Gut anatomy of the worker caste of Neotropical genera Cylindrotermes Holmgren and Hoplotermes Light (Infraorder Isoptera, Termitidae)

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REVISTA

BRASILEIRA

DE

Entomologia

AJournalonInsectDiversityandEvolution

w w w . r b e n t o m o l o g i a . c o m

Systematics,

Morphology

and

Biogeography

Gut

anatomy

of

the

worker

caste

of

Neotropical

genera

Cylindrotermes

Holmgren

and

Hoplotermes

Light

(Infraorder

Isoptera,

Termitidae)

Mauricio

Martins

Rocha

∗

,

Carolina

Cuezzo

UniversidadedeSãoPaulo,MuseudeZoologia,SãoPaulo,SP,Brazil

a

r

t

i

c

l

e

i

n

f

o

Articlehistory: Received10April2019 Accepted14June2019 Availableonline22June2019 AssociateEditor:DouglasZeppelini Keywords:

Entericvalve

Firstproctodealsegment Gizzard

Termite Workercaste

a

b

s

t

r

a

c

t

Studiesoverthelastsixdecadeshaveshownthatthegutanatomyofthetermiteworkercasteprovides avaluablesetoftaxonomiccharacters.However,thegutanatomyofmostAmericantermitinetaxais stilllittleknown.ThisstudyinvestigatedtheanatomyoftheworkergutoftheNeotropicaltermitine generaCylindrotermesHolmgrenandHoplotermesLight.Weprovidedescriptionsandillustrationsofthe gutinsituandthecuticularornamentationofthegizzard,entericvalve,andﬁrstproctodealsegmentfor bothgenera.

Introduction

With63generaand643livingspeciesworldwide(Constantino, 2019),theTermitinaeisaheterogeneousgroupoftermites, with-outdiagnosticcharacterssupportingitasacladeamongTermitidae subfamilies(Bourguignon et al.,2017). The subfamilyis repre-sented in theNeotropical region by 20 genera, occurringfrom thesouthernUnitedStates(GnathamitermesLight,1932;

Hoploter-mesLight,1933)tosouthernArgentina(OnkotermesConstantino,

2002). Three of these genera are cosmopolitan (Amitermes

Silvestri, 1901, Microcerotermes Silvestri, 1901, and Termes

Linnaeus,1758).

The South and Central American genus Cylindrotermes

Holmgren, 1906 was proposed based on specimens collected

inBolivia,duringtheNordenskiöldexpedition(1904–1905).One centurylater,Rochaand Cancello(2007)revisedthegenusand described three new species, based on the external morphol-ogy of the soldier and worker castes from Brazilian samples, and redescribed four of the ﬁve previously described species (exceptthetype speciesCylindrotermes nordenskioldi Holmgren, 1906).Noirot(2001,Fig.12d)illustratedanunidentiﬁedspecies of Cylindrotermes, with brief comments on internal structures. With a total of eight nominal species now recognized within Cylindrotermes,thegeographicaldistributionofthegenusranges

∗ Correspondingauthor.

E-mail:mmrocha.cupim@gmail.com(M.M.Rocha).

fromtherainforestof southernCentral America (CostaRica)to thesavannas(Cerrado)ofSouthAmerica.Cylindrotermesspecies formsmallcolonies,frequentlyfoundinsidedrydeadwood(small trunksanddrysticksontheground).Therearereportsofdamage torootsandstemsofsugarcanecausedbymembersofthegenus

(Mirandaetal.,2004).

Hoplotermesisanuncommonmonotypictermitegenus,with Hoplotermes amplus Light, 1933 as the type-species, described fromtheexternalmorphology ofthesoldierand worker castes of specimens from western Mexico; the imago caste remains unknown. The gut anatomy of Hoplotermes has never been described.ThegenushasbeenrecordedinCostaRica,Guatemala and Nicaragua (Krishna et al., 2013). Colonies of this species are found in wood, under rocks, or cow dung; they build earthen chambers and passageways in, on and near attacked

wood(Light,1933).

Characters of the worker caste digestive tube have come to be widely recognized as key resources for taxonomy over the last six decades (Noirot and Kovoor, 1958; Sands, 1992,

1998; Noirot,2001), and theirdescriptionhasbeenmandatory

for taxonomic studiesin thelast two decades.Although many generaare reasonablywell representedinmuseums,the inter-nal morphology of many species is still unknown. To help to ﬁlltheselacunae,weprovidedetaileddescriptionsand illustra-tionsof theworker gutof seven species of Cylindrotermes and H. amplus, including the gut coiling in situ and the cuticular ornamentationofthegizzard,entericvalve,andﬁrstproctodeal segment.

https://doi.org/10.1016/j.rbe.2019.06.002

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pectinatescales),(b)entericvalve,withoneofthecushionsoutlined(largearrow: proximalpad,smallarrow:distalportion).

Fig.2.Cylindrotermescaata:(a)detailofgizzardarmature(smallarrow:crop pecti-natescales),(b)entericvalve,withoneofthecushionsoutlined(largearrow: proximalpad,smallarrow:distalportion).

Materialandmethods

Materialexamined

WestudiedspecimensdepositedintheMuseudeZoologiada Universidadede São Paulo(MZUSP), São Paulo,Brazil.We dis-sectedandillustratedtheworkergutcoiling,gizzard,andenteric valveofHoplotermesamplus(MZUSP-15464,Guatemala,Jutiapa); Cylindrotermes brevipilosus Snyder, 1926(MZUSP-24534: Brazil, MatoGrossodoSulState,CostaRicamunicipality);Cylindrotermes

WeadoptedtheterminologyofNoirot(2001)forthegutcoiling, gizzard,andentericvalvecuticularornaments,andofRochaand

Constantini(2015)fortheﬁrstproctodealcuticularornaments.

Linedrawingscharacterizingthegutinsituandtheﬁrst proc-todealsegmentwerepreparedwithadrawingtubecoupledtoa stereomicroscope.Thegizzard,entericvalve,andﬁrstproctodeal segmentweremountedinglycerinonaslideandphotographed withanopticalmicroscope.Particularly,theentericvalveof Cylin-drotermeshasasmallsectionandatightlyattachedmusculature, whichischallengingtomountsatisfactorily,especiallysincethe numberofgood-qualityspecimensfordissectionwaslimited.

Results

Afterdetailedexamination,weobserveddifferentgut-coiling patternsinsituandaparticularcuticularornamentationforeach genera,CylindrotermesandHoplotermes,asfollows.

Cylindrotermes

Worker digestive tube (Figs. 1–8): Crop slightly asym-metrical, not voluminous, covered with ﬁnely pectinate scales

(Figs.1a,2a,3a,4a,5a,6aand7a,arrows).Gizzardwitha

com-plete armature arranged in a columnar belt with24 folds (six first-order,sixsecond-order,and12 third-order)andapulvillar beltwith12folds(sixfirst-order,sixsecond-order);Lengthratio betweenbeltsaboutone;pulvillioffirstorderconspicuous,with tinypectinate scales,and pulvilli ofsecond ordervery reduced

(Figs. 1a, 2a, 3a, 4a, 5a, 6a and 7a).Stomodeal valve inserted

apically in thetubularmesenteron. Mesenteric tongueexternal

Fig.3.Cylindrotermescapixaba:(a)detailofgizzardarmature(smallarrow:croppectinatescales),(b)entericvalve,withoneofthecushionsoutlined(largearrow:proximal pad,smallarrow:distalportion).

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Fig.4. Cylindrotermesﬂangiatus:(a)detailofgizzardarmature(smallarrow:croppectinatescales),(b)entericvalve,withoneofthecushionsoutlined(largearrow:proximal pad,smallarrow:distalportion).

Fig.5.Cylindrotermesmacrognathus:(a)detailofgizzardarmature(smallarrow:croppectinatescales),(b)entericvalve,withoneofthecushionsoutlined(largearrow: proximalpad,smallarrow:distalportion).

Fig.6.Cylindrotermesparvignathus:(a)detailofgizzardarmature(smallarrow:croppectinatescales),(b)entericvalve,withoneofthecushionsoutlined(largearrow: proximalpad,smallarrow:distalportion).

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pectinatescales),(b)entericvalve,withoneofthecushionsoutlined(largearrow: proximalpad,smallarrow:distalportion).

tothemesenteric arch,tapering broadly,and connected atthe mesenterononlybyaﬁliformmesenterictissue(Fig.8a,arrow). Two pairs of Malpighian tubules attached at the mesenteron-proctodeumjunction,withdistinctinsertionsonoppositesidesof thesegment(Fig.8e,arrow).Firstproctodealsegment(P1) tubu-larandelongated,formingamarkedloopbellowtherectum(P5) (Fig.8a–d),distallyattachedtothepaunch(P3)atthedorsal/right sideoftheabdomen(Fig.8b,arrow)anddeeply insertedatthe P3,surroundedbyathickbandofmuscle.P1withoutcuticular ornamentation.Enteric-valve armature (P2) weakly sclerotized, composedbythreenearlyequalcushions,eachcushionwithtwo distinctspine-covered areas,the padclosest toP1,withrobust spines(Figs.1b,2b,3b,4b,5b,6b,and7b,largearrows);andan elongatedportion,coveredwithsmallscatteredspines,closestto

reliablyidentiﬁed specimensforCylindrotermesnordenskioldi, to beconsideredherein,weexpectasimilarpattern.Cylindrotermes shareswiththeEthiopiangenusCephalotermesSilvestri,1912and thesoldierlessOrientalgenusProtohamitermesHolmgren,1912,the tubularP1formingaventralloop.InCephalotermestheP1formsa shorterloopthaninCylindrotermes,themesenterictonguelacksa ﬁliformconnection,andtheentericvalvehassixunequal

cush-ions(Noirot,2001, Figure12 D,E;Sands, 1998,Figs. 669–681),

Protohamitermesworkermandibles(Ahmad,1976,Fig.8A)differ considerablyfromthoseofCylindrotermesandCephalotermes,and theentericvalvehassixcushions(Ahmad,1976,Fig.10A).Another possiblyrelatedgeneraisOrientotermesAhmad,1976,asoldierless Orientaltermite(Bourguignonetal.,2017),butsincelittle infor-mationontheworkergutanatomyisavailable,itisnotpossible tomakeacompletecomparison.Themandiblesclearlydifferfrom thoseofCylindrotermes(Ahmad,1976,Fig.8B)andtheentericvalve hassixcushions(Ahmad,1976,Fig.11A).

Fig.8.Cylindrotermesparvignathusworkergutinsitu:(a)dorsal,(b)right(arrow:entericvalveinsertion),(c)ventral,and(d)leftviews,and(e)detailofmixedsegmentand Malpighiantubulesinsertion(arrow).Grayindicatesmesenterictissue;c:crop;M:mesenteron;MT:mesenterictongue,i:isthmus,P1:ﬁrstproctodealsegment(ileum); P3:thirdproctodealsegment(paunch);P4:fourthproctodealsegment(colon);P5:ﬁfthproctodealsegment(rectum).

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Fig.9.Hoplotermesamplusworker:(a)detailofgizzardarmature(smallarrow:croppectinatescales),(b)entericvalve(largearrows:cushions).

Fig.10.Hoplotermesamplusworkergutinsitu:(a)detailofstomodealvalveinsertion,(b)dorsal,(c)right,(d)ventraland(e)leftviews;(f)detailofenteric-valveinsertion indorsalview(P4andP5removed,arrow:enteric-valveinsertion).Grayareaindicatesmesenterictissue;c:crop;M:mesenteron;MT:mesenterictongue,i:isthmus,P1: ﬁrstproctodealsegment(ileum);P3:thirdproctodealsegment(paunch);P4:fourthproctodealsegment(colon);P5:ﬁfthproctodealsegment(rectum).

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Fig.11.HoplotermesamplusP1ornaments:(a)schematicdrawingofP1showingarrangementofspines,withbindicatingrobustspinesinproximalregion,andindicating sparseshortspineslaterallyanddistally;(b)detailofrobustproximalspines;(c)detailofsparseshortspines(MT,mesenterictonguespace).

Ontheotherhand,allNeotropicalapicotermitinegeneraand someOrientalgenera(e.g.,EuhamitermesHolmgren,1912)havea similarloopedP1(Noirot,2001;Fig.10H,L).Theydifferfrom Cylin-drotermesinhavingtheleftmandiblewithaconspicuousnotch beforethethirdmarginaltooth,andthemesenterictongue,when present,internaltothemesentericarch.

Hoplotermes

Workerdigestivetube(Figs.9–11):Cropslightly asymmetri-cal,moderateinsize,coveredwithfinelypectinatescales(Fig.9a, arrow).Gizzardwithacompletearmaturearrangedina colum-narbelt with24folds(six first-order,six second-order,and 12 third-order) and a pulvillar belt with 12 folds (six first-order, six second-order); Length ratio between belts about one; pul-villioffirstorderornamentedwithrecognizablepectinatescales, pulvilliofsecondorderinconspicuous(Fig.9a).Stomodealvalve insertionslightlydisplacedtotheinnerregionofthemesenteric arch(Fig.10a).Mesenterontubular,withasinglewell-developed mesenterictongueoflengthnearlyequaltohalfthatofthe mesen-tericarch,endingjustatthebeginningoftheP1dilatedportion (Fig.10d).ItwasnotpossibletoexaminetheMalpighiantubules, sincewehadfewspecimensfordissection.P1stronglydilated,its sizenearlyequaltotheP3(Fig.10c,dandf),andattachedtoit byanarrowconnectionatthedistalregionoftheabdomen,below theP5(Fig.10f,arrow);cuticularornamentationofP1withrobust spineslocatedaroundtheborder ofthemesenterictongueand justdistaltoit(Fig.11a,b),andwithsparseshortspineslaterally anddistally(Fig.11a,c).P2withasingleringofthreeelongated cushions(Fig.9b,arrows),weaklymarked,radiallyarranged,and coveredsparselywithsmallspines.P3welldeveloped,protruded throughthemesentericarch.Isthmusmarked(Fig.10b),P4witha shortU-turn(Fig.10c).

Comparisonsamongrecognizedtermitinegenera: Hoploter-mes, some species of Amitermes (Sands, 1998, Figs. 609–628), GenuotermesEmerson,1950,andmostsyntermitinegenera(see Rochaetal.,2017)haveastronglyinﬂatedgloboseorfusiformP1 andsimilarcuticularornamentationoftheP2,withspines orga-nizedindistinctpatterns(Rochaetal.,2017;Engeletal.,2009). Hoplotermesandsomesyntermitinegenerahaveasimilar distri-butionof spines in theP1 cuticular ornamentation (Rocha and

Constantini,2015),withthemostrobustspinesconcentratedin

theproximalportion. NootherAmericantermitinegenerahave thiscombinationofmorphologicalfeatures.

Discussion

NoirotandKovoor(1958)proposedtwogroupswithin

Termiti-nae,deﬁnedbytheanatomyofthegut:theThoracotermesandthe Termes,basedonlyonAfricantaxa.Later,Noirot(2001),withalarge number of specimens fromdifferentregions, distinguishedﬁve groups:thepantropicalTermesgroup,theOrientalPericapritermes group,thepantropicalAmitermesgroup,theEthiopian Cubiter-mesgroup[theThoracotermesgroupofNoirotandKovoor(1958)], and the African Foraminitermes group (hypothetical new sub-family). Engeletal. (2009) formallyraisedthelast two groups to subfamily rank, the Cubitermitinae and Foraminitermitinae. These subfamilies were supported as natural clades in studies

by Inward et al. (2007) and Bourguignon et al. (2017). Rocha

etal.(2019) recognizedin theAmericanregion,theAmitermes

group, comprising the genera Amitermes, Gnathamitermes, and Hoplotermes;theCavitermes-Termesgroup,withtheCavitermes subgroup, comprising Cavitermes Emerson, 1925,

Cornicapriter-mes Emerson, 1950, Dihoplotermes Araujo, 1961, Divinotermes

Carrijo and Cancello, 2011, Palmitermes Hellemans and Roisin,

2017,andSpinitermesWasmann,1897,andtheTermessubgroup, comprisingInquilinitermesMathews,1977,andTermes;the Neo-capritermes group,comprising Neocapritermes Holmgren, 1912, PlanicapritermesEmerson,1949(inSnyder,1949),and

Crepititer-mesEmerson,1925;andtheOrthognathotermesgroup,comprising

Dentispicotermes Emerson, 1949 (in Snyder, 1949) and Orthog-nathotermes Holmgren, 1910; while separating Cylindrotermes, Genuotermes, Microcerotermes, and Onkotermes as truly distinct genera.

Thepresentstudyexaminedindetailtheinternalmorphologyof theworkercasteofCylindrotermesandHoplotermes, complement-ingtheiroriginaldescriptions.Basedonthisnewinformation,we considerthatCylindrotermescannotbeassignedtotheAmitermes group (sensu Noirot, 2001). Evidence from the gut morphol-ogysupportsCylindrotermesasarelativeoftheEthiopiangenus Cephalotermes,ahypothesispreviouslyadvancedbyInwardetal.

(2007)andBourguignonetal.(2017),intheirphylogenetic

recon-structions.

Hoplotermes has not yet been included in any evolutionary scheme, but based on our morphological observations, it may berelatedtotheAmitermes–Drepanotermeslineage.Toovercome thislimitationandtoreachﬁrmconclusionsaboutthekinshipof Hoplotermes,westronglyrecommendedthatthistermitinegenus beincludedinfuturephylogeneticanalyses.Authorcontributions

M.M.R.conceivedtheresearchandeditedtheﬁgures;M.M.R. andC.C.collectedandanalyzeddataandwrotethepaper.

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Conﬂictsofinterest

Theauthorsdeclarenoconﬂictsofinterest.

Acknowledgements

WearegratefultoR.H.Scheffrahn(UniversityofFlorida) for donatingmaterialofH.amplus.Wereceivedﬁnancialsupportfrom theNationalCouncilforScientiﬁcandTechnologicalDevelopment, Brazil(CNPq)throughgrantPROTAX-001/2015toM.M.Rocha,and fromtheSãoPauloResearchFoundation,Brazil(FAPESP)through grant2013/05610-1toC.Cuezzo.

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