Efficacy of Visual Surveys for White-Nose Syndrome at Bat Hibernacula


previous item | next item

Citation
Efficacy of Visual Surveys for White-Nose Syndrome at Bat Hibernacula

Material Information

Title:
Efficacy of Visual Surveys for White-Nose Syndrome at Bat Hibernacula
Series Title:
Plos One
Creator:
Janicki, Amanda F.
Frick, Winifred F.
Kilpatrick, A. Marm.
Parise, Katy L.
Foster, Jeffrey T.
McCracken, Gary F.
Publication Date:
Language:
English

Subjects

Subjects / Keywords:
White-Nose Syndrome ( local )
Wns ( local )
Pseudogymnoascus Destructans ( local )
Bats ( local )
Bat Mortality ( local )
Bats In North America ( local )
Genre:
serial ( sobekcm )

Notes

Abstract:
White-Nose Syndrome (WNS) is an epizootic disease in hibernating bats caused by the fungus Pseudogymnoascus destructans. Surveillance for P. destructans at bat hibernacula consists primarily of visual surveys of bats, collection of potentially infected bats, and submission of these bats for laboratory testing. Cryptic infections (bats that are infected but display no visual signs of fungus) could lead to the mischaracterization of the infection status of a site and the inadvertent spread of P. destructans. We determined the efficacy of visual detection of P. destructans by examining visual signs and molecular detection of P. destructans on 928 bats of six species at 27 sites during surveys conducted from January through March in 2012–2014 in the southeastern USA on the leading edge of the disease invasion. Cryptic infections were widespread with 77% of bats that tested positive by qPCR showing no visible signs of infection. The probability of exhibiting visual signs of infection increased with sampling date and pathogen load, the latter of which was substantially higher in three species (Myotis lucifugus, M. septentrionalis, and Perimyotis subflavus). In addition, M. lucifugus was more likely to show visual signs of infection than other species given the same pathogen load. Nearly all infections were cryptic in three species (Eptesicus fuscus, M. grisescens, and M. sodalis), which had much lower fungal loads. The presence of M. lucifugus or M. septentrionalis at a site increased the probability that P. destructans was visually detected on bats. Our results suggest that cryptic infections of P. destructans are common in all bat species, and visible infections rarely occur in some species. However, due to very high infection prevalence and loads in some species, we estimate that visual surveys examining at least 17 individuals of M. lucifugus and M. septentrionalis, or 29 individuals of P. subflavus are still effective to determine whether a site has bats infected with P. destructans. In addition, because the proba

Record Information

Source Institution:
University of South Florida Library
Holding Location:
University of South Florida
Rights Management:
This item is licensed with the Creative Commons Attribution License. This license lets others distribute, remix, tweak, and build upon this work, even commercially, as long as they credit the author for the original creation.
Resource Identifier:
K26-00037 ( USFLDC: LOCAL DOI )
k26.37 ( USFLDC: LOCAL Handle )

USFLDC Membership

Aggregations:
University of South Florida
Karst Information Portal

Postcard Information

Format:
serial

Downloads

This item is only available as the following downloads:


Full Text

PAGE 1

RESEARCHARTICLEEfficacyofVisualSurveysforWhite-Nose SyndromeatBatHibernaculaAmandaF.Janicki1* ,WinifredF.Frick2,A.MarmKilpatrick2,KatyL.Parise3,Jeffrey T.Foster3 ¤,GaryF.McCracken11 DepartmentofEcologyandEvolutionaryBiology,UniversityofTennessee,Knoxville,Tennessee,United StatesofAmerica, 2 DepartmentofEcologyandEvolutionaryBiology,UniversityofCaliforniaSantaCruz, SantaCruz,California,UnitedStatesofAmerica, 3 CenterforMicrobialGeneticsandGenomics,Northern ArizonaUniversity,Flagstaff,Arizona,UnitedStatesofAmerica ¤ Currentaddress:DepartmentofMolecular,Cellular,andBiomedicalSciences,UniversityofNew Hampshire,Durham,NewHampshire,UnitedStatesofAmerica * ajanicki@vols.utk.eduAbstractWhite-NoseSyndrome(WNS)isanepizooticdiseaseinhibernatingbatscausedbythefungus Pseudogymnoascusdestructans .Surveillancefor P . destructans atbathibernacula consistsprimarilyofvisualsurveysofbats,collectionofpotentiallyinfectedbats,andsubmissionofthesebatsforlaboratorytesting.Crypticinfections(batsthatareinfectedbutdisplaynovisualsignsoffungus)couldleadtothemischaracterizationoftheinfectionstatus ofasiteandtheinadvertentspreadof P . destructans .Wedeterminedtheefficacyofvisual detectionof P . destructans byexaminingvisualsignsandmoleculardetectionof P . destructans on928batsofsixspeciesat27sitesduringsurveysconductedfromJanuarythrough Marchin2012 – 2014inthesoutheasternUSAontheleadingedgeofthediseaseinvasion. Crypticinfectionswerewidespreadwith77%ofbatsthattestedpositivebyqPCRshowing novisiblesignsofinfection.Theprobabilityofexhibitingvisualsignsofinfectionincreased withsamplingdateandpathogenload,thelatterofwhichwassubstantiallyhigherinthree species( Myotislucifugus , M . septentrionalis ,and Perimyotissubflavus ).Inaddition, M . lucifugus wasmorelikelytoshowvisualsignsofinfectionthanotherspeciesgiventhe samepathogenload.Nearlyallinfectionswerecrypticinthreespecies( Eptesicusfuscus , M . grisescens ,and M . sodalis ),whichhadmuchlowerfungalloads.Thepresenceof M . lucifugus or M . septentrionalis atasiteincreasedtheprobabilitythat P . destructans was visuallydetectedonbats.Ourresultssuggestthatcrypticinfectionsof P . destructans are commoninallbatspecies,andvisibleinfectionsrarelyoccurinsomespecies.However, duetoveryhighinfectionprevalenceandloadsinsomespecies,weestimatethatvisual surveysexaminingatleast17individualsof M . lucifugus and M . septentrionalis ,or29individualsof P . subflavus arestilleffectivetodeterminewhetherasitehasbatsinfectedwith P . destructans .Inaddition,becausetheprobabilityofvisuallydetectingthefunguswas higherlaterinwinter,surveysshouldbedoneasclosetotheendofthehibernationperiod aspossible. PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 1/12 OPENACCESS Citation: JanickiAF,FrickWF,KilpatrickAM,Parise KL,FosterJT,McCrackenGF(2015)Efficacyof VisualSurveysforWhite-NoseSyndromeatBat Hibernacula.PLoSONE10(7):e0133390. doi:10.1371/journal.pone.0133390 Editor: DaniloRusso,UniversitàdegliStudidiNapoli FedericoII,ITALY Received: February10,2015 Accepted: June26,2015 Published: July21,2015 Copyright: ©2015Janickietal.Thisisanopen accessarticledistributedunderthetermsofthe CreativeCommonsAttributionLicense ,whichpermits unrestricteduse,distribution,andreproductioninany medium,providedtheoriginalauthorandsourceare credited. DataAvailabilityStatement: Allrelevantdataare availablefromDryadDigitalRepository(doi: 10.5061/ dryad.8mm58 ). Funding: FundingwasprovidedbytheNational ScienceFoundationEcologyofInfectiousDiseases (DEB-1115895andDEB-1336290)toWFF,AMK, JTF,andGFM;andtheUniversityofTennessee DepartmentofEcologyandEvolutionaryBiology.The fundershadnoroleinstudydesign,datacollection andanalysis,decisiontopublish,orpreparationof themanuscript.

PAGE 2

IntroductionDiseasesurveillanceinwildlifeisoftenlimitedbydiagnostictechniquesthatarecost-effective, rapid,andfeasibleforuseonwildanimals[ 1 , 2 ].Fordiseaseswherehostsdisplayvisiblesymptoms,visualsurveysareoftencost-effectiveandcanbeappealingforsurveillancebecausethey typicallyimposeminimaldisturbanceonhostpopulations[ 3 , 4 ].However,ifhostshavecryptic infectionsthatarenotobservable,thenvisualsurveyswillhavelimitedutilityforreliablyidentifyinghabitatsharboringinfectedindividuals(aprimarygoalofdiseasesurveillance)andwill underestimateinfectionprevalence.Estimatingtheefficacyofvisualsurveysforaparticular diseaseisnecessarytodeterminewhetherthislow-costandminimallydisruptivesurvey methodisanappropriatesurveillanceapproach. White-NoseSyndrome(WNS)isarapidlyspreadingepizooticdiseasethathascausedwidespreaddeclinesinsixspeciesofhibernatingbatsinNorthAmerica,raisingsubstantialconcern abouttheriskofextirpationandextinctionofspecies[ 5 – 8 ].WNSiscausedbythefungalpathogen, Pseudogymnoascusdestructans [ 9 – 11 ],whichinfectsandkillsbatsduringhibernation [ 12 ]bydisruptingphysiology[ 13 – 15 ]andnaturaltorporarousalpatterns[ 10 , 16 ].Thedisease wasnamedWNSbecausethefacesandwingsofsomeinitiallydocumentedbatswerevisibly coveredinwhite,powderyfungalgrowth[ 17 ].Thediseasewasfirstdetectedinacavenear Albany,NewYorkin2006,andbythespringof2015WNShadbeenconfirmedinsevenspeciesofbatsin26U.S.statesandfiveCanadianprovinces[ 18 ].Althoughtheexactoriginof P . destructans remainsunclear,recentgeneticdatasuggestthefunguswasintroducedtoNorth AmericafromtheWesternPalearctic[ 19 , 20 ]. VisualsurveillanceforWNSisconductedinhundredsofcavesandmineseachyearandis theprimarysurveillancestrategyrecommendedbytheU.S.FishandWildlifeServiceWNS NationalResponsePlanandtheCanadianWildlifeHeathCooperativeWNSNationalPlan [ 21 , 22 ].SurveillanceforWNSconsistsprimarilyofsearchingforbatswithvisiblefungalinfectionsof P . destructans (e.g.visiblefungusonskintissues),andsubmittingbatswithsuspected infectionforlaboratorytestingbyhistopathology[ 23 ].Histopathologyisusedtoconfirmthe presenceofepidermalcuppingerosionsandlesionsonthewingmembranediagnosticofWNS disease[ 23 ].ReportingofhibernaculawithWNSisusedtotrackdiseasespreadaswellas informmanagementdecision-making,suchasrestrictinghumanaccesstositesorrequiring decontaminationprotocolstoreducepotentialspreadofthefungusbyhumans[ 24 ]. Batsbecomeinfectedwith P . destructans beforethefungusonskintissuesbecomesvisible tothehumaneye.Thesecrypticinfectionscouldeasilybemissedduringvisualsurveys,causing sitestobefalselyclassifiedas ‘ uninfected ’ wheninfactthepathogenispresentandbatsare infected.Falselyreportingasiteasnothavingbatsinfectedwith P . destructans couldleadto underestimatesoftheimpactofdiseaseonbatpopulations,andunrestrictedhumanaccess withoutdecontaminationcouldleadtoinadvertentspreadof P . destructans .FalsevisualdetectionsofP . destructans causedbyotherfungisuchas Trichophytonredellii [ 25 ]couldalsooccur andcouldleadtounnecessarykillingofbatsforsubmissionforhistopathology.Therecent developmentofaqPCRassay[ 26 ]todetect P . destructans DNAfromepidermalswabsamples frombatsprovidesanopportunitytodeterminetheaccuracyandefficacyofvisualsurveysfor detectingthepresenceofthepathogenathibernaculaandtheprevalenceofinfectionondifferentbatspecies.AlthougharangeofdifferentfactorscanaffectDNAquantityextractedfrom swabs(e.g.extractionefficiency),thisqPCRassayhasbeenshowntobebothhighlyspecificto P . destructans andhighlysensitive,makingitanaccurateandusefulmethodtodetermineif batsareinfectedandforestimatingprevalence[ 27 , 28 ]. Ourmainobjectivewastodeterminetheaccuracyofvisuallydetectinginfectionsof P . destructans atbathibernacula.Here,wedefineinfectionasthepresenceof P . destructans DNA EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 2/12 CompetingInterests: Theauthorshavedeclared thatnocompetinginterestsexist.

PAGE 3

detectedbyqPCRfromswabsamplescollectedfrombats.Weestimatedtheprobabilityoffailingtovisuallydetectinfectionsonbatsthattestedpositivefor P . destructans byqPCR(i.e.the probabilityofaninfectionbeingcryptic).Wehypothesizedthatcrypticinfectionswouldbe lesslikelyinbatswithhigherpathogenloads,andasaresult,crypticinfectionswouldbemore likelyinspecieswithlowerpathogenloads[ 12 ].Wealsocomparedwhetherthepresenceor absenceofparticularbatspeciesatahibernaculumincreasedtheprobabilityofvisuallydetecting P . destructans onbats.MaterialsandMethods SamplecollectionWeexaminedthepresenceof P . destructans insixspeciesin27hibernaculainfourstatesinthe southeasternUnitedStates( Fig1 )duringwinterhibernationfromJanuarythroughMarchin 2012 – 2014.Weswabbed928batsofsixspeciesoverthreeyearswithanaverageof22bats (range:5 – 50)ofonetosixspeciespresentineachhibernaculum.Batswereswabbedfivetimes ontheirmuzzleandforearmwithpolyester-tippedswabsdippedinsterilewater.Priorto Fig1.Mapofsamplecollection. Amapof27hibernaculainfourstateswherehibernatingbatsweresampledfromJanuary-Marchin2012 – 2014.Shading designatestheyearthatWNSandmolecularevidenceof P . destructans wereconfirmedinaU.S.county[ 18 ]. doi:10.1371/journal.pone.0133390.g001 EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 3/12

PAGE 4

swabbing,wenotedwhetherfunguswasvisibleonthebat ’ sskintissues(muzzle,ears,forearms, anduropatagium)whilethebatwasinhand.Allbatswerereleasedaftersamplingatthesite wheretheyhadbeenroosting.SwabswerestoredinRNAlatertopreserveDNAandkeptrefrigeratedorfrozenuntiltesting. AllbathandlingproceduresfollowedguidelinesapprovedbytheAmericanSocietyof MammalogistsandtheUniversityofTennesseeInstitutionalAnimalCareandUseCommittee. DecontaminationproceduresissuedbyU.S.FishandWildlifeServicewerefollowedforallcavinggear[ 24 ].PermitsforthisresearchwereobtainedfromMissouriDepartmentofConservation(15184,15471,and15871),TennesseeWildlifeResourcesAgency(3716),andU.S.Fish andWildlifeService(TE71613A-0).Otherbatsampleswerecollectedincollaborationwith stateagencypersonnelwithpermitsfromAlabamaWildlifeandFreshwaterFisheriesandKentuckyDepartmentofFishandWildlifeResources.SampletestingSwabsamplesandstandardswereextractedwithDNeasyBloodandTissueextractionkits (Qiagen,Valencia,CA)withmodificationsforfungalextractionsthatincludedtheadditionof lyticaseduringthelysisstep[ 28 ].Eachextractionplatehad16negativecontrolwells(100% P . destructans negative)distributedthroughouttheplate.DNAsampleswereanalyzedbyrealtimePCRusingmethodsdevelopedbyMulleretal.[ 26 ],usingacut-offof40cyclesforapositivedetection.Cyclethresholdvalues(Ctvalue)wereusedtocalculatefungalloads,innanograms,usingtheequationload=10((22.04942-Ctvalue)/3.34789),whichwasderivedfromserial dilutionsofaquantifiedstandardofisolate P . destructans 20631 – 21.Seventy-fivepercentof sampleswereruninduplicateandasamplewasconsidered P . destructans positiveifeitheror bothrunswerepositive.FungalloadswereaveragedacrossbothrunsafterconversionfromCt values.StatisticalanalysisVisualdetectionof P . destructans onbats. Weusedgeneralizedlinearmodelswitha binomialdistributiontodetermineiftheprobabilityofvisuallydetecting P . destructans ona batwasassociatedwithfungalload,whensamplingoccurred,andifdetectionprobabilitydifferedamongspecies.Weusedabias-reductionmethod(packagebrglminRv.3.1.2)todeal withthecompleteseparationpresentinthedata(insomespeciesnovisualdetectionsofthe fungusweremade).WeusedthenumberofdayssinceJanuary1sttoaccountfordifferencesin timingofsamplingasvisibilityofinfectionmayincreaselaterintheseason[ 27 , 29 , 30 ].Wefit twelve apriori modelswithcombinationsofmain,additive,andinteractiveeffectsrepresenting ourhypothesesandusedAkaikeinformationcriterion(AIC)modelselectioncriteriatodeterminethebest-fittingmodel.Weestimatedtheprobabilityoffalselydetectingvisualinfection usingbatsthattestednegativebyqPCRbutwerenotedwithvisiblewhitefungusinthefield. Wecomparedwhetherfalsedetectiondifferedamongspeciesusingalikelihoodratiotestto compareanullmodeltoonewithspeciesincluded. Visualsurveysforsite-leveldetectionof P . destructans onbats. Weusedgeneralizedlinearmodelswithbinomialdistributionsinwhicheachsitevisitwasadatapointtodetermine whethertimingofsurvey,samplingeffort,andspeciesofbatsexaminedinfluencedvisual detectionof P . destructans onbatsatasite.For Myotislucifugus , Myotisseptentrionalis ,and Perimyotissubflavus ,wealsodeterminedwhethertheprevalenceofinfectionofbatswithvisual infectionsinfluencedthelikelihoodofvisuallydetectingthefungusduringasitevisit.AllstatisticalanalyseswereconductedinProgramRv.3.1.2. EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 4/12

PAGE 5

Results Pathogenloadsandvisualdetectionof P . destructans onbatsSeventy-sevenpercent(306/397)ofbatsthattestedpositivefor P . destructans byqPCRhadno visiblesignsof P . destructans ,demonstratingthattheprobabilityoffalsenegatives(i.e.failing tovisuallydetect P . destructans onbatsthathadthepathogen)ishigh( Table1 ).TheprobabilityofobservingvisiblewhitefungusonabatthatwasqPCRnegativewaslow(14/531or2.6%) anddidnotdifferamongspecies(likelihoodratiotest: =5.10,df=5; P =0.40).Thebest-fittingmodeloftheprobabilityofvisualdetectionincludedfungalload,samplingdate,andan additivespecieseffect(AICweight=0.55;Fittedequationfor M . lucifugus =Pr(Detection)~ -12.9(±1.5)+1.77(±0.2)log10(load)+0.02(±0.01)(dayssinceJanuary1);For M . septentrionalis and P . subflavus theinterceptequaled-14.01(±1.6);Forthethreeotherspecies ( Eptesicusfuscus , Myotisgrisescens ,and Myotissodalis )theinterceptequaled-13.47(±1.6)), suggestingthattheprobabilityofvisuallydetecting P . destructans onabatincreasedwithpathogenloadmeasuredbyqPCR,buttheslopedidnotdifferamongspecies( Table2 , Fig2 ).The probabilityofvisuallydetecting P . destructans increasedwiththenumberofdayssinceJanuary 1standtherewasonlyweaksupportthatthiseffectdifferedamongspecies( Table2 ). Visibleinfectionsoccurredmostfrequentlyinthreespecies( M . lucifugus , M . septentrionalis , and P . subflavus )thathadthehighestfungalloadsand M . lucifugus hadasignificantlylower detectabilitythreshold(e.g.higherintercept)comparedto M . septentrionalis and P . subflavus , whichwerenotsignificantlydifferentfromeachother( Fig2 ).Loadsontheotherthreespecies ( E . fuscus , M . grisescens ,and M . sodalis )wereusuallytoolowtoresultinvisibleinfection ( Fig2 ).EfficacyofvisualsurveysathibernaculaFortypercent(17/43)ofsiteswhereatleastonebattestedpositivefor P . destructans byqPCR hadnobatswithvisualsignsof P . destructans andwouldhavebeenclassifiedas ‘ uninfected ’ basedsolelyonvisualsurveys.Thelikelihoodofdetectingthepresenceof P . destructans atasite withvisualsurveysincreasedwiththenumberofbatsexaminedforthethreespeciesthatfrequentlyexhibitvisualinfections( M . lucifugus , M . septentrionalis ,and P . subflavus )(Pr(Detection)~-0.90+0.12(±0.051)#mylu.myse.pesu.sampled;N=43; P =0.02),andtherewasvery weaksupportfortheinfluencebywhenavisitoccurredbetweenJanuaryandMarchorexaminationofotherspecies( Table3 ).Theprobabilityofvisualdetectionof P . destructans atasite increasedwithprevalenceofinfectionfor P . subflavus (Pr(Detection)~-1.1+3.13(±0.051)Prevalence;N=32; P < 0.01),butnotfor M . lucifugus (Pr(Detection)~-1.1+1.0(±1.8) Table1.Fractionofbatswithvisiblefungusonbatstestedfor Pseudogymnoascusdestructans by qPCR. SpeciesFractionofbatswithvisiblefungus qPCR+qPCREptesicusfuscus 0.10(1/10)0.0(0/30) Myotisgrisescens 0.04(1/26)0.02(5/201) Myotislucifugus 0.35(24/69)0.06(3/50) Myotisseptentrionalis 0.24(7/29)0.0(0/22) Myotissodalis 0.0(0/21)0.05(4/76) Perimyotissub avus 0.24(58/242)0.01(2/152) Samplesizesareshowninparentheses. doi:10.1371/journal.pone.0133390.t001 EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 5/12

PAGE 6

Prevalence;N=13; P =0.56)or M . septentrionalis ,thelatterofwhichhadaprevalenceof100% atallsites( Fig3 ).Visualsurveysthatincludeeither17 M . lucifugus or17 M . septentrionalis have a99%likelihoodofdetecting P . destructans ifitispresentatthesite.For P . subflavus ,examining atleast29batsisrequiredtohavea99%chanceofdetecting P . destructans ifitispresent.DiscussionOurresultssuggestthatcrypticinfectionsarewidespreadandthatsolelyusingvisiblesignsof P . destructans greatlyunderestimatesinfectionprevalenceinbatsevenduringmidtolatewinter(January-March)whenthemajorityofsurveillancesurveysforWNSareconducted.Cryptic infectionsweresocommoninsomespecies( E . fuscus , M . grisescens ,and M . sodalis )thatvisual surveyswereonlyusefulfordetecting P . destructans atasiteifotherspecies( M . lucifugus , M . septentrionalis ,and/or P . subflavus )alsowerepresentandexamined.Thehigherpercentagesof thelatterthreespeciesthatdisplayedvisible P . destructans ,combinedwiththehighinfection prevalenceinthesespecies,resultedinaveryhighlikelihoodthat P . destructans wasdetectedat asitewheneverthesebatspecieswerepresent. Ourresultsalsoshowthattheprobabilityofvisualdetectionincreaseswithfungalloadof P . destructans ,anddifferencesinfungalloadsamongspeciesexplainmostofthedifferencesin theprobabilityofobservingvisible P . destructans onbats.Thisislikelybecausehigherloads indicatealargernumberofconidiaandhyphaeonthebatsandagreaterlikelihoodofthefungusbeingvisible.Thisisconsistentwiththefindingthattheprobabilityofvisualdetectionof P . destructans onbatswashigherlaterinthehibernationseasonwhenthefungushashadsufficienttimetogrowonthebatsandisatmaximalloads[ 12 ],suggestingthatvisualsurveys shouldbescheduledlateinhibernationtobemaximallyeffective.Ourfindingsaresimilarto patternsofvisualprevalenceinEuropewherevisibleinfectionsalsopeakedinlatehibernation [ 29 ].Hibernationseasonlengthmayinfluencevisualdetectiongiventhatmostbatsbecome infectedatthestartofhibernationandfungalloadsincreaseoncebatsaretorpid[ 12 ].Thus, infectionsmaybecomevisiblesoonerinnorthernlatitudeswherebatslikelyenterhibernation earlier[ 31 ]. Evenwiththesamefungalload,somespeciesweremorelikelytoexhibitvisible P . destructans ( Fig2 ).Visible P . destructans wasdetectedatsignificantlylowerloadson M . lucifugus thanotherspecies( Fig2 ),perhapsbecausetheirdarkerskinprovidesmorevisualcontrastwith Table2.Modelselectionresultsforvisualdetectabilityof Pseudogymnoascusdestructans onbats. Model AICAICweights species+load+date 0.00.55 species * date+load2.00.20 species+date * load2.10.19 species+load5.20.04 load7.10.02 species * load+date9.70.00 species * load18.30.00 species * load * date29.00.00 species+date118.80.00 species128.60.00 date132.40.00 pd.visible.bat~null142.00.00 Modelsarerankedby AICandthebestttingmodelisshowninbold. doi:10.1371/journal.pone.0133390.t002 EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 6/12

PAGE 7

thewhitefungus. Myotislucifugus and M . septentrionalis ,whenpresent,arethebest ‘ sentinels ’ orindicatorsofthepresenceof P . destructans whensurveyingforvisiblesigns,andsurveying Fig2.Visualdetectabilityof Pseudogymnoascusdestructans onbatscomparedtofungalloads. Solidlinesshowpredictedrelationshipsfromthe best-fitmodel( Table2 )anddashedlinesshowthe95%confidencebandsforearly(January1st;bluelines)andlate(March31st;greenlines)samplingdates. Individualcirclesarebatsthattestedpositivefor P . destructans byqPCRanddid(y-axisvalueof1)ordidnot(0)havevisibleevidenceof P . destructans . doi:10.1371/journal.pone.0133390.g002 Table3.Modelselectionresultsforvisualdetectabilityof Pseudogymnoascusdestructans athibernacula. #mylu.myse.pesu.sampledreferstothesumofthenumberofbatsofthreespeciessampled( M . lucifugus – mylu, M . septentrionalis – myse, P . subflavus – pesu). Model AICAICweights #mylu.myse.pesu.sampled 0.00.56 date+#mylu.myse.pesu.sampled1.30.29 null4.90.05 all.bats.sampled5.00.05 all.bats.sampled+date5.60.03 date6.60.02 Modelsarerankedby AICandthebestttingmodelisshowninbold. doi:10.1371/journal.pone.0133390.t003 EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 7/12

PAGE 8

P . subflavus canalsobeuseful.Incontrast,fungalloadsin E . fuscus , M . grisescens ,and M . sodalis aresimplytoolowtoconsistentlyresultinvisible P . destructans .Differencesinfungalloads andinfectionintensityamongspeciessuggestsinterestingdifferencesineithertransmission, hibernatingbehaviors,and/ordiseasesusceptibilityamonghibernatingspeciesexposedto P . destructans [ 5 , 32 , 33 ]. Currently,visualsurveysareroutinelyusedtodeterminewhether P . destructans hasinvaded newhibernacula[ 21 , 22 ].Ourresultsshowthattheefficacyofthesevisualsurveysdependson whichspeciesarepresentatasiteandhowmanybatsareexaminedforvisiblefungus.For example,thepresenceof M . lucifugus or M . septentrionalis increasestheprobabilitythat P . destructans canbedetectedvisuallyatasiteandthatthesecanbeusedas ‘ sentinel ’ speciesfor thepresenceof P . destructans ( Fig3 ).Ourresultssuggestthatwithamoderatesurveyeffortof examiningeither20(ifsurveying M . lucifugus or M . septentrionalis )or30(ifonly P . subflavus Fig3.Detectionofvisible Pseudogymnoascusdestructans onbatsathibernaculaandthefractionofbatswith P . destructans atthatsiteas determinedbyqPCR. Circlesrepresentsiteswhereaspecieswassampled,withredcirclesindicatingsiteswhereatleastoneindividualofthatspecieshad visiblefungusandblackcirclesindicatingsiteswherenoindividualsofthatspecieswereobservedwithvisiblefungus.Thesizeofthecirclesissc aledtothe numberofbatssampledatasite.Thex-axisshowstheproportionofbatsthatwerepositivefor P . destructans byqPCRandthey-axisshowswhetherat leastoneindividualbatatthatsiteofanyspecieswasnegative(0)orpositive(1)forvisiblefungalinfections.Prevalenceofinfectionwasasigni ficant predictorfordetectionofvisibleinfectionsatasiteforasinglespecies, P . subflavus .Solidblacklineandgrayshadingfor P . subflavus representthebest-fit lineand95%confidencebandfortherelationshipbetweenprevalenceofinfectionanddetectionofvisibleinfectionsathibernacula. doi:10.1371/journal.pone.0133390.g003 EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 8/12

PAGE 9

areexamined)individualsatasite,thenvisualsurveyscanindeedbeeffectiveatdetermining whetherbatsareinfectedwith P . destructans atahibernaculum.Atsiteswithspeciesthatrarely orneverhavevisiblesignsof P . destructans ,suchas E . fuscus , M . grisescens ,or M . sodalis ,visual surveysareineffective.Toensurevisualsurveillanceiseffectiveatdeterminingwhether P . destructans hasinvadednewsites[ 21 , 22 ],futuresurveillanceguidelinesshouldincorporate thesespecificrecommendationsonspeciesandsamplesizesrequiredforeffectivesurveillance efforts. Thewidespreadoccurrenceofcrypticinfectionsinallspecieshasdirectrelevancetomanagementandsurveillanceofthisdisease[ 34 , 35 ].Visualsurveyscanbeaneffectiveandrelativelylow-costpartofsurveillanceactivities,especiallyinareaswhereroutinewintercolony countsarealreadyconducted[ 36 ],onlyaslongassitescontainsufficientnumbersofbats ( > 20)ofspeciesthatexhibitvisualinfections(e.g. M . lucifugus , M . septentrionalis ,and/or P . subflavus ).Further,visualsurveysofindividualbatsaremosteffectivelateinthehibernation season.However,fordetectionof P . destructans onspecieswithpredominatelycrypticinfectionsandtoaccuratelymeasureprevalence,swabsamplingandtestingsampleswithmolecular methodsareneeded[ 12 , 26 , 27 ].Ultraviolet(UV)illuminationhasrecentlybeenproposedfor WNSsurveillancebasedoncomparisonswithhistologicalexaminationofbatssubmittedfor testingbasedonvisualsignsofWNSandbatscollectedinareaswherethefungushasbeen presentforseveralyears[ 37 ].WedidnotexaminebatsunderUVilluminationandacomparisonofthismethodwithmoleculartestingofswabsampleswouldbeusefultodetermine whetherUVilluminationiseffectivefordetectingcrypticinfectionsontheleadingedgeoffungalinvasion.Currently,theU.S.FishandWildlifeServiceWNSNationalResponsePlanand theCanadianWildlifeHealthCooperativeWNSNationalPlansurveillanceprotocolsrely entirelyonvisualsurveillance[ 21 , 22 ],butourfindingssuggestthatcombiningswabsampling andvisualsurveyswouldimprovenationalsurveillanceofthisdisease. TherearecurrentlynoactivemanagementstrategiesforcontrolormitigationofWNS otherthancaveclosures[ 21 , 34 ].However,activitiessuchascullinghavebeenconsideredasa meanstopreventthespreadofthediseasetonewregions[ 38 ].Theoccurrenceofcrypticinfectionsdemonstratesthatcullingvisiblyinfectedbatswillbeineffectiveathaltingthespreadof P . destructans ,supportingearlymodelingefforts[ 38 ].Further,recentevidencesuggeststhat cullinginfectedindividuals,evenusingahighlysensitivemethod(e.g.qPCRonswabsamples), willbeineffectivebecauseP . destructans isoftenwidespreadintheenvironmentayearafter thefungusreachesasite,andcanpersistatsitesandintheabsenceofbatsforlongperiods[ 27 , 38 – 41 ].Ourfindingsthatcrypticinfectionscommonlyoccuratbathibernaculasuggestthat althoughthespreadof P . destructans acrossNorthAmericaisconsistentwithspreadbybats [ 42 – 44 ],restrictingrecreationalaccessandrequiringfieldhygieneprotocolstodecontaminate gearwillreducepotentialhuman-mediatedspread.AcknowledgmentsWethank3anonymousreviewersforinsightfulfeedbackonearlierdraftsandpersonnelfrom thefollowingagencies/organizationsforassistanceinthefieldandaccesstohibernacula:U.S. FishandWildlifeService,U.S.NationalParkService,TennesseeWildlifeResourcesAgency, TennesseeValleyAuthority,MissouriDepartmentofConservation,AlabamaWildlifeand FreshwaterFisheries,KentuckyDepartmentofFishandWildlifeResources,SoutheasternCave Conservancy,theNatureConservancy,theMissouriBatCensus,andEnvironmentalSolutions andInnovations.WethankDarwinBrackforassistancewithswabsamplecollection.We thankKevinDressandNicoletteJankeforlabworkonDNAextractionsandqPCR.Wethank DavidBlehertandJeffLorchforproviding P . destructans samplesforcreatingstandards. EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 9/12

PAGE 10

AuthorContributionsConceivedanddesignedtheexperiments:AFJWFFAMKGFM.Performedtheexperiments: AFJKLP.Analyzedthedata:AFJWFFAMKGFM.Wrotethepaper:AFJWFFAMKJTF GFM.References1. StallknechtDE.Impedimentstowildlifediseasesurveillance,research,anddiagnostics.In:ChildsJE, MackenzieJS,RichtJA,editors.Wildlifeandemergingzoonoticdiseases:thebiology,circumstances andconsequencesofcross-speciestransmission.Berlin:Springer;2007.pp.445 – 461. 2. SleemanJM.Hasthetimecomeforbigscienceinwildlifehealth?Ecohealth.2013;10:335 – 338.doi: 10.1007/s10393-013-0880-0 PMID: 24136386 3. HochachkaWM,DhondtAA.Density-dependentdeclineofhostabundanceresultingfromanewinfectiousdisease.ProcNatlAcadSciUSA.2000;97:5303 – 5306.PMID: 10792031 4. BlanchongJA,SamuelMD,GoldbergDR,ShadduckDJ,LehrMA.Persistenceof Pasteurellamultocida inwetlandsfollowingaviancholeraoutbreaks.JWildlDis.2006;42:33 – 39.PMID: 16699146 5. LangwigKE,FrickWF,BriedJT,HicksAC,KunzTH,KilpatrickAM.Sociality,density-dependenceand microclimatesdeterminethepersistenceofpopulationssufferingfromanovelfungaldisease,whitenosesyndrome.EcolLett.2012;15:1050 – 1057.doi: 10.1111/j.1461-0248.2012.01829.x PMID: 22747672 6. FrickWF,PuechmailleSJ,HoytJR,NickelBA,LangwigKE,FosterJT,etal.DiseasealtersmacroecologicalpatternsofNorthAmericanbats.GlobalEcolBiogeogr.2015;doi: 10.1111/geb.12290 7. FrickWF,PollockJF,HicksAC,LangwigKE,ReynoldsDS,TurnerGG,etal.Anemergingdisease causesregionalpopulationcollapseofacommonNorthAmericanbatspecies.Science.2010; 329:679 – 682.doi: 10.1126/science.1188594 PMID: 20689016 8. ThogmartinWE,Sanders-ReedCA,SzymanskiJA,McKannPC,PruittL,KingRA,etal.White-nose syndromeislikelytoextirpatetheendangeredIndianabatoverlargepartsofitsrange.BiolConserv. 2013;160:162 – 172. 9. LorchJM,MeteyerCU,BehrMJ,BoylesJG,CryanPM,HicksAC,etal.Experimentalinfectionofbats with Geomycesdestructans causeswhite-nosesyndrome.Nature.2011;480:376 – 379.doi: 10.1038/ nature10590 PMID: 22031324 10. WarneckeL,TurnerJM,BollingerTK,LorchJM,MisraV,CryanPM,etal.InoculationofbatswithEuropean Geomycesdestructans supportsthenovelpathogenhypothesisfortheoriginofwhite-nosesyndrome.ProcNatlAcadSciUSA.2012;109:6999 – 7003.doi: 10.1073/pnas.1200374109 PMID: 22493237 11. MinnisAM,LinderDL.Phylogeneticevaluationof Geomyces andalliesrevealsnocloserelativesof Pseudogymnoascusdestructans ,comb.nov.,inbathibernaculaofeasternNorthAmerica.Fungal Biol.2013;117:638 – 649.doi: 10.1016/j.funbio.2013.07.001 PMID: 24012303 12. LangwigKE,FrickWF,ReynoldsR,PariseKL,DreesKP,HoytJR,etal.Hostandpathogenecology drivetheseasonaldynamicsofafungaldisease,white-nosesyndrome.ProcRSocBBiolSci.2015; 282:20142335. 13. WarneckeL,TurnerJM,BollingerTK,MisraV,CryanPM,BlehertDS,etal.Pathophysiologyofwhitenosesyndromeinbats:amechanisticmodellinkingwingdamagetomortality.BiolLett.2013; 9:20130177.doi: 10.1098/rsbl.2013.0177 PMID: 23720520 14.CryanPM,MeteyerCU,BlehertDS,LorchJM,ReederDM,TurnerGG,etal.Electrolytedepletionin white-nosesyndromebats.JWildlDis.2013;49:398 – 402.doi: 10.7589/2012-04-121 PMID: 23568916 15. VerantML,MeteyerCU,SpeakmanJR,CryanPM,LorchJM,BlehertDS.White-nosesyndromeinitiatesacascadeofphysiologicdisturbancesinthehibernatingbathost.BMCPhysiol.2014;14:10.doi: 10.1186/s12899-014-0010-4 PMID: 25487871 16. ReederDM,FrankCL,TurnerGG,MeteyerCU.Frequentarousalfromhibernationlinkedtoseverityof infectionandmortalityinbatswithWhite-NoseSyndrome.PLoSOne.2012;7:e38920.doi: 10.1371/ journal.pone.0038920 PMID: 22745688 17. BlehertDS,HicksAC,BehrM,MeteyerCU,Berlowski-ZierBM,BucklesEL,etal.BatWhite-NoseSyndrome:Anemergingfungalpathogen?Science.2009;323:227.doi: 10.1126/science.1163874 PMID: 18974316 18. U.S.FishandWildlifeService.White-nosesyndromemap.2015.Available: http://www. whitenosesyndrome.org/resources/map EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 10/12

PAGE 11

19. PuechmailleSJ,FrickWF,KunzTH,RaceyPA,VoigtCC,WibbeltG,etal.White-nosesyndrome:is thisemergingdiseaseathreattoEuropeanbats?TrendsEcolEvol.2011;26:570 – 576.doi: 10.1016/j. tree.2011.06.013 PMID: 21835492 20. LeopardiS,BlakeD,PuechmailleSJ.White-NoseSyndromefungusintroducedfromEuropetoNorth America.CurrBiol.2015;25:R217 – R219.doi: 10.1016/j.cub.2015.01.047 PMID: 25784035 21. U.S.FishandWildlifeService.Anationalplanforassistingstates,federalagencies,andtribesinmanagingWhite-NoseSyndromeinbats.2011.Available: https://www.whitenosesyndrome.org/sites/ default/files/white-nose_syndrome_national_plan_may_2011.pdf 22. CanadianWildlifeHealthCooperative.AnationalplantomanageWhiteNoseSyndromeinbatsinCanada.2015.Available: http://www.cwhc-rcsf.ca/docs/BatWhiteNoseSyndrome-NationalPlan.pdf . 23. MeteyerCU,BucklesEL,BlehertDS,HicksAC,GreenDE,Shearn-BochslerV,etal.Histopathologic criteriatoconfirmwhite-nosesyndromeinbats.JVetDiagnInvest.2009;21:411 – 414.PMID: 19564488 24. ShelleyV,KaiserS,ShelleyE,WilliamsT,KramerM,HamanK,etal.Evaluationofstrategiesforthe decontaminationofequipmentfor Geomycesdestructans ,thecausativeagentofWhite-NoseSyndrome(WNS).JCaveKarstStud.2013;75:1 – 10. 25. LorchJM,MinnisAM,MeteyerCU,RedellJA,WhiteJP,KaarakkaHM,etal.Thefungus Trichophyton redellii sp.nov.causesskininfectionsthatresembleWhite-NoseSyndromeofhibernatingbats.JWildl Dis.2015;51:36 – 47.doi: 10.7589/2014-05-134 PMID: 25375940 26. MullerLK,LorchJM,O ’ ConnorM,GargasA,BlehertDS.Batwhite-nosesyndrome:areal-timeTaqManpolymerasechainreactiontesttargetingtheintergenicspacerregionof Geomycesdestructans . Mycologia.2013;105:253 – 259.doi: 10.3852/12-242 PMID: 22962349 27. LangwigKE,HoytJR,PariseKL,KathJ,KirkD,FrickWF,etal.Diseasedynamicsofwhite-nosesyndromeinvasion,MidwestUSA.EmergInfectDis.2015;21:1023 – 1026.doi: 10.3201/eid2106.150123 PMID: 25989230 28. ShueyMM,DreesK,LindnerDL,KeimP,FosterJT.AhighlysensitiveqPCRassayforthedetection anddifferentiationof Pseudogymnoascusdestructans and Pseudogymnoascus species.ApplEnviron Microbiol.2014;80:1726 – 1731.doi: 10.1128/AEM.02897-13 PMID: 24375140 29. PuechmailleSJ,WibbeltG,KornV,FullerH,ForgetF,MühldorferK,etal.Pan-Europeandistributionof white-nosesyndromefungus( Geomycesdestructans )notassociatedwithmassmortality.PLoSOne. 2011;doi: 10.1371/journal.pone.0019167 30. SachanowiczK,St pie A,CiechanowskiM.Prevalenceandphenologyofwhite-nosesyndromefungus PseudogymnoascusdestructansinbatsfromPoland.CentEurJBiol.2014;doi: 10.2478/s11535013-0280-z 31. NorquayKJO,WillisCKR.Hibernationphenologyof Myotislucifugus .JZool.2014;294:85 – 92. 32. FrankCL,MichalskiA,McDonoughAA,RahimianM,RuddRJ,HerzogC.TheresistanceofaNorth Americanbatspecies( Eptesicusfuscus )toWhite-NoseSyndrome.PLoSOne.2014;doi: 10.1371/ journal.pone.0113958 33. JohnsonJS,ReederDM,LilleyTM,CzirjákGÁ,VoigtCC,McMichaelJW,etal.Antibodiesto Pseudogymnoascusdestructans arenotsufficientforprotectionagainstwhite-nosesyndrome.EcolEvol. 2015;5:2203 – 2214.doi: 10.1002/ece3.1502 PMID: 26078857 34. LangwigKE,VoylesJ,WilberMQ,FrickWF,MurrayKA,BolkerBM,etal.Context-dependentconservationresponsestoemergingwildlifediseases.FrontEcolEnviron.2015;13:195 – 202. 35. FoleyJ,CliffordD,CastleK,CryanPM,OstfeldRS.Investigatingandmanagingtherapidemergence ofwhitenosesyndrome,anovel,fatal,infectiousdiseaseofhibernatingbats.ConservBiol.2011; 25:223 – 231.doi: 10.1111/j.1523-1739.2010.01638.x PMID: 21284732 36. LoebSC,RodhouseTJ,EllisonLE,LausenCL,ReichardJD,IrvineKM,etal.AplanfortheNorth Americanbatmonitoringprogram(NABat).USDAForestServiceResearchandDevelopmentSouthernResearchStation.2015.Available: http://www.srs.fs.usda.gov/pubs/gtr/gtr_srs208.pdf . 37. TurnerGG,MeteyerCU,BartonH,GumbsJF,ReederDM,OvertonB,etal.Nonlethalscreeningof bat-wingskinwiththeuseofultravioletfluorescencetodetectlesionsindicativeofwhite-nosesyndrome.JWildlDis.2014;50:566 – 573.doi: 10.7589/2014-03-058 PMID: 24854396 38. HallamTG,McCrackenGF.ManagementofthepanzooticWhite-NoseSyndromethroughcullingof bats.ConservBiol.2011;25:189 – 194.doi: 10.1111/j.1523-1739.2010.01603.x PMID: 21054529 39. LorchJM,MullerLK,RussellRE,O'ConnorM,LindnerDL,BlehertDS.Distributionandenvironmental persistenceofthecausativeagentofwhite-nosesyndrome, Geomycesdestructans ,inbathibernacula oftheEasternUnitedStates.ApplEnvironMicrobiol.2013;79:1293 – 1301.doi: 10.1128/AEM.0293912 PMID: 23241985 EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 11/12

PAGE 12

40. HoytJR,OkoniewskiJ,LangwigKE,FrickWF,StoneWB,KilpatrickAM.Long-termpersistenceof Pseudogymnoascusdestructans ,thecausativeagentofwhite-nosesyndrome,intheabsenceofbats. Ecohealth.2014;doi: 10.1007/s10393-014-0981-4 41. ReynoldsHT,IngersollT,BartonHA.Modelingtheenvironmentalgrowthof Pseudogymnoascus destructans anditsimpactonthewhite-nosesyndromeepidemic.JWildlDis.2015;51:318 – 331.doi: 10.7589/2014-06-157 PMID: 25588008 42. WilderAP,FrickWF,LangwigKE,KunzTH.Riskfactorsassociatedwithmortalityfromwhite-nosesyndromeamonghibernatingbatcolonies.BiolLett.2011;7:950 – 953.doi: 10.1098/rsbl.2011.0355 PMID: 21632616 43. MaherSP,KramerAM,PulliamJT,ZokanMA,BowdenSE,BartonHD,etal.Spreadofwhite-nosesyndromeonanetworkregulatedbygeographyandclimate.NatCommun2012;3:1306 – 1308.doi: 10. 1038/ncomms2301 PMID: 23250436 44. O ’ ReganSM,MagoriK,PulliamJT,ZokanMA,KaulRRB,BartonHD,etal.Multi-scalemodelofepidemicfadeout:willlocalextirpationeventsinhibitthespreadofwhite-nosesyndrome?EcolAppl.2015; 25:621 – 633. EfficacyofVisualSurveysforWNS PLOSONE|DOI:10.1371/journal.pone.0133390July21,2015 12/12


printinsert_linkshareget_appmore_horiz

Download Options

close


  • info Info

    There are only PDFs associated with this resource.

  • link PDF(s)



Cite this item close

APA

Cras ut cursus ante, a fringilla nunc. Mauris lorem nunc, cursus sit amet enim ac, vehicula vestibulum mi. Mauris viverra nisl vel enim faucibus porta. Praesent sit amet ornare diam, non finibus nulla.

MLA

Cras efficitur magna et sapien varius, luctus ullamcorper dolor convallis. Orci varius natoque penatibus et magnis dis parturient montes, nascetur ridiculus mus. Fusce sit amet justo ut erat laoreet congue sed a ante.

CHICAGO

Phasellus ornare in augue eu imperdiet. Donec malesuada sapien ante, at vehicula orci tempor molestie. Proin vitae urna elit. Pellentesque vitae nisi et diam euismod malesuada aliquet non erat.

WIKIPEDIA

Nunc fringilla dolor ut dictum placerat. Proin ac neque rutrum, consectetur ligula id, laoreet ligula. Nulla lorem massa, consectetur vitae consequat in, lobortis at dolor. Nunc sed leo odio.