a new land planarian species of geoplana (platyhelminthes,...

A new land planarian species of Geoplana (Platyhelminthes,Tricladida, Geoplanidae) from the Valdivian temperate rainforestof southern Chile

Jos� Horacio Grau*, 1 and Fernando Carbayo2

1 Museum f�r Naturkunde Berlin, Leibniz-Institut f�r Evolutions- und Biodiversit�tsforschung an der Humboldt-Universit�t zu Berlin,Invalidenstraße, 43, 10115 Berlin, Germany

2 Escola de Artes, Ci�ncias e Humanidades, Universidade de S�o Paulo. S�o Paulo, Brazil

Introduction

Land planarians are a group of carnivorous free-livingplatyhelminths (Jennings 1971) with a worldwide distri-bution, to the exclusion of the polar region (Winsoret al. 1998). They are predators in soil ecosystems,where they inhabit humid forest floors and can reachhigh levels of diversity in the tropics (Winsor et al.1998). The Neotropical land planarian fauna is com-posed of representatives belonging to three subfamilies,Rhynchodeminae, Microplaninae and Geoplaninae; thelatter being the largest and exclusive to the Neotropics(Ogren & Kawakatsu 1998). As external features arefew in number in land planarians, and some of these,such as the color pattern of the body might show in-traspecific variation, internal characteristics are themost important for unequivocal identification, and forproviding systematic and phylogenetic data. Geoplani-nid species, whose internal morphology presently re-mains unknown, are placed in the genus Pseudogeopla-na Ogren & Kawakatsu, 1990; a taxon specificallyerected to accomodate species inquirendae and nominadubia (Ogren & Kawakatsu 1990).

Geoplana Stimpson, 1857 is the richest genus withinGeoplaninae (ca. 280 species) and is found widelythroughout the Neotropical region. Twenty-five nominalspecies are known from Chile, five of them belongingto Geoplana (see Grau & Carbayo 2010). Here we de-scribe a new Chilean Geoplana species from the Valdi-vian temperate rainforest.

Material and methods

A single specimen was collected in January 2007 in the ArboretumPark of the Universidad Austral de Chile, in the campus Island Teja(39�480 S; 73�150 W) at 5 meters above sea level, Valdivia, Los Riosregion, southern Chile. The worm was collected by hand, and thendirectly put into a plastic tube containing 90 % ethanol for fixationand preservation. Fragments of the anterior region, pre-pharyngeal re-gion, pharynx and copulatory apparatus were sectioned and dehy-drated in ascending ethanol series, and subsequently embedded inParaplast. Sagittal, horizontal and transverse serial sections (8 mmthick) of different body regions were stained with Mallory-Cason tri-chrome and Masson stains (Winsor 1998).

The ratio of the height of the cutaneous musculature to the heightof the body (cutaneous muscular index, CMI or mc:h) was calculatedafter Froehlich (1955). Illustrations were made using a microscope

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Zoosyst. Evol. 87 (2) 2011, 327–334 / DOI 10.1002/zoos.201100010

Received 5 January 2011Accepted 1 March 2011Published 23 September 2011

Key Words

ContinenticolaGeoplaninaeflatworm

Abstract

A new species, Geoplana valdiviana sp. n. (Platyhelminthes, Tricladida), from southernChile is described. The species is characterized by having the cutaneous musculatureventrally reaching up to the central nervous system, the parenchymal muscle layersgathered in bundles, a penis papilla with the dorsal insertion anterior to the ventralinsertion, shell glands opening only into the common glandular duct, and oviductsgreatly widened in their ascending portion. The two former features are also present inGeoplana chanca and Pasipha ercilla, and might be an evidence of close phylogeneticrelationship between them.

* Corresponding author, e-mail: [email protected]

with a drawing tube attachment. The holotype is deposited in the Mu-seum f�r Naturkunde Berlin, Germany, under the acronym ZMB (forZoologisches Museum Berlin).

Abbreviations used in figurescml, cutaneous muscle layer; cns, central nervous system; co, commonglandular ovovitelline duct; dd, dorsal double diagonal parenchymalmuscle layer; de, dorsal epithelium; di, dorsal insertion of the pharynx;e, eyes; ej, ejaculatory duct; fa, female genital atrium; g, gonopore; gl,glands; i, intestine; m, mouth; od, ovovitelline duct; phm, pharyngealmusculature; php, pharyngeal pouch; pp, penis papilla; pv, prostatic ve-sicle; sb, sub-intestinal transverse parenchymal muscle layer; sbn, sub-cutaneous nervous system; sd, sperm duct; sg, shell glands; sp, supra-in-testinal transverse parenchymal muscle layer; t, testes; va, vagina; ve,ventral epithelium; vi, ventral insertion of the pharynx; vit, vitellaria.

Results

Geoplana valdiviana sp. n.

Material. Holotype, ZMB 11245. Valdivia (Chile), January 2007. F.C�diz, col. Anterior region: transverse sections on 24 slides. Anteriorregion 2: horizontal sections on six slides. Pre-pharyngeal region:transverse sections on four slides. Pharynx: sagittal sections on sixslides. Copulatory apparatus: sagittal sections on ten slides. Remain-der of body in 80 % ethanol.

Diagnosis. Geoplana species with body lanceolate andflat, 24 mm in length. Blackish dorsum provided with across-shaped whitish design on the cephalic region, anda median yellow band on the rest of the back. Eyesdorsal with clear halos. Glandular margin present. CMI,18–19 %. Testes dorsal, underneath supra-intestinaltransverse parenchymatic muscle layer; dorsal insertionof penis papilla anterior to ventral one. Ascending por-tion of ovovitelline ducts greatly widened. Shell glandsopening only into common glandular duct.

Type locality. Valdivian temperate rainforest (39�480 S; 73�150 W) sur-rounding the city of Valdivia in southern Chile. The park where theworm was collected bears some patches of native forest that suffersfrom low human disturbance.

Etymology. The specific name refers to the city of Val-divia, southern Chile, where the specimen was col-lected.

DescriptionExternal morphology. Body of medium size, after fixa-tion 24 mm long, 3.3 mm wide, 0.9 mm thick. Parallel

body margins throughout most of the body length andgradually tapering towards the pointed anterior tip; theposterior one is rounded. Dorsum general appearancegrey-blackish, with a wide yellow median band, andscattered with white eye halos on the lateral region(Fig. 1). The anterior 1.5 mm region is blackish, with athin median white stripe almost at the end of which atransverse stripe (also white) runs perpendicular, thusforming a cross pattern near the anterior tip. A medianyellowish band with scattered small dark pigment dotsruns posteriorly from the median white stripe up toshortly before the posterior end. The yellowish band isoutlined by black stripes that become dark-grey towardsthe body margins. White eye halos spread dorsally oneach side of the yellow median band. Ventrally thebody is whitish, with the anterior tip darker than rest ofthe under-surface. A faint grey median line runs alongthe ventral surface.

Eyes present at the anterior tip arranged in an irregu-lar single marginal row. Posteriorly eyes gradually mul-tiplying and extending dorsally as much as 1/3 of bodywidth from each body side along the entire bodylength. Sensory pits running marginally along the firstthird of the body in a single row. Ciliated creeping soleextending to about 80 % of body width.

Epidermis and its secretions. Dorsal epitheliumthicker than ventral (Figs 2B–C). Granular erythrophilsecretions opening through dorsal body surface andthrough body margins. Amorphous cyanophil glandsopening through dorsal and ventral surfaces. Rhabdito-gen cells distributed dorsally and ventrally, being moreabundant on the dorsal surface. Towards the anterior re-gion of the body granular erythrophil secretions accu-mulate between the intestinal branches and open mainlythrough the margins and the ventral surface (Fig. 2A).Towards the anterior region cyanophil glands also be-come more abundant.

Cutaneous musculature. The cutaneous musculatureis formed by the three typical layers found in Geoplani-nae, i.e., circular, double diagonal and longitudinal. Thelongitudinal layer is the most developed and is packedin thick and dense bundles. The dorsal layer is 50–60 mm thick (Fig. 2B); the ventral, which reaches up tothe ventral nervous plate, is 80–90 mm thick (Figs 2C,

Grau, J. H. & Carbayo, F.: New species of Geoplana (Platyhelminthes, Tricladida) from southern Chile328

Figure 1. Geoplana valdiviana sp. n. Fixed holotype. Photograph of the dorsal side of the body. Anterior end pointig to the left.The posterior end is bent upwards.

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4E). As no ventral submuscular peripheral nerve netwas seen, it is presumed here to be located very closeto the central nerve plate. Towards the anterior and pos-terior ends of the body the muscle layers graduallyweaken until they disappear without specializations.

Parenchymal musculature. Three muscle layers arepresent; dorsal double diagonal (30–40 mm thick),transverse supra-intestinal (30–40 mm thick) and trans-verse sub-intestinal (30–45 mm thick). All three par-enchymal muscle layers constituted by fibers, mostly

Zoosyst. Evol. 87 (2) 2011, 327–334 329

Figure 2. Geoplana valdiviana sp. n. Holotype. Photomicrographs of cross-sectioned pre-pharyngeal region. A. General view;B. Dorsal musculature; C. Ventral musculature.

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gathered into bundles: double diagonal (4–8 fibers perbundle), transverse supra-intestinal (5–12 fibers perbundle) and the transverse sub-intestinal (8–17 fibersper bundle) (Figs 4A–E). Towards the anterior and pos-terior body tips the parenchymal muscle layers gradu-ally weaken without specializations; anteriorly, the

transverse sub-intestinal layer becomes thinner moregradually (Fig. 3A) than the other layers.

Pharynx. Mouth located at the end of the anteriorthird of the pharyngeal pouch. Pharynx bell-shaped,tilted ventrally at an angle of 30�, and occupying mostof pharyngeal pouch. Dorsal insertion of the pharynx


Figure 3. Geoplana valdiviana sp. n. Holotype. A. Photomicrograph of cross section of cephalic region; B. Pharynx, photomicro-graph of the sagittal section of the pharyngeal region; C. Pre-pharynx, reconstruction drawing.


approximately at the level of the mouth (Fig. 3B).Pharyngeal pouch musculature composed of a circularmusculature followed by decussate diagonal fibers.Outer pharynx musculature composed of a layer ofsub-epithelial circular fibers followed by a layer of

longitudinal fibers. Inner pharynx musculature 3–4 times thicker than outer, and composed of a thicksub-epithelial layer of circular fibers followed by alayer of longitudinal fibers. Esophagus very short orabsent.

Zoosyst. Evol. 87 (2) 2011, 327–334 331

Figure 4. Geoplana valdiviana sp. n. Holotype. Photomicrographs of the pre-pharyngeal region. A–B. Frontal section of dorsalcutaneous musculature. Anterior end of the body towards top of the photo; C. Frontal section of ventral cutaneous musculature.Anterior end of the body towards top of the photo; D. Frontal section of dorsal cutaneous musculature. Anterior end of the bodytowards top of the photo; E. Sagittal section of ventral cutaneous musculature.


Male copulatory apparatus. Testes globular, measur-ing about 200�250 mm, and located dorsally betweenthe intestinal branches and the supra-intestinal trans-verse parenchymal muscle layer (Fig. 3C). The testesextend from the ovaric region to shortly before the

pharynx. Sperm ducts placed above the sub-intestinalparenchymal muscle layer and slightly external to theoviducts. The sperm ducts are distally expanded andfull of spermatozoa. Slightly posterior to the pharynxthese ducts curve dorsally and then medially to connect


Figure 5. Geoplana valdiviana sp. n. Holotype. A. Copulatory apparatus, photomicrograph of sagittal section; B. Copulatory appa-ratus, interpretative drawing based on sagittal sections; C. Photomicrograph of a sagittal section showing widened oviduct.


to the end of the anterior third of the prostatic vesicle(Fig. 5B). The sperm ducts are lined with cuboidal andciliated epithelium. The prostatic vesicle is located im-mediately behind the pharyngeal pouch and consists ofa long, horizontal tubular cavity. Its lining epithelium istall columnar and highly vacuolated. Granular erythro-phil secretions surround and penetrate the prostatic ve-sicle. Within the penial bulb the prostatic vesicle con-tinues as a thick ejaculatory duct, 60–70 mm in caliber,opening through the tip of the penis papilla. The ejacu-latory duct is lined with a tall columnar epithelium sur-rounded by a thin reticular muscle layer. The penis pa-pilla is horizontal, cylindrical to conical in shape, andslightly oriented sideways, having the dorsal insertionlocated anterior to the ventral; it occupies most of themale atrium (Figs 5A–B). The penis papilla is linedwith cuboidal nucleated epithelium; it is surrounded bya thin, circular musculature. Numerous necks of glandscontaining amorphous cyanophil secretions openthrough the base of the penis papilla. The penis papillaand the male atrium are lined with cuboidal nucleated,non-ciliated epithelium. The male atrium is slightly lar-ger than the female atrium. The gonopore canal runs ata posteriorly descending angle.

Female reproductive system. The ovaries are globular(130�150 mm). They are located at the end of theanterior third of the body, above the central nerve plateand directly underneath the transverse sub-intestinalparenchymal muscle layer. The oviducts arise from thedorsal side of the ovaries, slightly external to them, andrun posteriorly at the same level. Anterior to the maleatrium they ascend and, dorsally to the gonopore, uniteinto a long and horizontal common glandular ovovitel-line duct. The ascending portions of the oviducts aregreatly widened, up to ten times, due to the fact that itscolumnar epithelium becomes very tall (60–100 mm;Figs 5B–C). The vagina arises dorso-posteriorly fromfemale atrium, and bends dorso-anteriorly to receivecommon glandular ovovitelline duct. The female atriumis a horizontal cavity about three times as long as wide.

The oviduct musculature is composed of cross-orga-nized muscle fibers. The shell glands open only intothe common glandular ovovitelline duct. The female at-rium is lined with a tall columnar and vacuolatedepithelium which is surrounded by interwoven longitu-dinal and circular muscle fibers. The common muscularcoat is formed by longitudinal and oblique muscle fi-bers, and surrounds the male atrium, the female atriumand the common glandular ovovitelline duct.

Discussion

The new species belongs to the subfamily Geoplaninaeas it shows all the main diagnostic characters, i.e. abroad, ciliated creeping sole, well-developed cutaneouslongitudinal musculature gathered into bundles, anddorsal testes. This subfamily contains 17 genera (Ogren& Kawakatsu 1990; Carbayo & Leal-Zanchet 2003;

Carbayo 2010), including a collective genus Pseudogeo-plana Ogren & Kawakatsu, 1990 accommodating spe-cies only known from their external morphology.

The external morphology of G. valdiviana sp. n.shows no similarity to other species allocated to Pseu-dogeoplana because of the combination of body shapeand color pattern; particularly the cross-shaped whitishdesign on the anterior end, and a yellowish midbodyline on the rest of the dorsum.

The new species presents a combination of morpho-logical features that matches the diagnosis of the genusGeoplana, namely the presence of a penis papilla; thedorsal approach of the female canal; the absence ofglandular ridges on the penis papilla; and the absenceof cephalic glandulo-muscular organs and sensory pa-pillae (also absent in the type species Geoplana vaginu-loides (Darwin, 1844), pers. obs.).

Within Geoplana, the new species resembles onlyGeoplana chanca E. M. Froehlich, 1978 in the generalaspect of the copulatory apparatus and in two additionalunique features, i.e., (a) an uncommonly thick andstrong cutaneous musculature in which the longitudinalmuscle fibers are gathered into dense bundles, ventrallyreaching up to the central nervous system; and (b) par-enchymal musculature strongly developed and alsogathered into bundles (as seen by F. Carbayo in thetype material of G. chanca). Geoplana valdiviana sp. n.can be distinguished from G. chanca mainly due to thefact that in the latter the dorsal insertion of the penispapilla is at the level of the ventral insertion, the distalportion of the ovovitelline ducts receives the shellglands openings, and the common glandular ovovitel-line duct is very short. Externally, the new species alsodiffers from G. chanca in that in this species the medialband extends from the anterior to the posterior tip, andin that it does not show the whitish cross-shaped designon the back. Furthermore, the body of G. chanca isbroader than G. valdiviana sp. n.

These shared features might be evidence of a close re-lationship of G. valdiviana sp. n. to G. chanca. Strik-ingly, a species belonging to another genus, Pasipha er-cilla (Froehlich E. M., 1978), also from Chile, presentssimilar features in the cutaneous and parenchymal mus-cular systems. As far as we know there are no other Geo-planinae species with parenchymal muscle fibers ar-ranged into bundles throughout the body. These featuresmight be an evidence of close phylogenetic relationshipfor the three Chilean species. In the Brazilian Geobiasubterranea (Schultze & M�ller, 1857) the greatly devel-oped cutaneous and parenchymal musculature are likelyanalogous since these specializations – ostensibly anadaptation to life underground (Froehlich C. G. 1955) –are restricted to only the anterior third of the body.

The Chilean land planarian fauna is still poorly studiedand appears to harbor a large morphological diversity.There is a need to study in detail the cutaneous and par-enchymal muscle systems in order to elucidate whetherthe peculiarities found in this new species support the hy-pothesis of a Chilean subclade within Geoplaninae.

Zoosyst. Evol. 87 (2) 2011, 327–334 333


Acknowledgements

We would like to express our gratitude to Franscico Javier C�dizLorca for collecting the specimen, to Dr. Birger Neuhaus, curatorof Vermes in the Museum f�r Naturkunde Berlin (MFN) and theMFN for the research facilities granted to JHG, to Jutta Zeller, forthe professional assistance with the histological procedures at theMFN, to Dr. Jason Dunlop for linguistic corrections, and the tothe two anonymous referees for their valuable comments and sug-gestions.

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a new land planarian species of geoplana (platyhelminthes,...

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