bagworm bags as portable armour against invertebrate predators · bags are generally known to...

Submitted 18 August 2015Accepted 22 January 2016Published 15 February 2016

Corresponding authorShinji Sugiura sug-iurashinjigmailcom

Academic editorSusan Bertram

Additional Information andDeclarations can be found onpage 11

DOI 107717peerj1686

Copyright2016 Sugiura

Distributed underCreative Commons CC-BY 40

OPEN ACCESS

Bagworm bags as portable armouragainst invertebrate predatorsShinji SugiuraGraduate School of Agricultural Science Kobe University Kobe Japan

ABSTRACTSome animals have evolved the use of environmental materials as lsquolsquoportable armourrsquorsquoagainst natural enemies Portable bags that bagworm larvae (Lepidoptera Psychidae)construct using their own silk and plant parts are generally believed to play an importantrole as a physical barrier against natural enemies However no experimental studieshave tested the importance of bags as portable armour against predators To clarify thedefensive function I studied the bagworm Eumeta minuscula and a potential predatorCalosoma maximoviczi (Coleoptera Carabidae) Under laboratory conditions allbagworm larvae were attacked by carabid adults but successfully defended themselvesagainst the predatorsrsquo mandibles using their own bags The portable bags which arecomposedmainly of host plant twigsmay function as a physical barrier against predatormandibles To test this hypothesis I removed the twig bags and replaced somewith herbleaf bags all bag-removed larvae were easily caught and predated by carabids whileall bag-replaced larvae could successfully defend themselves against carabid attacksTherefore various types of portable bags can protect bagworm larvae from carabidattacks This is the first study to test the defensive function of bagworm portable bagsagainst invertebrate predators

Subjects Animal Behavior Ecology Entomology Evolutionary Studies ZoologyKeywords Carabidae Physical defense Portable cases Predation Psychidae

INTRODUCTIONAnimals have evolved defensive armour to protect themselves from predators for examplearmadillos and crabs have hardened their exoskeletons hedgehogs and sticklebackshave developed spines and snails have developed shells as defensive armour (Edmunds1974 Eisner 2003 Emlen 2014) Conversely many animals have evolved the use ofenvironmentalmaterials as defensive armour (Edmunds 1974) For example phytophagousinsects accumulate host plant secondary metabolites in their bodies to defend themselveschemically against their natural enemies (eg Eisner Esiner amp Siegler 2005) and hermitcrabs use gastropod shells as lsquolsquoportable armourrsquorsquo against predators (Edmunds 1974)

The larvae of holometabolous insects are vulnerable to enemy attacks because of their softbodies and have developed various types of defensive armour (Greeney Dyer amp Smilanich2012) For example the spines and hairs of caterpillars constitute physical defences againstpredators (Dyer 1995 Dyer 1997 Murphy et al 2009 Sugiura amp Yamazaki 2014) Someinsect larvae construct lsquolsquoportable casesrsquorsquo using their own silk thread excrement andorenvironmental materials (eg plant parts and stones) Such case-bearing behaviour hasbeen found in three holometabolous insect orders (Root amp Messina 1983) Trichoptera

How to cite this article Sugiura (2016) Bagworm bags as portable armour against invertebrate predators PeerJ 4e1686 DOI107717peerj1686

(eg caddisfly larvae of the suborder Integripalpia Holzenthal et al 2007) Coleoptera(eg leaf beetle larvae of the subfamilies Clytrinae Cryptocephalinae Chlamisinae andLamprosomatinae Brown amp Funk 2005 Chaboo Brown amp Funk 2008) and Lepidoptera(eg moth larvae of the superfamilies Incurvarioidea and Tineoidea Stehr 1987) Physicaldefence against predators using portable cases has been tested experimentally in Trichoptera(Otto amp Svensson 1980 Ferry et al 2013) and Coleoptera (Root amp Messina 1983 Brown ampFunk 2010) but not in Lepidoptera

The bagworm family Psychidae (Lepidoptera Tineoidea) includes ca 1000 species andall of their larvae construct portable cases (Rhainds Davis amp Price 2009) The materialsused for constructing bags differ among bagworm species eg treeherbgrass leaveslichens twigs petioles bark fragments wood debris and sand particles (Sugimoto 2009aSugimoto 2009b) The portable bags are generally believed to play an important role asportable armour against natural enemies (Rhainds Davis amp Price 2009) For example bagshave been reported to function as a physical barrier against parasitoid attack the ovipositorof an ichneumonid parasitoid was too short to reach pupae of the bagworm Thyridopteryxephemeraeformis (Haworth) inside the larger bags and the parasitism rate was inverselycorrelated with bag size (Cronin amp Gill 1989) However bagworm larvae and pupae insidebags are generally known to suffer heavier parasitism by more diverse parasitoids thanare other external-feeding caterpillars (Hawkins 1994) suggesting that bagworm bagsmay not be effective armour against parasitoids Rather predators such as birds andpredacious arthropods may impose a selective pressure on the evolution or maintenanceof bags Although the impacts of predators have been reported in some bagworm species(Rhainds Davis amp Price 2009 Pierre amp Idris 2013) no experimental studies have testedthe importance of bags and materials used for bags as defensive armour against predatorsClarifying the defensive function of bags would contribute to further understanding ofhow portable armour has evolved in animals

To test whether portable bags can protect bagworms from predator attacks Calosomaadults (Coleoptera Carabidae) were observed attacking larvae of a bagworm species underlaboratory conditions Adults of the carabid genus Calosoma hunt lepidopteran larvae andpupae (Forsythe 1982Weseloh 1985 Bruschi 2013) providing a good model predator forinvestigating the defensive behaviour of lepidopteran larvae (Sugiura amp Yamazaki 2014)In this study I first investigated the defensive success or failure of bagworm larvae againstcarabid attacks Second I tested whether bag-removed larvae could defend themselvesagainst carabids in order to clarify the importance of bags Furthermore I investigated theeffects of bag replacement (with a different type of bag) on the defensive success of bagwormlarvae against carabid attacks to elucidate the importance of materials for constructingbags

MATERIALS AND METHODSStudy speciesTo clarify the defensive function of portable bags I used the bagworm species Eumetaminuscula Butler (Psychidae) and the potential predator Calosoma maximoviczi Morawitz(Carabidae)

Sugiura (2016) PeerJ DOI 107717peerj1686 214

Larvae of E minuscula feed on leaves of various woody species including bothangiosperms and gymnosperms and construct portable bags using their own silk thread andleaf fragmentspetiolestwigs of host plants (Figs 1A and 1B Kobayashi amp Taketani 1993Sugimoto 2009b) In Japan E minuscula overwinters asmiddle-instar larvae and pupates inearly summer (Kobayashi amp Taketani 1993) Various natural enemies are known to attackE minuscula larvae and pupae inside the bags (Kobayashi amp Taketani 1993) including 25parasitoid wasp species (Nishida 1983) three parasitoid fly species (Shima 1999) one antspecies (Nishida 1983) and one bird species (Ikeda 1988) For laboratory experiments allE minuscula larvae were collected from the forest edge in Shimosasori Takarazuka Hyogo(3455primeN 13518primeE 190 m above sea level) in late May 2015 Active larvae were used inlaboratory experiments although unhatched eggs of parasitoid flies (Diptera Tachinidae)were found on some active larvae Before the experiments I measured the fresh weightof each E minuscula larva and its bag to the nearest 01 mg using an electronic balance(PA64JP Ohaus Tokyo Japan) I also used slide callipers to measure the bag length larvallength and head capsule width of E minuscula to the closest 01 mm Sampled larvaewere determined to be 6th or 7th (last) instar based on the head capsule width (range25ndash38 mm cfNishida 1983) The bags were ca 18 times the length of the larvae (Fig 1Bmean larval body length 177 plusmn 25 mm (mean plusmn SD) mean bag length 324 plusmn 49 mmn= 36) and as heavy as larvae (mean fresh larval weight 2453 plusmn 711 mg mean fresh bagweight 2450 plusmn 744 mg n= 36) All bags were composed mainly of plant twigs (Figs 1Aand 1B)

Calosoma maximoviczi adults exclusively hunt lepidopteran larvae on both the groundand vegetation (Kamata amp Igarashi 1995 Sugiura amp Yamazaki 2014) This carabid speciesuses its mandibles to catch and injure caterpillars and then feeds on them (Sugiura ampYamazaki 2014) Since C maximoviczi adults can attack caterpillars of various speciesand size under laboratory conditions C maximoviczi adults are considered appropriatefor investigating the defence behaviour of lepidopteran larvae against generalist predators(Sugiura amp Yamazaki 2014) For laboratory experiments all adults of C maximovicziwere collected from a secondary forest in Nunobiki Kobe Hyogo (3442primeN 13411primeE60ndash170 m above sea level) in early May 2015 I have not observed C maximoviczi adultsattacking bagworms under field conditions however the habitat and active season partlyoverlap between E minuscula larvae and C maximoviczi adults in this sampling regionsuggesting that E minuscula larvae can encounter C maximoviczi adults on trunks ortwigs of woody plants Active adults of C maximoviczi which attacked caterpillars underlaboratory conditions were used in the laboratory experiments Before the experiments Imeasured the fresh weight of each adult of C maximoviczi to the nearest 01 mg using anelectronic balance I also used slide callipers to measure the body and mandible lengths ofC maximoviczi to the closest 01 mm

The insects used in this study were not endangered or protected species in the samplingregion The experiments were undertaken according to the Kobe University AnimalExperimentation Regulations The experiments also comply with the current laws ofJapan


Figure 1 Photos of the bagworm Eumeta minuscula and its potential predator Calosoma maximoviczi(A) Eumeta minuscula bags on shrubs (B) An E minuscula larva and the inside of its bag (C) A bagwormand a carabid on bamboo material under laboratory conditions (D) A bag protecting the larva from acarabid attack (E) A bag-removed larva eaten by a carabid (F) A replaced bag protecting the larva from acarabid attack

Laboratory experimentsTo test the defensive function of portable bags I conducted the following experiment ina well-lit laboratory (25 C) in late May 2015 A bagworm larva and a carabid adult wereplaced on bamboo material (width 7 mm height 15 mm Figs 1C and 2) which modelledtree twigs and trunks because both carabids and bagworms forage on tree twigs and trunksunder field conditions The bamboo material was looped (Fig 2 length 700 mm diameter


Figure 2 The arena used in the experiments A Eumeta minuscula larva and a Calosoma maximovicziadult were placed on bamboo material

200 mm) so that bagworms could encounter carabids in all trials The looped bamboomaterial was also surrounded by a plastic circular cylinder (diameter 220 mm height120 mm)

During a 10-min period I observed (1) whether a carabid attacked a bagworm larvaand (2) whether the carabid finally injured the bagworm I deemed that a bagworm larvacould not defend itself against a carabid adult when the carabid was observed to catchand injure the larva within the 10-min period When an adult carabid gave up attackinga bagworm without injuring it I deemed that the bagworm successfully defended itselfagainst the carabid I also continued observing further attacks by the carabid within the10-min period I used 15 adults (4 females and 11 males) of C maximoviczi (mean plusmn SDbody weight 4471plusmn 1047 mg mean body length 251plusmn 20 mm mean mandible length19 plusmn 01 mm n= 15) to conduct three types of experiments (Table 1) body weightand length significantly differed among three types of experiments (one-way analyses ofvariance body weight F = 43 P = 004 body length F = 41 P = 004) while mandiblelength did not differ (F = 09 P = 044)

Experiment 1 Normal E minuscula larvae (bag treatment control) were provided asthe first prey to five adult C maximoviczi (Table 1) To clarify the importance of bags asa defensive barrier against predators I provided bag-removed E minuscula larvae (bagtreatment removed) as the second prey to the same five carabid individuals just after theexperiment with the first prey (Table 1) I used a pair of scissors to remove the bags fromE minuscula larvae first prey that successfully defended itself against carabid attacks wasalso used as the second prey To investigate which bagworm individuals or bag treatmentscould affect the defence success of bagworms I used the same bagworms in differenttreatments The removal of bags from middle- and late-instar bagworms has been known


Table 1 Defensive success or failure of the bagworm Eumeta minuscula against the potential predator Calosoma maximoviczi under laboratory conditions

Predator (C maximowiczi) First prey (E minuscula) Second prey (E minuscula)a

Nob Sex Weight(mg)

No b Bagtreatmentc

Weight(mg)d

Defencee Numbers ofattacksf

Nob Bagtreatmentc

Weight(mg)d


Experiment 1C1 Male 3844 E1 Control 7113 Success 1 E1 Removed 2635 Failure 1C2 Male 4263 E2 Control 4366 Success 4 E2 Removed 2265 Failure 1C3 Male 3433 E3 Control 4853 Success 3 E3 Removed 2541 Failure 1C4 Female 5304 E4 Control 2858 Success 4 E4 Removed 1381 Failure 1C5 Male 6042 E5 Control 6992 Success 1 E5 Removed 3267 Failure 1

Experiment 2C6 Female 4209 E6 Removed 3382 Failure 1 E11 Control 2368 Success 3C7 Male 5721 E7 Removed 2185 Failure 1 E12 Control 5057 Success 1C8 Male 6004 E8 Removed 2384 Failure 1 E13 Control 3305 Success 1C9 Male 4463 E9 Removed 1183 Failure 1 E14 Control 6180 Success 1C10 Female 5666 E10 Removed 1647 Failure 1 E15 Control 3771 Success 1

Experiment 3C11 Male 3690 E16 Replaced 4273 Success 2 E16 Removed 2304 Failure 1C12 Male 4184 E17 Replaced 4794 Success 3 E17 Removed 3590 Failure 1C13 Female 3729 E18 Replaced 3409 Success 3 E18 Removed 2054 Failure 1C14 Male 3990 E19 Replaced 4496 Success 1 E19 Removed 3368 Failure 1C15 Male 2527 E20 Replaced 3135 Success 4 E20 Removed 2028 Failure 1

NotesaThe second prey was provided to each predator after my observation of the predator behaviour in response to the first preybDifferent code numbers showed that different individuals were usedcBag treatment control normal bags removed bags were removed experimentally replaced normal (twig) bags were replaced with soft (herb leaf) bags (see text)dTotal fresh weight (including bags) was shown for control and bag-replaced larvae while fresh body weight (except bags) was measured for bag-removed larvaeeDefence success and failure of E minuscula indicated predation failure and success by C maximoviczi respectivelyfTotal number of attacks by C maximoviczi on E minuscula Two or more attacks indicated that a carabid attacked a bagworm again after giving up its first attack

Sugiura(2016)PeerJD

OI107717peerj1686

614

to render bagworms susceptible to drought and starvation eg bag-removed larvae diedafter several days (Kaufmann 1968) However my preliminary observations showed thatbag-removed E minuscula did not die within the 10-min period due to drought andstarvation

Experiment 2 Bag-removed E minuscula larvae were provided as the first prey to fiveadult C maximoviczi (Table 1) I provided control E minuscula larvae as the second preyto the same five carabids just after the experiment with the first prey (Table 1) Different E minuscula larvae were used as the second prey I conducted this experiment to avoid anypotential systematic effects of the first prey on responses to the second prey by carabids

Experiment 3 Bag-replaced E minuscula larvae (bag treatment replaced) were providedas the first prey to five adultC maximoviczi (Table 1) To clarify the importance ofmaterialsfor constructing bags I replaced the normal (tight) bags with soft bags I used a pair ofscissors to remove the bags from 10 E minuscula larvae The larvae were placed individuallyin plastic Petri dishes (90mmdiameter 30mmhigh) withminced leaves of the herb speciesArtemisia indica varmaximowiczii (Asteraceae) I used a pair of scissors to mince the leaves(mean fragment length 44 plusmn 17 mm n= 27) Five of 10 E minuscula larvae constructedsufficiently large bags (bag length gt 25 mm) using their own silk thread and the leaffragments (Fig 3) one day after placement The replaced bags were ca 15 times the lengthof the larvae (mean larval body length 186 plusmn 27 mm mean bag length 279 plusmn 12 mmn= 5) and half as heavy as larvae (mean fresh larval weight 2669 plusmn 751 mg mean freshbag weight 1353plusmn 358 mg n= 5) Such replacement with a different type of bag has beenconducted in another bagworm species (Kaufmann 1968) Five larvae constructing newbags were used as the first prey in this experiment Just after conducting the experimentwith the first prey (ie bag-replaced larvae) I provided bag-removed E minuscula larvaeas the second prey to the same five carabids (Table 1) The first prey that had successfullydefended itself against carabid attacks was also used as second prey To investigate whichbagworm individuals or bag treatments could affect the defence success of bagworms Iused the same bagworms in different treatments

All adult carabids attacked each bagworm within the 10-min period Even whenbagworms did not actively walk carabids were observed to attack and bite motionlessbags Larval weight bag weight total (larval+ bag) weight bag length and larval length ofthe E minuscula used in this study did not differ among the three experiments (one-wayanalyses of variance F = 02ndash11 P = 038ndash084)

Fisherrsquos exact testswere used to compare the success rate of defence by bagworms betweencontrol bag-removed and bag-replaced treatments Considering the independence of thedata I excluded the data for the second prey from the analysis All analyses were performedusing R ver 2151 (R Development Core Team 2012)

RESULTSExperiment 1 all control E minuscula larvae (n= 5) were attacked by C maximovicziadults but successfully defended themselves against the predator attacks (Table 1 Figs 1Cand 1D) When bagworm larvae were attacked by carabids the larvae quickly retracted


Figure 3 Predation success of the carabid Calosoma maximoviczi and defensive success of the bag-worm Eumeta minuscula for different bag treatments (control bag-removal and bag-replacement)

their heads and thoraxes into their bags to escape from the attacks (Fig 1D Movie S1)Carabids frequently bit the bags but could not injure the larvae due to the bag protection(Fig 1D) Finally all of the carabids gave up attacking the larvae Three of five bagwormlarvae were attacked by carabids again within the 10-min period but successfully defendedthemselves against further attacks (Table 1) The other (two) bagworms remained retractedafter the first carabid attack and were not attacked again (Table 1) All of the bag-removedlarvae were easily caught and injured by the same individual carabids (Fig 1E and Table 1Movie S1) The dorsal lateral or ventral abdomens of larvae were the locations injured bycarabid mandiblesExperiment 2 all bag-removed larvae (n= 5) were easily caught and predated by carabids(Table 1) All control larvae (n= 5) were attacked by the same individual carabids butsuccessfully defended themselves against the attacks due to bag protection (Table 1) Onebagworm was attacked by the carabid again within the 10-min period but successfullydefended itself against further attacks (Table 1) Other bagworms remained retracted afterthe first carabid attack and were not attacked again (Table 1)

Experiment 3 all bag-replaced larvae (n= 5) were attacked by carabids but successfullydefended themselves against the attacks (Fig 1F and Table 1) Carabids frequently bitthe soft bags but could not injure the larvae due to the bag protection (Movie S1)Four of five bagworms were attacked by carabids again within the 10-min period butsuccessfully defended themselves against further attacks (Table 1) The other bagwormremained retracted after the first carabid attack and was not attacked again (Table 1) Allthe bag-removed larvae were easily predated by the same individual carabids (Table 1)

The success rate of bagwormdefence differed significantly among bag treatments (Fig 3)the defensive success rate of control bag-removed and bag-replaced larvae was 100 0and 100 respectively (Table 1 Fisherrsquos exact test control vs bag-removal P = 00008control vs bag-replacement P = 10 bag-removal vs bag-replacement P = 00008)


DISCUSSIONPortable cases of bagworms are generally believed to play an important role as a physicaldefence against natural enemies (Rhainds Davis amp Price 2009) however no studies havetested their effectiveness experimentally This study demonstrated that bags could protectE minuscula larvae from C maximoviczi attacks (Table 1 and Fig 3) This is the firststudy to test the defensive function of portable cases against invertebrate predators inLepidoptera Although the bag defence of a single bagworm species was shown in thisstudy my experiment showed that bags made from two different materials (ie twigand herb leaf bags) could effectively defend bagworms against the predator (Table 1and Fig 3) Accordingly bags made of other materials may also function as defensivearmour against invertebrate predators although further studies are needed Studies haveclarified the defensive function of portable cases in the two holometabolous insect ordersTrichoptera (Otto amp Svensson 1980 Ferry et al 2013) and Coleoptera (Root amp Messina1983 Brown amp Funk 2010) Case-bearing behaviours are considered to have evolvedindependently in Trichoptera and Lepidoptera (Holzenthal et al 2007 Malm Johansonamp Wahlberg 2013) although trichopterans and lepidopterans branched from a commonancestor (Holzenthal et al 2007) This study clarified the defensive function in the orderLepidoptera strengthening the hypothesis that case-bearing behaviour has repeatedlyevolved for anti-predator defence in insects

I observed attackndashdefence behaviour in 30 pairs of the predator C maximoviczi and theprey E minuscula (Table 1) However I excluded the data for the second prey from theFisherrsquos exact tests because the same individuals of C maximoviczi and E minuscula wereused in different experiments (Table 1) Such data could be analysed using a generalisedlinear mixed model (GLMM) with a binomial error distribution and a logit link withdefensive success or failure (0 or 1) by bagworms as a binary response bag treatmentsas fixed factors and carabid individuals as a random effect However all bagwormssuccessfully defended themselves in at least one treatment group (Table 1) therebyextending parameters to infinity when all values in a category were 0 or 1 (cf Sugiura ampYamazaki 2014) Therefore the GLMM could not be conducted in this study Althoughthe sample size for Fisherrsquos exact tests was too small (n= 5treatment) the combined datashowed robust results ie all control group bagworms (n= 15) could successfully defendagainst carabid attacks and all bag-removed larvae (n= 10) failed to defend against theattacks (Fig 3)

No carabid species have been observed preying on bagworm larvae under fieldconditions However I showed that bagworms could perfectly defend against carabidattacks (Table 1 and Fig 3) Such perfect defence by bagworms suggests very few chances toobserve carabid predation on bagworms under field conditions Other natural enemies areknown to impact bagworms (Ellis et al 2005 Rhainds Davis amp Price 2009) For examplebirds have been considered to regulate bagworm populations (Horn amp Sheppard 1979)However birds may not prefer bagworms over non-bagged caterpillars because of theincreased handling cost (ie time taken to remove bags cf Moore amp Hanks 2000)Furthermore the large bags of bagworms have been observed to prevent parasitoid


oviposition (Cronin amp Gill 1989) However more diverse parasitoid species and higherparasitism rates have been reported for case-bearing caterpillars than bare caterpillars(Hawkins 1994) In fact a relatively large number of parasitoid species is known toparasitise the bagworm E minuscula (Nishida 1983) This may be related to the lsquolsquorefugiarsquorsquohypothesis ie caterpillars that are unlikely to be eaten by predators can provide enemy-freespace for parasitoids (Gentry amp Dyer 2002 Stireman amp Singer 2003) Therefore indirectinteractions among predators and parasitoids via shared prey may alter selection pressureson bag evolution in bagworms Studies have used predators from various groups includingants bugs and wasps to test the effectiveness of caterpillar defences against natural enemies(Dyer 1995 Dyer 1997 Murphy et al 2009) however I used a single predator species inthis study Many interaction factors such as attack size strategy and natural history of thepredators may cause the variation in defensive effectiveness in caterpillars Consequentlybagworm defences against predators other than carabid beetles should be tested to clarifythe selective agents leading to the evolution of portable bags

Bagworm bags may have other functions (Rhainds et al 2009) For example bagscan provide microclimate conditions that protect immature bagworm from desiccationor that accelerate development (Barbosa Waldvogel amp Breisch 1983 Smith amp Barrows1991 Rivers Antonelli amp Yoder 2002 Rhainds et al 2009) In addition constructing bagscan magnify their relative size to arthropod predators eg bags were ca 18 times thelength of larvae in E minuscula (Fig 1B) The size magnification by bag construction canprovide protection through increased effectiveness of physical or behavioural defencesagainst arthropod predators because predation by arthropods is generally negativelysize-dependent (Remmel Davison amp Tammaru 2011 Greeney Dyer amp Smilanich 2012)However one study indicated that C maximoviczi eventually attacked various sizes andspecies of lepidopteran larvae (body weight 333ndash5667 mg body length 126ndash346 mmSugiura amp Yamazaki 2014) Furthermore I observed C maximoviczi adults attackinglarge hawk moth larvae under laboratory conditions (body weight 72886ndash168669 mgbody length 843ndash1127 mm) although they did not successfully prey on the large larvae(Sugiura unpublished data) Therefore the different predation rate by C maximovicziadults between control and bag-removed larvae (Table 1 and Fig 3) was not caused bythe size difference between control and bag-removed larvae but by the presenceabsenceof bags Furthermore the cryptic appearance can also serve as camouflage (Rhainds etal 2009) and although this study did not test the importance of cryptic appearance forbagworms C maximoviczi adults were frequently observed to attack and bite motionlessbags of E minuscula larvae This suggests that carabids can use scent as well as appearanceto locate prey Therefore the cryptic appearance of bagworms was unlikely to influencemy results Taken together bagworm bags may have various types of functions that are notmutually exclusive Portable cases that have more than one function may be selected morefrequently and evolve more rapidly than those with a single function

ACKNOWLEDGEMENTSI thank the editor and two reviewers for critical comments on my manuscript


ADDITIONAL INFORMATION AND DECLARATIONS

FundingThis researchwas partly supported by aGrant-in-Aid for Scientific Research (no 25292034)The funders had no role in study design data collection and analysis decision to publishor preparation of the manuscript

Grant DisclosuresThe following grant information was disclosed by the authorGrant-in-Aid for Scientific Research 25292034

Competing InterestsThe author declares there are no competing interests

Author Contributionsbull Shinji Sugiura conceived and designed the experiments performed the experimentsanalyzed the data contributed reagentsmaterialsanalysis tools wrote the paperprepared figures andor tables reviewed drafts of the paper

EthicsThe following information was supplied relating to ethical approvals (ie approving bodyand any reference numbers)

The experiments were undertaken according to the Kobe University AnimalExperimentation Regulations

Data AvailabilityThe following information was supplied regarding data availability

Raw data is shown in Table 1

Supplemental InformationSupplemental information for this article can be found online at httpdxdoiorg107717peerj1686supplemental-information

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the bagworm Thyridopteryx ephemeraeformis (Lepidoptera Psychidae) CanadianEntomologist 115855ndash858 DOI 104039Ent115855-7

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Brown CG Funk DJ 2010 Antipredatory properties of an animal architecture howcomplex faecal cases thwart arthropod attack Animal Behaviour 79127ndash136DOI 101016janbehav200910010


Bruschi S 2013 Calosoma of the world Bologna Natura Edizioni ScientificheChaboo CS Brown CG Funk DJ 2008 Faecal case architecture in the gibbosus

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Dyer LA 1995 Tasty generalists and nasty specialists Antipredator mechanisms intropical lepidopteran larvae Ecology 761483ndash1496 DOI 1023071938150

Dyer LA 1997 Effectiveness of caterpillar defense against three species of invertebratepredators Journal of Research on the Lepidoptera 3448ndash68

EdmundsM 1974Defense in animals Harlow LongmanEisner T 2003 For love of insects Cambridge The Belknap Press of the Harvard

University PressEisner T Esiner M Siegler M 2005 Secret weapons defences of insects spiders scorpions

and other many-legged creatures Cambridge The Belknap Press of the HarvardUniversity Press

Ellis JAWalter AD Tooker JF Ginzel MD Reagel PF Lacey ES Bennett AB GrossmanEM Hanks LM 2005 Conservation biological control in urban landscapesmanipulating parasitoids of bagworm (Lepidoptera Psychidae) with flowering forbsBiological Control 3499ndash107 DOI 101016jbiocontrol200503020

Emlen DJ 2014 Animal weapons the evolution of battle New York HoltFerry EE Hopkins GR Stokes AN Mohammadi S Brodie ED Gall BG 2013 Do all

portable cases constructed by caddisfly larvae function in defense Journal of InsectScience 13Article 5 DOI 1016730310130501

Forsythe TG 1982 Feeding mechanisms of certain ground beetles (Coleoptera Cara-bidae) The Coleopteristsrsquo Bulletin 3626ndash73

Gentry GL Dyer LA 2002 On the conditional nature of neotropical caterpillar defensesagainst their natural enemies Ecology 833108ndash3119DOI 1018900012-9658(2002)083[3108OTCNON]20CO2

Greeney HF Dyer LA Smilanich AM 2012 Feeding by lepidopteran larvae is dangerousa review of caterpillarsrsquo chemical physiological morphological and behavioraldefenses against natural enemies Invertebrate Survival Journal 97ndash34

Hawkins BA 1994 Pattern and process in host-parasitoid interactions CambridgeCambridge University Press

Holzenthal RW Blahnik RJ Prather AL Kjer KM 2007 Order Trichoptera Kirby 1813(Insecta) caddisflies Zootaxa 1668639ndash698

Horn DJ Sheppard RF 1979 Sex ratio pupal parasitism and predation in two decliningpopulations of the bagworm Thyridopteryx ephemeraeformis (Haworth) (Lepi-doptera Psychidae) Ecological Entomology 4259ndash265DOI 101111j1365-23111979tb00583x


Ikeda K 1988 Predation on the bagworms Eumeta minuscula and Eumeta japonica by theJapanese White-eye Zosterops japonicus in winter Forest Pest 3728ndash31 (in Japanese)

Kamata N Igarashi Y 1995 An example of numerical response of the carabid beetleCalosoma maximowicziMorawitz (Col Carabidae) to the beech caterpillarQuadricalcarifera punctatella (Motschulsky) (Lep Notodontidae) Journal of AppliedEntomology 119139ndash142 DOI 101111j1439-04181995tb01259x

Kaufmann T 1968 Observations on the biology and behavior of the evergreen bagwormmoth Thyridopteryx ephemeraeformis (Lepidoptera Psychidae) Annals of theEntomological Society of America 6138ndash44 DOI 101093aesa61138

Kobayashi F Taketani A eds 1993 Forest insects Tokyo Yokendo (in Japanese)Malm T Johanson KAWahlberg N 2013 The evolutionary history of Trichoptera

(Insecta) a case of successful adaptation to life in freshwater Systematic Entomology38459ndash473 DOI 101111syen12016

Moore RG Hanks LM 2000 Avian predation of the evergreen bagworm (LepidopteraPsychidae) Proceedings of the Entomological Society of Washington 102350ndash407

Murphy SM Leahy SMWilliams LS Lill JT 2009 Stinging spines protect slugcaterpillars (Limacodidae) from multiple generalist predators Behavioral Ecology21253ndash160 DOI 101093behecoarp166

Nishida E 1983 Biologies and parasite complexes of two bagworms Eumeta japonicaand Eumeta minuscula (Lepidoptera Psychidae) in Japan Kontyucirc 51394ndash411

Otto C Svensson BS 1980 The significance of case material selection for the survival ofcaddis larvae Journal of Animal Ecology 49855ndash865 DOI 1023074231

Pierre EM Idris AH 2013 Studies on the predatory activities of Oecophylla smaragdina(Hymenoptera Formicidae) on Pteroma pendula (Lepidoptera Psychidae) in oilpalm plantations in Teluk Intan Perak (Malaysia) Asian Myrmecology 5163ndash176

RDevelopment Core Team 2012 R a language and environment for statistical comput-ing Vienna R Foundation for Statistical Computing

Remmel T Davison J Tammaru T 2011 Quantifying predation on folivorous insectlarvae the perspective of life-history evolution Biological Journal of the LinneanSociety 1041ndash18 DOI 101111j1095-8312201101721x

Rhainds M Davis DR Price PW 2009 Bionomics of bagworms (Lepidoptera Psychi-dae) Annual Review of Entomology 54209ndash226DOI 101146annurevento54110807090448

Rivers DB Antonelli AL Yoder JA 2002 Bags of the bagworm Thyridopteryxephemeraeformis (Lepidoptera Psychidae) protect diapausing eggs from water lossand chilling injury Annals of the Entomological Society of America 95481ndash486DOI 1016030013-8746(2002)095[0481BOTBTE]20CO2

Root RB Messina FJ 1983 Defensive apdaptation and natural enemies of a case-bearing beetle Exema canadensis (Coleoptera Chrysomelidae) Psyche 9067ndash80DOI 101155198347471

Shima H 1999Host-parasite catalog of Japanese Tachinidae (Diptera)Makunagi ActaDipterologica Suppl 11ndash108


SmithMP Barrows EM 1991 Effects of larval case size and host plant species oncase internal temperature in the bagworm Thyridopteryx ephemeraeformis (Ha-worth)(Lepidoptera Psychidae) Proceedings of the Entomological Society of Wash-ington 93834ndash838

Stehr FW 1987 Immature insects vol 1 Dubuque KendallHuntStireman III JO Singer MS 2003 Determinants of parasitoid-host associations

insights from a natural tachinid-lepidopteran community Ecology 84296ndash310DOI 1018900012-9658(2003)084[0296DOPHAI]20CO2

SugimotoM 2009a A comparative study of larval cases of Japanese Psychidae (Lepi-doptera) (1) Japanese Journal of Entomology (New Series) 121ndash15 (in Japanese withEnglish summary)

SugimotoM 2009b A comparative study of larval cases of Japanese Psychidae (Lepi-doptera) (2) Japanese Journal of Entomology (New Series) 1217ndash29 (in Japanese withEnglish summary)

Sugiura S Yamazaki K 2014 Caterpillar hair as a physical barrier against invertebratepredators Behavioral Ecology 25975ndash983 DOI 101093behecoaru080

Weseloh RM 1985 Predation by Calosoma sycophanta L (Coleoptera Carabidae) evi-dence for a large impact on gypsy moth Lymantria dispar L (Lepidoptera Lymantri-idae) pupae Canadian Entomologist 1171117ndash1126 DOI 104039Ent1171117-9


(eg caddisfly larvae of the suborder Integripalpia Holzenthal et al 2007) Coleoptera(eg leaf beetle larvae of the subfamilies Clytrinae Cryptocephalinae Chlamisinae andLamprosomatinae Brown amp Funk 2005 Chaboo Brown amp Funk 2008) and Lepidoptera(eg moth larvae of the superfamilies Incurvarioidea and Tineoidea Stehr 1987) Physicaldefence against predators using portable cases has been tested experimentally in Trichoptera(Otto amp Svensson 1980 Ferry et al 2013) and Coleoptera (Root amp Messina 1983 Brown ampFunk 2010) but not in Lepidoptera

The bagworm family Psychidae (Lepidoptera Tineoidea) includes ca 1000 species andall of their larvae construct portable cases (Rhainds Davis amp Price 2009) The materialsused for constructing bags differ among bagworm species eg treeherbgrass leaveslichens twigs petioles bark fragments wood debris and sand particles (Sugimoto 2009aSugimoto 2009b) The portable bags are generally believed to play an important role asportable armour against natural enemies (Rhainds Davis amp Price 2009) For example bagshave been reported to function as a physical barrier against parasitoid attack the ovipositorof an ichneumonid parasitoid was too short to reach pupae of the bagworm Thyridopteryxephemeraeformis (Haworth) inside the larger bags and the parasitism rate was inverselycorrelated with bag size (Cronin amp Gill 1989) However bagworm larvae and pupae insidebags are generally known to suffer heavier parasitism by more diverse parasitoids thanare other external-feeding caterpillars (Hawkins 1994) suggesting that bagworm bagsmay not be effective armour against parasitoids Rather predators such as birds andpredacious arthropods may impose a selective pressure on the evolution or maintenanceof bags Although the impacts of predators have been reported in some bagworm species(Rhainds Davis amp Price 2009 Pierre amp Idris 2013) no experimental studies have testedthe importance of bags and materials used for bags as defensive armour against predatorsClarifying the defensive function of bags would contribute to further understanding ofhow portable armour has evolved in animals

To test whether portable bags can protect bagworms from predator attacks Calosomaadults (Coleoptera Carabidae) were observed attacking larvae of a bagworm species underlaboratory conditions Adults of the carabid genus Calosoma hunt lepidopteran larvae andpupae (Forsythe 1982Weseloh 1985 Bruschi 2013) providing a good model predator forinvestigating the defensive behaviour of lepidopteran larvae (Sugiura amp Yamazaki 2014)In this study I first investigated the defensive success or failure of bagworm larvae againstcarabid attacks Second I tested whether bag-removed larvae could defend themselvesagainst carabids in order to clarify the importance of bags Furthermore I investigated theeffects of bag replacement (with a different type of bag) on the defensive success of bagwormlarvae against carabid attacks to elucidate the importance of materials for constructingbags

MATERIALS AND METHODSStudy speciesTo clarify the defensive function of portable bags I used the bagworm species Eumetaminuscula Butler (Psychidae) and the potential predator Calosoma maximoviczi Morawitz(Carabidae)
















Nob Sex Weight(mg)

No b Bagtreatmentc

Weight(mg)d


Nob Bagtreatmentc

Weight(mg)d






Sugiura(2016)PeerJD

OI107717peerj1686

614





























































































Nob Sex Weight(mg)

No b Bagtreatmentc

Weight(mg)d


Nob Bagtreatmentc

Weight(mg)d






Sugiura(2016)PeerJD

OI107717peerj1686

614















































































bagworm bags as portable armour against invertebrate predators · bags are generally known to...

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