anattached duct leading to the surface of thetoadskinandwhencompressedeitherduringa predatory attack orwhen handled roughly,toxinisforcedoutoftheducts.Whenthawingfrozen toads the glands can leak toxin(freezing puts pressure on vesicles), andalthoughthemajorityoftoxinremainswithinthe gland/vesicles, to avoid losing toxin it isimportant to not wash the excised glands.Thawed toads are slippery, and the toxin isquite poisonous, so it is important to usewaterproof gloves under solid work gloves,and a sharp knife. When excising glands weare careful to cut deep enough to get all thevesicles intact, and to avoid skin contact. Toprevent eye splashes we always wear safetyglasses.Excisedglandsarebaggedandfrozenindefinitely, and can be defrosted in batcheswhenneededtobeprocessedintobaits.

2.6 HowdoestheCTCshipexcisedparotoidglands?Whereitisnecessarytotransportexcisedparotoidglandsthiscaneitherbedonebyhanddelivery,orbypost,ideallywiththeglandsfrozen.Theeffortandcostofmailingafrozenshipmentismoreor lessthesameregardlessofthenumberofglands,asmostoftheweight is inthepackaging.Thestrategyweuseisasfollows;

• Excisedglandsarefrozenasasingleblock.• Frozenglandsaresealedinatleasttwolayersofwatertightziplockbags(removingallair).• Bagged,frozenglandsarepackedinasmall“hard”eskywithseveralfrozenfreezerpacks.• Theeskylidissecurelytappedshuttoavoidanyleakage.• Theeskyispackagedinanaddressedcardboardshippingbox,withabsorbentpadding.• Thecardboardboxcontainingtheeskyisshippedbyovernight“cold”courier.

2.7 HowdoestheCTCformulatetadpoleattractant?Todeliverattractant ina formthatcanbeused tocatchcane toad tadpoles requires a cheap, safe, environmentally sustainable, slow release formulation. This iscurrently achieved by absorbing a calibrated quantity of attractant on commercially available aquarium air-stones,alongwithgreenfooddye.Thedried,green,attractantloadedair-stonesaredistributedasBufoTabs.

2.8 Why does the CTC add green food dye to BufoTabs? When added to a trap the green food dye rapidlywashesoff,makingiteasiertorecognizeused(spent)fromunused(unspent)BufoTabs.

2.9 HowcanIgetsomeBufoTabs? The patent for BufoTabs has been licensed, and efforts are underway toregister,manufacture anddistribute these as a commercial product. As thismay take 2-4 years, the CTCwasestablished as an interimmeasure. The CTC is currently not in a position to release BufoTabs direct to CTCMembers,however,weareworkingcloselywithCTCAffiliatestoachieveamanageddistribution.IfyoubelongtoorknowofanorganisationwhomaybeinterestedinbecomingaCTCAffiliate,pleaseencouragethemtovisitthewebsiteandsubmitanexpressionofinterest.

2.10 Are BufoTabs safe for the user? Like any pest control product the answer is a qualified yes. As the

Figure 1. Adult toad parotoid gland

CTCFAQGuide(16July2018)

attractantchemistryiscomprisedofanaturaldefensivetoxin,BufoTabsshouldonlybeusedbyadultsover18years, and users should avoid skin contact or ingestion. All end-users should read and heed advice indocumentationsuppliedwithBufoTabs,which includesaUQMaterialShipping form,andMaterialSafetyDataSheet(MSDS).

2.11 AreBufoTabssafefortheenvironment?Yes.BufoTabsseektomimicthechemicalplumereleasedbycanetoadeggs.Ataquantitative level,eachadult toadproducessufficientattractant toprepare5-20BufoTabs.Putanotherway,asingledeadtoadwouldrelease5-20timesmoretoxinintotheenvironmentthanasingleBufoTab.It’salsoworthnotingasingleBufoTabcancatchasmanyas1,000to10,000toxictadpoles,withanetreductionofenvironmentaltoxinload.

2.12 Isthereariskofby-catch?Allexperiencetodateisthatby-catch(iefish)isnotanissue.Thisisinpartduetothespeciesselectivenatureofthepheromoneattractant,furtheraidedbysmallfunnelentry(1cm).Itisalsoimportant tonotethat tadpole trapsdonotkill tadpoles,so in theunlikelyevent thatby-catch isencounteredthereisampleopportunitytomakenoteofandreleasethespeciesinvolved.

2.13 AretherelimitationsontheuseofBufoTabs?BufoTabsmustbeusedassupplied, for thepurposeofcanetoadtadpoletrapping,andnotbemodified,repurposed,sold,ordistributedoutsideCTCAffiliateorganisations.When using BufoTabs in a particular location please ensure you have authorisation, and follow all directivesfromlandownersand/orauthorities.

2.14 DoBufoTabshavealongshelflife?BufoTabscanbestoredindefinitely,ifkeptdryandinarelativelycool(<30°C)location,awayfromdirectsunlight.ItisstronglyrecommendedthatBufoTabsbestoredintheoriginalpackaging.

2.15 HowlongdoBufoTabsworkinatadpoletrap?WhenaBufoTabisaddedtoatrap,theattractantisslowlyreleased to generate an attractant plume that extends through the funnels, outside and away from the trap,attractingcanetoadtadpolesbacktothesource(insidethetrap).Ifthetrap,BufoTabandattractantplumeareleftundisturbed(i.e.thetrapisnotremovedfromthewater),aBufoTabshouldgenerateasustainedattractantplume for at least 24 h, depending on environmental conditions. If during this period (i.e. 2 h) the trap issuccessfulinattractingtadpolesandisremovedfromthewater,anddrainedtofilteroffthetadpolecatch,thisdisturbs and negates any established plume, severely reducing the effective duration of the “partially” usedBufoTab.Toaccommodatetheseissueswerecommendthefollowingsequenceofevents;

• PositionthetrapandaddanewBufoTab.• Monitorthetrapoverthenext2-24h.• Removethetrapfromthewateratanytime(2-24h)wherethereisagoodcatchoftadpoles.• Drain,harvestanddisposeofthetadpolesandtheusedBufoTab.• Ifthetadpolenumbersinthewaterbodywarrant,repeat(i)to(iv)inthesamelocation.

Alternatively,ifyourtrapdesignallowsforit,youcanavoiddisturbingtheattractantplumebyleavingthetrap(andassociatedattractantplume)inplaceandcarefullynettingthetadpolesdirectlyfromthetrapwithasmalldabnet.Thiswaythetrapcanbelefttorunforthefull24hwithperiodictadpoleharvestsbynetting.Ifyoudousethisapproachbeverycarefultoavoidaccidentallykilling(crushing)anytadpolesinthetrap.Deadtadpolesreleaseanalarmchemicalthatcouldnegatetheattractant.

2.16 How do I build a tadpole trap? Tadpole traps (funnel traps) are typicallyconstructed by attaching plastic funnels to opposite ends of a plastic box, with thesmall diameter end projecting into the box (Figure . It is important that the joinbetween funnel andboxbewater tight, and that theboxhasnootherholes or gapsthat will allow leakage of attractant. When a bait is added to a trap the scent trailextends out through the funnels. Tadpoles sense the trail and move up theconcentrationgradienttowardsthetrap.Whenencounteringthetrapthetadpolesare"funnelled"intothetrap.Onceinthetraptheexitissmall,andintheabsenceofareasontoexit(ienoreversescenttrail),tadpolesstayinthetrap.

Figure 2. Tadpole trap

CTCFAQGuide(16July2018)

2.17 WhenshouldIuseatadpoletrap?Thebesttimetousetadpoletrapsisduringperiodsofsignificantcanetoad breeding, which are typically weather dependent. Traps work best when deployed in areas with hightadpoledensity.

2.18 WhereandhowshouldIuseatadpoletrap?Therangeof thescent trail releasedbybaited traps is finite,butdoesextendfurtherfromthetrapwithtime.Itisimportanttoavoidcompetingscenttrails,thatcouldoccurifmultipletrapsareplacedincloseproximity.Itisalsohelpfultoscantheareaimmediatelyaroundtraps,andremoveanydeadadulttoadsfromthewater.Deadadultscanreleaseacompetingscenttrail.Itisalsoimportanttoavoidplacingtrapsinflowingwater(ierunningstreams)orinwaterbodiesexperiencinganinflowofwater(ierainrunoff),asdisturbedwaterwilldisturbthescenttrail.Trapsarebestplacedinstill,shallownearshorewater, sitting on the bottom, with the top 20% above water, and funnels fully submerged. Traps should becoveredwithalidandshadecloth,toavoidoverheating.Itisimportantthattadpolesbecapturedandremovedfromthetrapalive,asdeadtadpolesreleaseanalarmchemicalthatwillover-ridetheattractantsignal.

2.19 I’vecaughttadpoles,sowhatshouldIdowiththem?Canetoadtadpolesshouldbeharvestedlivefromtrapsbyeitherfilteringthroughaclothorsieve,orbyasmallhandheldnet.Onceharvested,livecanetoadtadpolesshouldbetransferredwithaminimumvolumeofwatertoaplasticbagoranothersuitablesealedcontainer.Inanadaptationofthecool-freezemethodapprovedforeuthanizingadulttoads,thesealedbag/containerofcanetoadtadpolesshouldbecooled(e.g.onice)untiltadpolesarenon-motile,afterwhichitcanbetransferredtoafreezerandfrozensolid.Beawarethatdeadcanetoadtadpolesaretoxicandifleftintheenvironmentmaybescavenged and poison native predators. Be sure to bag and bin dead cane toad tadpoles along with usedBufoTabs.

2.20 WhatdoIdowithausedBufoTab?Bagandbinalongwiththedeadtadpoles.

2.21 Dofrogtadpolesgetattractedtotraps?Allevidence todatesuggests thatBufoTabsonlyattractandtrapcane toadtadpoles.Thisdoesnotmeanthatotherspeciescannotrandomlywander into the trap,but there iscurrentlynoevidenceofsignificantaccidentaltrappingoffrogtadpoles.Nevertheless,itisalwaysgoodpracticetomonitor traps forunwantedby-catch, and in theunlikelyevent thatby-catch is encountered takenoteandrelease(alive).

2.22 Willtadpoletrappingdeliveranenvironmentalbenefit?Thisisagoodquestion,andthejuryisstillout.Theanswerwilllikelyonlybecomeevidentafterwerolloutthetechnologyforasustainedperiodoftime,andassessimpact.Thatsaid,weareconfidenttadpoletrappingcanremoveverylargenumbersoftoxictadpolesfromtheenvironment,andthiswillundoubtedlyreducetoxin load. Insomeecosystemstheremovalofcompetingcanetoadtadpoleswillalsoreleasehabitatandresources,tothebenefitofotheraquaticspecies.Wearehopefulthatthelarge-scaleremovaloftadpoles,coordinatedwithhandcaptureandcullingofadulttoads,willsignificantlyreducelocalcanetoadpopulations.Thiswillmostlikelybeevidentinandaroundmanagedwaterbodiessuchasfarmdamsandpublicwaterfeatures(i.e.streams,lakes,ponds,golfcourses,wetlandsetc…).

2.23 WilltadpoletrappingeradicatecanetoadsinAustralia?Canetoadtadpoletrappingisavaluabletool,butitalone will not eradicate cane toads from Australia. It can be used to remove tadpoles, and hopefully reducejuvenile and adult cane toad populations, in and around accessible water features – delivering localizedenvironmentalreturns.As localas thesereturnsmaybe, theyarestillwins in thewaragainst the toad.By itsvery nature, tadpole trapping is labour intensive (albeit less so than adult capture) and cannot becomprehensivelyrolledoutoveracontinentasvastasAustralia.Ithasaroletoplay,butweneedtocontinuetheresearch,tofindadditionalapproachestoachieveanationwidesolutiontotheCaneToadChallenge.

CTCFAQGuide(16July2018)

3.ADULTCANETOADS

3.1 Whatisthemosthumanewaytokilladulttoads? The safest andmost humaneway for thepublic to killadult toads is the cool/freezemethod. Thismethod involves bagging toads and cooling them in a fridge untilcomatose (i.e. notmoving), followedby storing in a freezeruntil frozen solid. For further informationon thismethoddownloadandread“Iscoolingthenfreezing”ahumanewaytokillamphibiansandreptiles?”

3.2 Aretheremethodsforkillingadulttoadsthatarenotrecommended?Anymethod that is inhumane innotacceptable.This includesgolf clubs,malletsandotherbashing implements.Wealsocautionagainst theuseofchemicals (i.e.Dettol,Hopstop, salt, bleach…)as theseare indiscriminateandwill kill speciesother thancanetoads.Equallyproblematic,ifdispersedintheenvironmentsomechemicalscanseriouslydegradetheviabilityofaquaticecosystems.

3.3 DoestheCTCneeddeadtoads? Yes. TheCTC extracts cane toad tadpole attractant fromdead adult canetoads.Todothisweneedasupplyofdeadtoads.

3.4 WheredoestheCTCsourceitsdeadtoads?Wecanonly receive toads thathavebeencollectedandkilledhumanelybyapprovedCTCAffiliates,aspartofongoingcanetoadcontrolactivities-allowingustodivertdeadtoads from landfill, to produce attractant. CTCAffiliates thatwould like to be approved to supply dead toadsshould contact the CTC to confirm that their protocols are humane, and to receive advice on processing andshipping.TheCTCcannotacceptdeadtoadsfrommembersofthepublic,orfromunapprovedCTCAffiliates.

CTCFAQGuide(16July2018)

4IMAGESOFSUCCESSFULTRAPSINACTIONQueenslandFrogSocietyTrap(imagebyJuliaSquires)

HeadlandGolfClub(imagebyJohnSquires)