A new technique in the excavation of ground-nest bee burrows (Hymenoptera: Apoidea)

RevistaBrasileiradeEntomologia62(2018)1–4

REVISTA

BRASILEIRA

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Entomologia

Short

Communication

A

new

technique

in

the

excavation

of

ground-nest

bee

burrows

(Hymenoptera:

Apoidea)

Diego

Marinho

_,

_Juliana

_Andrade

_,

_Rodrigo

_O.

_Araujo

_,

_Felipe

_Vivallo

a_{UniversidadeFederaldoRiodeJaneiro,MuseuNacional,DepartamentodeEntomologia,QuintadaBoaVista,RiodeJaneiro,RJ,Brazil} b_{UniversidadCatólicadelMaule,FaculdaddeCienciasBásicas,DepartamentodeCiênciasBiológicasyQuímicas,Talca,Chile}

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Articlehistory:

Received17October2017 Accepted13November2017 Availableonline21November2017 AssociateEditor:RodrigoGonc¸alves Keywords: Apidae Apoidea Broodcells Methodology Solitarybees

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Beeshaveadiversifiednaturalhistory,thusthemethodsappliedtostudysuchdiversityarevaried.When itcomestostudiesofnestingbiology,beeswhichnestinpre-existingcavitieshavebeenreasonablywell studiedsinceresearchersstartedusingtrap-nests.However,beeswhosenestsarebuiltundergroundare poorlystudiedduetothedifficultyoffindingnaturalnestingareasandtheabsenceofamethodthat facilitatesbeenestexcavation.Thelatterisevidencedbythelackofaccuratedescriptionsinliteratureof hownestsareexcavated.Inthisstudywetestedcylindricalrubberrefillsoferaserpenasanewmaterialto beusedasatracerofundergroundnestgalleriesinanaturalnestingareaoftwospeciesofEpicharisKlug, 1807(Apidae).Wecomparedthistechniquedirectlywithplasterinpowderformmixedwithwaterand ourresultswithothermethodologicalstudiesdescribingalternativemethodsandmaterials.Therubber refilltechniqueovercamethemainissuespresentedbymaterialssuchasplaster,moltenmetalalloysand bioplastic,namely:deathoftheorganismsbyhightemperaturesand/orformationofplugsandmaterials undulyfollowingtherootsinsidethegalleries.

©2017SociedadeBrasileiradeEntomologia.PublishedbyElsevierEditoraLtda.Thisisanopen accessarticleundertheCCBY-NC-NDlicense(http://creativecommons.org/licenses/by-nc-nd/4.0/).

Beesareinsectswitha verydiversifiednaturalhistory.Most speciesaresolitary(about85%)andeachfemaletakescareofits ownbroodcells(Batra,1984;Michener,2007).Theirnestsarebuilt withseveralmaterials,likeresin,leaves,Dufour’sglandsecretions, etc.andonseveralsubstrates,suchasrocks,wood,plantbranches, insidepre-existingcavitiesordirectlyintheground(Batra,1984). Consideringthisbionomicaldiversity,themethodsusedtostudy theseorganismsarealsodiversified(Linsleyetal.,1952;Michener etal.,1955).

ApidsandMegachilidswhichnestinpre-existingcavitiesinthe wood,forinstance,havetheirbiologybetterunderstooddueto theadventofthetrap-nestmethod(Krombein,1967).However, themajorityofbeesbuildtheirbroodcellsunderground (Batra, 1984),andsubsequently,littleisknownabouttheprocessofnest constructionandassociatedaspectsofthebiologyofthesespecies, mostlyduetothedifficultyoffindingnaturalnestingareasandto obtaindatacollectionfromtheirundergroundnests.

Several methodological studies were developed regarding Hymenoptera ground-nesting habits (Michener et al., 1955; WilliamsandLofgren,1988),butthemajorityofthemwerefocused

∗ Correspondingauthor.

E-mail:diego.mp89@gmail.com(D.Marinho).

onants(Tschinkel,2010,2013).Forstudiesthatproposespecific methodsforcollectingundergroundbeenests,thematerialsused areplaster(Linsleyetal.,1952;Micheneretal.,1955;Nordenetal., 1994),moltenmetalalloys,paraffinwax(Linsleyetal.,1952)and bioplastic(Howell,1960).

From1960onwards,tothebestofourknowledge,no method-ological studies have been published focusing on methods of collectingundergroundbeenests,evidencingthelackofliterature concerningalternativewaystostudysuchnests.Furthermore, sev-eralstudiesonthebiologyofground-nestingbeesdonotdescribe in detail how excavation is performed (see Gaglianone, 2005; Gaglianone etal., 2015;Rocha-Filho etal., 2008;Roubik,1980; Rozen,2016;ThieleandInouye,2007),hinderingthepossibilityof comparisonofthemethodsused.Inthiscontext,weaimtopropose anewpracticalmethodfortracingthenesttunnelsandcollecting broodcellsfromundergroundnests.Additionally,wecompareit withothermethodologicalstudiespresentinliterature.

Nestsexcavation

ThestudywascarriedoutinanestaggregationareaofEpicharis analisLepeletier,1841andEpicharisfasciataLepeletier&Serville, 1828atthe“JardimBotânicodoRiodeJaneiro”(225814S,43 1318W),StateofRiodeJaneiro,Brazil.Nestexcavationswere https://doi.org/10.1016/j.rbe.2017.11.003

0085-5626/©2017SociedadeBrasileiradeEntomologia.PublishedbyElsevierEditoraLtda.ThisisanopenaccessarticleundertheCCBY-NC-NDlicense(http:// creativecommons.org/licenses/by-nc-nd/4.0/).

2 D.Marinhoetal./RevistaBrasileiradeEntomologia62(2018)1–4 performedduringfourhoursontwodifferentdates:onFebruary

6th,2017whenE.analiswasatitspeakactivityandonApril6th, whenE.fasciatawasnesting.Severalfemalesofbothspeciesshared thesingleaggregationarea,butnestingactivitieswere indepen-dent.

Inthefirstexpedition,weusedpowderofplasterandplaster mixedwithwater,inordertotrackthegalleryofthenestsuntilthe broodcellchambersbuiltbythebeesaccordingwiththe methodol-ogyoriginallyproposedbyMicheneretal.(1955)andsubsequently widelyusedbyothersresearchers(Danforth,1989;Eickwortand Eickwort,1969;Osgood,1989;Wcisloetal.,1993).

Duringthesecondvisit tothenestingareawetestedrubber refillsfromtheGomPenMaped® brand(Fig.1A),usedineraser pens.Eachrubberrefillhasacylindricalshape,0.7cmdiameter and11.0cmheight,highflexibility,anaveragecostofUS$2.50and canbeeasilyfoundatstationers.Acrossitsheight,wemadegraded marksincentimetersusingaballpointpen.Fordeeperbroodcells whereasinglerubberrefillcannotreach,wegluedseveralofthem toeachotherbytheirextremitiesusingcyanoacrylateoftheSuper Bonder®brand(Fig.1B).Wetestedthetracerwithuptofourrubber refillsduetothedepthofthebroodcellsintheground.

ExcavationsofE.analiscellswereperformedduringthefirst visit,whenwetestedplaster,butwithlittleprogress.Thetrack ofthenestgallerywaseasilylostusingthatmaterialinpowder

form; the debris and soil around the tunnel kept falling dur-ing the excavation using the shovel. When the plaster was applied mixed with water, the cast hardened in about one hour,butacquiredafragileconsistencyandbrokeeasilyduring excavation. We obtained six brood cells of E. analis using this method.

Theexcavationusingrubberrefill(henceforth “rubber”)was employed during the second visit to the same area, when E. fasciata wasnesting. The procedurewas performedas follows: theentrance of thenest tunnel wasspotted visually; the rub-ber was inserted and conducted inside the gallery. Using the rubber as a tracer, we dug a parallel transect using a shovel (Fig.1B);thisway,thenestarchitecturewaspreservedinorder tobedrawnandmeasured, since therubberretainsthetunnel structure, serving as a tracer for the deeper parts of the nest gallery.

Aftertheschemeofthenestarchitecturewasdone,anareaof approximately35cmofdiameterwasdugaroundthegallerytraced bytherubber(Fig.1C).Thisposteriorexcavationwasperformed inordertofindthebroodcellsplacedrandomlyaroundthemain gallerybythefounderbees.Then,broodcellswereremoved manu-allyandwewereabletocollect14ofthem,fourofE.analisandten ofE.fasciata.Noharmwasdonetothebeecellswhenexcavating throughthistechnique.Vouchersofthebroodcellsandemerged

Fig.1.Rubberrefillsusedastracerforexcavation:(A)withgradedmarks,and(B)gluedtoeachotherbytheirextremitiestracinganEpicharisfasciatanestgallery.(C) PosteriorgalleryexcavationinofEpicharisfasciatanestsinordertoobtainrandomlyplacedbroodcellsaroundtherubberrefilltracer.

D.Marinhoetal./RevistaBrasileiradeEntomologia62(2018)1–4 3 specimensweredepositedintheEntomologicalCollectionofthe

MuseuNacionaldoRiodeJaneiro(MNRJ).

Tracermethodsfornestexcavation

The method chosen for excavating nests may vary greatly dependingonthescopeandtheavailableinfrastructuretoperform thestudy.Themostusedmaterialisplaster,mainlybecauseis eas-ilyobtainedandhandled(Linsleyetal.,1952).However,wenoticed thesameproblemsreportedbyHowell(1960):wecontinuallylost thetrackofthetunnelsduetoloosedebristhatkeptfallingfrom thegallery;thecastformedwhenweapplieditwithwaterwas easilybroken;andmuchtimewasspentwaitingforittoharden. Theprocedureemployedusingrubbersharethesameadvantages andovercomesallthedisadvantagesofplaster:thereisnoneedfor water,nowaitingtimeandthetunnelisneitherlostnortherubber isbrokenduringtheexcavation.

Othermethodsdescribedinliteraturearemetalalloys,dental plaster,paraffinwaxandbioplasticwhichcastthestructureofthe undergroundnests;however,theeggsorimmaturelarvaemaybe killedduetothehightemperatureofthesematerialswhenthey areemployed(Micheneretal.,1955;Tschinkel,2010),withthe exceptionofbioplastic(Howell,1960).Inthisstudy,thecell archi-tectureofboth speciesobservedwouldfavorthosematerialsto floodintobroodcells,sinceweobservedsomeapertures, proba-blyforlarval-breathing.Instudiesaimingtostudythebiologyof beesorevenotherinsectsthatnestintheground,thissideeffect isnotdesirable,sincethecollectedbroodcellsaretakentothe lab-oratoryforfurtherobservationssuchasmonitoringandstudying immaturedevelopment.Castsofthetunnelstructurescannotbe madeusingtherubbertechnique,butweconsideritalesser dis-advantage.Additionally,theuseofmetalalloysisdifficultbecause itrequiresmuchsafetyequipmentwhenhandlingitinthefield (Tschinkel,2010).

In spite of bioplastic being a better alternative for the cast methodsdiscussedabove,Howell(1960)observedsomeissues: difficultytoexcavatethecastformed,bio-plasticreadilyfollowing rootsinthepathofthegallery,andtheformationofplugsbecause ofitsunevensolidification.Ourmethodalsoovercamethese dis-advantages,sincetherubberwasnotimpededtogothroughthe gallerybyroots,andaslongasitisalreadysolid,noplug-formation issuesoccur.

Thenewmethodpresentedcanbeusedtostudyunderground nestingbeeswithseveralsizes,aswecanfindrubberwith diame-tersvaryingfrom0.2to0.7cm.Wealsopointoutthatthistechnique canbeusedtootherground-nestingHymenopterabesidesbees. Thisisalsoalowcostmethod,duetounitvaluesrangingfromUS$ 2.2toUS$3.8,dependingonthedesireddiameter.Incomparison withothermethodssuchasmoltenmetalalloys,thecostis rea-sonablysmall,sinceonewouldnotneedtoolssuchasS-hooks,stir barsandhotgloves,whichmayturnfieldworkcostly(Tschinkel, 2010).

Studiesonotherground-nestingHymenoptera,suchasants, provideseveralcomparabletechniquesregardingspecific objec-tivesonantbiologystudiesandsuchcomparisonsarestatedas beneficialforthefield(Tschinkel,2010).Suchcomparable meth-odsarelackinginundergroundbeenestinvestigationsandevenin otherfieldresearchregardingpre-existingcavitiesbeenesters,as pointedoutbyMacIvor(2017)inhisreviewoncavity-nestboxesfor solitarybees.Overtime,severaltoolsmaybeusableforthestudy ofground-nestingbeesandwepointouttheneedfor methodol-ogydescriptionsemployedinnestingbiologystudies.Thedetailed descriptiononhowbroodcellsareexcavatedfromunderground

nestsmaycontributefortheestablishmentofbettermethodsto studyground-nestingbees.

Contributors

Allauthorsconceivedtheresearchandwenttofieldtoperform theexperiments.Thefirstauthorwrotethepaperandallauthors contributedequallyreviewingcriticallythemanuscriptfor impor-tantintellectualcontent.Allauthorsapprovedthefinalversionof thismanuscript.

Conflictsofinterest

Nonedeclared. Acknowledgments

We thank the Jardim Botânico do Rio de Janeiro, especially to Maria Lucia Franc¸a Teixeira, for the arrangements to allow ouraccesstothenestingarea;theConselhoNacionalde Desen-volvimento Científico eTecnológico (CNPq) for the scholarship grantedforthefirstandsecondauthorsofthisstudy,andClarisse FigueiredoforEnglishproofreading.ThispaperispartoftheSIGMA projectN◦21565MN/UFRJandthecontributionnumber18fromthe HYMN.

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